Term
| [CAM] What are the cellular requirements for multicellular organisms to survive? |
|
Definition
| 1. cells must be organized into distinct tissues 2. cells must adhere to other cells in the same tissue 3. different tissues need to be separated from one another |
|
|
Term
| [CAM] components of multicellular maintenance |
|
Definition
| ECM, cell adhesion molecules, cell junctions |
|
|
Term
| [CAM] what is the extracellular matrix? |
|
Definition
| network of proteins and carbohydrates that is secreted (contains collage, fibronectin...) |
|
|
Term
| [CAM] what is the basic definition of cell adhesion molecules? |
|
Definition
| they are proteins, on the surface of cells, that connect cells to other cells (Cadherins) or cells to the ECM (integrins) |
|
|
Term
| [CAM] What are some characteristics of CAM mediated cell adhesion? |
|
Definition
| low affinity, but very strong |
|
|
Term
| [CAM] Why are CAM mediated cell adhesions so strong? |
|
Definition
| 1. number of linkages 2. linkages with the cytoskeleton (actin, intermediate filaments) |
|
|
Term
| [CAM] What are the types of CAM's? |
|
Definition
| Cadherins, Immunoglobulins, Integrins, Selectins |
|
|
Term
| [CAM] What is the structure like of Cadherin? |
|
Definition
| single-pass integral membrane (NOT TM) that forms a homodimer |
|
|
Term
| [CAM] What is the function of Cadherin? |
|
Definition
| homophilic cell-cell interactions, form part of Adherens junctions and Desmosomes |
|
|
Term
| [CAM] Are Cadherins Ca++ dependent? |
|
Definition
|
|
Term
| [CAM] what is the basic structure of immunoglobulins? |
|
Definition
| single-pass integral membrane protein (NOT TM), does NOT dimerize, has Ig domain |
|
|
Term
| [CAM] What is the function of IgCAM's? |
|
Definition
| NCAM: homophilic cell-cell adhesion via sialic acid (the more sialylation, LESS adhesion) ICAM: heterophilic cell adhesion (ICAMS on endothelium bind integrins on rbc's) |
|
|
Term
| [CAM] Do IgCAM's depend on Ca++? |
|
Definition
| NO!!! IgCAM's are Ca++ independent |
|
|
Term
| [CAM] What is the basic structure of Integrins? |
|
Definition
| form heterodimers of alpha and beta subunits; alpha has 4 Ca++ binding sites, is bisected connected by a disulfide bond, is TM; beta has 1 Ca++ binding site, has Cys rich domain, is TM; head groups in extracellular domain bind ECM; C-termini (intracellular) bind actin |
|
|
Term
| [CAM] What ECM substrates do integrins bind to? |
|
Definition
| collagen, laminin, fibronectin, proteoglycans |
|
|
Term
| [CAM] Are integrins dependent on Ca++? |
|
Definition
| Yes, or on any divalent cation! (Ca++, Mg++...) |
|
|
Term
| [CAM] What is the basic structure of selectins? |
|
Definition
| composed of perlecan domain and lectin domain; lectin binds carbohydrates and glycoproteins |
|
|
Term
| [CAM] What is the function of the lectin domain in selectings? |
|
Definition
| binds carbohydrates and glycoproteins in ECM |
|
|
Term
| [CAM] What is the function of Selectins? |
|
Definition
| responsible for leukocyte-endothelium interaction and heterophilic cell cell interaction in general |
|
|
Term
| [CAM] Are Selectins Ca++ dependent? |
|
Definition
|
|
Term
| [CAM] Describe the model for Adherens formation via E-Cadherin adhesion |
|
Definition
| each cell contains monomers of Cadherins, which dimerize on the same cell... these dimers then polymerize even further with other dimers on the same cell; these polymers then polymerize with Cadherin polymers on the adjacent cell (all polymerization is Ca++ mediated... the higher Ca++ concentration, the more polymerization there is) |
|
|
Term
| [CAM] Experiment 1: Cadherins and Ca needed for Adherens formation |
|
Definition
| grew L-cells (mouse fibroblasts) which do not express Cadherin: -Cadherin gene +Ca... no Adherens; +Cadherin +Ca... Adherens formation; +Cadherin -Ca... no Adherens |
|
|
Term
| [CAM] Experiment 2: E-Cadherins mediate cell-cell adhesion according to type of Cadherin and amount of Cadherin secreted |
|
Definition
| Different types of Cadherins: use different fluorescent labels on different Cadherins (E vs. N) -- E aggregate to the center, N aggregate to the periphery; make cells expressing different amounts of Cadherin -- those expressing high Cadherin aggregate at center, those expressing low Cadherin aggregate to periphery [SORTING OUT] |
|
|
Term
| [CAM] What are the three major groups of ECM proteins? |
|
Definition
| Proteoglycans (perlecans -- selectins), collagen (sheet forming, fibular forming), Multiadhesive (laminin, fibronectin (binds integrins), nidogen/entactin) |
|
|
Term
| [CAM] Experiment 2: mixed green and red fluorescently labelled cells expressing proteoglycan and aggregation factor from the same sponge species or from a different species...what happens? |
|
Definition
| when both from same species: results in yellow, because the red and green mix. when from different species: results in green and red clusters, because aggregation is species specific |
|
|
Term
| [CAM] Experiment 4: control: grow salivary gland with fibronectin. experimental: grow salivary gland with anti-fibronectin. |
|
Definition
| control: salivary gland develops normally. Experimental: salivary gland develops abnormally. fibronectin is necessary for normal development. Also, anti-integrin results in same morphology as anti-fibronectin. |
|
|
Term
| [CAM] Experiment 5: Grew 1. WT mice 2. collagen 2 null mice 3. perlecan null mice |
|
Definition
| WT grew normally, collagen 2 and perlecan null mice resulted in dwarfism... very underdeveloped. ECM components are crucial for development! |
|
|
Term
| [CAM] Describe what happens when a signaling integrin is activated |
|
Definition
| when ligand binds to integrin, causes conformation change (bent over vs erect). integrins and RTK's coactivate each other, and together, activate SRC protein pathway and PKB pathway, which leads to cell proliferation, cell survival, cytoskeletal organization, cell migration, and gene transcription |
|
|
Term
| [CAM] What pathways does the signaling integrin activation lead to? |
|
Definition
| SRC protein pathway, PKB pathway |
|
|
Term
| [CAM] What processes are turned on when signaling integrins/RTK's are activated? |
|
Definition
| cell proliferation, cell migration, cell survival, gene transcription, cytoskeletal organization |
|
|
Term
| [CAM] Describe the mechanism for signaling integrin activation? |
|
Definition
| unbound state is a heterodimer and is bent over, which brings the two subunits close together. ligand binds and causes the heterodimer to become erect, allowing the two subunits to separate. This conformational change causes the interactions with adapter proteins to change, leading to downstream effects |
|
|
Term
| [CAM] What are the 4 types of epithelial cells that we discussed? |
|
Definition
| simple columnar, simple squamous, transitional, stratified squamous |
|
|
Term
| [CAM] where are simple columnar epithelial cells found? |
|
Definition
| cervical tract, digestive tract |
|
|
Term
| [CAM] where are simple squamous epithelial cells found? |
|
Definition
| blood vessels, body cavities |
|
|
Term
| [CAM] where are transitional epithelial cells found? |
|
Definition
| bladder, function in its contraction and expansion |
|
|
Term
| [CAM] Where are stratified squamous epithelial cells found? |
|
Definition
|
|
Term
| [CAM] What are the functions of Cell Junctions? |
|
Definition
| 1. bind cells together 2. form water and salt tight seal between cells 3. couples cells metabolically and electrically 4. anchor cells to ECM |
|
|
Term
| [CAM] What are the main types of cell junctions? |
|
Definition
| Anchoring Junctions, Tight junctions, Gap junctions |
|
|
Term
| [CAM] What are the types of anchoring junctions? |
|
Definition
| Adherens Junctions, Desmosomes, hemidesmosomes |
|
|
Term
| [CAM] For Aderens junctions, what kind of interaction is it? |
|
Definition
|
|
Term
| [CAM] For Adherens junctions, what is it formed by? |
|
Definition
| Cadherins: Cadherin termini bind to p-120 Catenin and petaCatenin, then betaCatenin binds to alphaCatenin, then alphaCatenin binds to F-actin ---> attaches to actin in the cytoskeletons of the two cells |
|
|
Term
| [CAM] In Adherens junctions, what function do alpha- and betaCatenins serve? |
|
Definition
| adapter molecules by joining Cadherins with F-actin |
|
|
Term
| [CAM] What does a loss of E-Cadherin lead to? |
|
Definition
| loss of Adherens junctions --> motile cells --> cancer |
|
|
Term
| [CAM] For Desmosomes, what kind of interaction is it? |
|
Definition
|
|
Term
| [CAM] For Adherens junctions, what is the function of these junctions? |
|
Definition
| shape, tension, signaling |
|
|
Term
| [CAM] For Desmosomes, what kind of function do they mediate? |
|
Definition
| strength, durability, signaling |
|
|
Term
| [CAM] For Desmosomes, what is it formed by? |
|
Definition
| Desmosomal Cadherins: desmoglein, desmocollin |
|
|
Term
| [CAM] What are the desmosomal Cadherins, and what do they do? |
|
Definition
| Desmoglein, Desmocollin; form Desmosomes |
|
|
Term
| [CAM] In Desmosomes, what do the desmosomal Cadherins do? |
|
Definition
| interact with with special cytosolic proteins (plakoglobin, desmoplakin, plakophilins) to form cytoplasmic plaque |
|
|
Term
| [CAM] What special cytosolic proteins are involved in desmosomes? (hint: form cytoplasmic plaque) |
|
Definition
| plakoglobin, desmoplakin, plabophilin |
|
|
Term
| [CAM] In desmosomes, what attaches to cytoplasmic plaques? |
|
Definition
| Intermediate filaments...refer to picture |
|
|
Term
| [CAM] In hemidesmosomes, what kind of interaction is it? |
|
Definition
|
|
Term
| [CAM] In hemidesmosomes, what is it made of? |
|
Definition
| integrins (alpha 6 beta 4) |
|
|
Term
| [CAM] in hemidesmosomes, what do the integrins attach to? |
|
Definition
| intermediate filaments of the cytoskeleton |
|
|
Term
| [CAM] in hemidesmosomes, what is the function? |
|
Definition
| shape, rigiditiy, signaling |
|
|
Term
| [CAM] in hemidesmosomes, name some subunit compositions of integrins and their associated ligands |
|
Definition
| alpha6beta4 ion epithelial cells; laminin. alpha11bbeta3 in serum proteins such as fibrinogen: RGD sequence. fibronectin: platelets |
|
|
Term
| [CAM] compare hemidesmosomes to focal adhesions |
|
Definition
| focal adhesions: non-epithelial cells, connect to actin and fibronectin. hemidesmosomes: epithelial cells, connect to intermediate filaments and laminin (ECM) |
|
|
Term
| [CAM] What kind of cells do hemidesmosomes deal with? |
|
Definition
|
|
Term
| [CAM] Tight Junctions, what is their function? |
|
Definition
| To form a water-tight and salt-tight seal, and thus divides the cell into 2 surfaces -- apical and basolateral -- each surface has its own set of proteins, lipids, functions --> essential for cell polarity |
|
|
Term
| [CAM] Where are tight junctions present? |
|
Definition
| epithelia in epidermis, intestines, pancreatic ducts |
|
|
Term
| [CAM] Experiment 6: fluorescently labeled apical lipids... what happens? |
|
Definition
| apical lipids remain apical... unless the tight junction is disrupted --> tight junctions block protein and lipid diffusion from one surface to the other |
|
|
Term
| [CAM] What are the molecules that make up tight junctions? |
|
Definition
| occludin, ZO proteins (anchor to actin), claudin (TM protein), JAM |
|
|
Term
| [CAM] In tight junctions, what do occludins do? |
|
Definition
| interact with actin via ZO adapter proteins |
|
|
Term
| [CAM] in tight junctions, what do JAM's do? |
|
Definition
| are junction adhesion molecules from the IgCAM family |
|
|
Term
| [CAM] Experiment 7: What was used to determine where the tight junction was? (stained black, but could not penetrate tight junction) |
|
Definition
| lanthanum hydroxyde, between cells |
|
|
Term
| [CAM] Tight Junctions, what kind of interaction? |
|
Definition
|
|
Term
| [CAM] Gap Junctions, what kind of interaction? |
|
Definition
|
|
Term
| [CAM] What are Gap junctions made of? |
|
Definition
| connexins, innexins, pannexins |
|
|
Term
| [CAM] Does the Gap junction connect to the cytoskeleton? |
|
Definition
| possibly, but would have to be indirectly, via adaptor proteins or other types of junctions |
|
|
Term
| [CAM] What is the funciton of gap junctions? |
|
Definition
| communication between cells, transport of small molecules from cell to cell, dispersion of signals (Ca, cAMP, ions...) |
|
|
Term
| [CAM] Gap junctions are basically hexagonal cylindrical pores that connect adjacent cells |
|
Definition
|
|
Term
| [CAM] What is the structure of gap junctions in vertebrates? |
|
Definition
| 12 homo- or heteroconnexins per channel, so 6 connexins per cell (invertebrates-innexins, pannexins - connexins and innexins together) |
|
|
Term
| [CAM] What is the size of the largest molecule that can pass through a gap junction? |
|
Definition
| 1.2kD (ions, vitamins, signaling molecules, small peptides) |
|
|
Term
| [CAM] How can the permeability of gap junctions be regulated? |
|
Definition
| pH, Ca concentration, phosphorylation of connexins/innexins |
|
|
Term
| [CAM] How can one regulate the permeability of gap junctions using Ca? |
|
Definition
| high intracellular Ca leads to channel closing; open channel by decreasing intracellular Ca (Ca diffusion important for coordinate contraction in heart) |
|
|
Term
| [CAM] Gap junction: hexameric ring 1 (cell 1) + hexameric ring 2 (cell 2). Each ring crosses membrane four times. |
|
Definition
|
|
Term
| [ECM] What are the major components of the ECM? |
|
Definition
| Basal Lamina, Collagens, Proteoglycans + GAG's, fibronectin, adhesive interactions in motile in normal cells by integrins |
|
|
Term
| [ECM][Basal Lamina] What is the Basal Lamina specialized for? |
|
Definition
| selective permeability, cell attachment (provides scaffold), migration) |
|
|
Term
| [ECM][Basal Lamina] Basal lamina is light or dense ECM? |
|
Definition
|
|
Term
| [ECM][Basal Lamina] What tissues does the basal lamina surround and connect? |
|
Definition
| epithelial cells (such as in the capillaries), muscle, fat, Schwann cells |
|
|
Term
| [ECM][Basal Lamina] Functions of the basal lamina? |
|
Definition
| separates endothelial cells and connective tissue; filters (in kidneys, intestine, capillaries), provides highways for cell migration, acts as a scaffold for cell regeneration |
|
|
Term
| [ECM][Basal Lamina] What are the basic components of the Basal Lamina? |
|
Definition
| Type 4 collagen(more flexible than fibrillar collagens, and assembles into sheet-like multilayered network) Perlecan(heparan sulfate proteoglycan) Laminin |
|
|
Term
| [ECM][Basal Lamina] Describe laminin's 3 chains |
|
Definition
| 3 polypeptides: beta binds lipids, alpha?, gamma binds lipids and collagens |
|
|
Term
| [ECM][Basal Lamina] laminin serves as what for cells? |
|
Definition
| laminin is what cells bind to in order to bind the basal lamina |
|
|
Term
| [ECM][Basal Lamina] What component of the basal lamina do cells bind to? |
|
Definition
|
|
Term
| [ECM][Basal Lamina] what molecules does the basal lamina laminin interact with? |
|
Definition
| Type 4 collagen (what it weaves in and out of), perlecans, integrins (form adhesive structures), entactin (a glycoprotein) |
|
|
Term
| [ECM][Basal Lamina] How are the three polypeptides in laminin linked? |
|
Definition
|
|
Term
| [ECM][Collagens] What is the basic structure of collagen? |
|
Definition
| right handed triple helix (of which the alpha chain is left handed); chains consist of Gly-Pro-X repeats. Gly in the center, Pro provides twisting ability |
|
|
Term
| [ECM][Collagens] What are the amino acid repeats in the alpha chain of collagens? |
|
Definition
| Gly-Pro-X gly in the center, pro provides twisting ability |
|
|
Term
| [ECM][Collagens] How are the different types of collagens distinguished from each other? |
|
Definition
| length of alpha chain, modifications to alpha chain, presence of segments that interrup the triple helix |
|
|
Term
| [ECM][Collagens] Describe Type 1 collagen |
|
Definition
| alpha length: 300nm; present in skin, tendons, bone, ligaments, dentin, interstitial tissues |
|
|
Term
| [ECM][Collagens] Describe Type 6 collagen |
|
Definition
| laterally associated with type 1, found in interstitial tissues, contains some globular domains |
|
|
Term
| [ECM][Collagens] Describe Type 9 collagen |
|
Definition
| laterally associated with type 2 collagen, has a globular N-terminus, has bound GAG, found in cartilage and vitreous humor |
|
|
Term
| [ECM][Collagens] Describe Type 4 Collagen |
|
Definition
| forms the network for ALL basal laminae |
|
|
Term
| [ECM][Collagens] What do mutations in Type 4 collagen cause? |
|
Definition
| pulmonary hemorrhage, renal failure |
|
|
Term
| [ECM][Collagens] Describe how Collagen 4 is assembled |
|
Definition
| a collagen monomer (the triple helix) has a head and a tail. If a dimer is formed, take two and put them tail to tail. If a tetramer is formed, take 4 and put them head to head -- to form the network for ECM |
|
|
Term
| [ECM][Collagens] Describe the Biosynthesis of Collagens 1,2,3 |
|
Definition
| ribosomes translate and translocate collagens into the ER. The peptides are then made into a triple helix by disulfide bonds, N glycosylation, and hydroxylation. Hsp47 interacts with the procollaten. Then it is transported to the Golgi, where multiple procollagens are laterally associated. They are then exoctytoses, and are cleaved to form collagens, which are assembled and crosslinked to form fibrillar collagen -- tendon and cartilage |
|
|
Term
| [ECM][Collagens] In the biosynthesis of collagen, what is the purpose of the propeptides? |
|
Definition
| They prevent the collagens from polymerizing too early. They are cleaved when the procollagens are exocytosed. |
|
|
Term
| [ECM][Collagens] Describe the organization of tendons |
|
Definition
| Fibrillar Collagen 1 associated with collagen 4 |
|
|
Term
| [ECM][Collagens] describe the structure of cartilage |
|
Definition
| fibrillar collagen 2 associated with collagen 6 and chondroitin sulfate |
|
|
Term
| [ECM][Proteoglycans+GAG] what are proteoglycans? |
|
Definition
| core protein with covalently attached GAG's |
|
|
Term
| [ECM][Proteoglycans+GAG] How are proteoglycans made? |
|
Definition
| the core protein is made in the ER, which is then transported to the Golgi, where it is GAG-glycosylated, sulfated at Ser residues (except hyaluronan of course) |
|
|
Term
| [ECM][Proteoglycans+GAG] What are the two main types of proteoglycans? |
|
Definition
| TM proteins (syndecans) and secreted PG's |
|
|
Term
| [ECM][Proteoglycans+GAG] What are the differences between proteoglycans and glycoproteins? |
|
Definition
| PG's are much longer and unbranched, and have a huge proportion of carbohydrates (up to 95% carbohydrate), can be huge (Aggrecan). GP on the other hand are mostly protein, shorter, branched... |
|
|
Term
| [ECM][Proteoglycans+GAG] Describe glycosaminoglycans GAG's |
|
Definition
| disaccharide polymers of GlcNac/GalNac and GlcUA/Gal that are very highly sulfated, so have a very negative charge, LARGE |
|
|
Term
| [ECM][Proteoglycans+GAG] What are the types of GAG's? |
|
Definition
| Hyaluronan, Chondroitin sulfate, heparan sulfate, keratan sulfate |
|
|
Term
| [ECM][Proteoglycans+GAG] What is Hyaluronan, and describe it |
|
Definition
| Hyaluronan is a type of GAG. It is O-glycosylated to a core protein's Ser residues. It resists compression, is NOT sulfated, is NOT protein-linked. In contrast to the other GAG's, it is formed on the cell surface, NOT the Golgi, binds a lot of water. Forms the core of cartilage. |
|
|
Term
| [ECM][Proteoglycans+GAG] What is the one GAG that is formed differently from all the others, and where does this take place? |
|
Definition
| Hyaluronan, it is formed on the cell surface instead of in the Golgi. |
|
|
Term
| [ECM][Proteoglycans+GAG] Describe chondroitin sulfate |
|
Definition
| It is formed in the Golgi, it is O-glycosylated, it is heavily sulfated, it is small (N<250) |
|
|
Term
| [ECM][Proteoglycans+GAG] Describe Heparan sulfate |
|
Definition
| heavily sulfated, N = 200, functions in anticlotting by activating antithrombin 3, secreted by mast cells, is O-glycosylated |
|
|
Term
| [ECM][Proteoglycans+GAG] Describe Keratan sulfate |
|
Definition
| heavily sulfated, very small N = 30-40, N-glycosylated instead of O-glycosylated like the rest of the GAG's, found in the cornea |
|
|
Term
| [ECM][Proteoglycans+GAG] Describe the structure of cartilage. |
|
Definition
| Hyaluronan forms the core. attached to it are long linking sugars, to which keratan sulfate, then chondroitin sulfate (longer) attach. look at the picture. Resists compression because of Hyaluronan |
|
|
Term
| [ECM][Proteoglycans+GAG] Describe the O-glycosylation of chondroitin sulfate |
|
Definition
| chondroitin sulfate repeats are attached to GlcUA, which is attached to the 3 linking sugars: Gly-X-Gly, which are attached to a Ser residue on the core protein |
|
|
Term
| [ECM][Proteoglycans+GAG] take a look at the O-glycosylation explanations in the lecture |
|
Definition
|
|
Term
| [ECM][Proteoglycans+GAG] What are the functions of PG's/GAG's? |
|
Definition
| 1. form a hydrated space, since they're so heavily hydrated, the GAG's form gels of varying pore size and charge that resist compression (cartilage...remember... Collagen does NOT resist compression). 2. modulate secreted proteins such as chemokines and proteases (fibroblast GF binds to heparan sulfate to cluster and activate receptors) |
|
|
Term
| [ECM][Fibronectin] What is fibronectin? |
|
Definition
| fibrillar, large, multi-adhesive ECM protein that binds many things |
|
|
Term
| [ECM][Fibronectin] What does fibronectin bind? |
|
Definition
| fibrous collagens, heparan sulfate (a GAG), integrins, itself |
|
|
Term
| [ECM][Fibronectin] What are the functions of fibronectin? |
|
Definition
| anchoring of cells to the matrix by binding integrins at focal adhesions and fibrillar adhesions, which are bound to the intermediate filaments inside the cell; regulation of cell shape and cytoskeleton; navigation of cells in development and wounds |
|
|
Term
| [ECM][Fibronectin] Fibronectin has the RGD sequence. What is this, what is it for, and what happens when it is not present? |
|
Definition
| RGD = arginine - glycine - aspartic acid, this is a binding site, it serves integrin binding, so if it is not present, cell adhesion to other cells reduced -- cancer |
|
|
Term
| [ECM][Fibronectin] Describe the structure of fibronectin |
|
Definition
| it is a dimer with repeating 1,2,3 modules that serve as binding sites for different things |
|
|
Term
| [ECM][Fibronectin] Describe the binding domains of fibronectin |
|
Definition
| 1: binds fibrin, heparan sulfate 2. binds collagen 3. functions in fibril assembly, fibrillogenesis RGD. binds integrins. |
|
|
Term
| [ECM][Fibronectin] Describe the mechanism for fibril assembly. |
|
Definition
| Fibronectin secretion doesn't automatically lead to fibril assembly. involves binding domain 3. fibrillogenesis only takes place on the surface of some cells. so, fibronectin regulates cell structure, and cells regulate fibronectin structure. |
|
|
Term
| [ECM][Adhesive Interactions] What kind of adhesive interactions do integrins partake in? |
|
Definition
| integrins cluster into adhesive structures in NON-epithelial cells and link ECM to microfilaments (adherens junctions) and intermediate filaments (hemidesmosomes). Remember how integrins are activated. |
|
|
Term
| [ECM][Adhesive Interactions] Describe the structure of the Dystrophin glycoprotein complex in skeletal muscle cells. |
|
Definition
| The DGC (dystrophin glycoprotein complex) comprises three subcomplexes: alpha beta dystroglycan, sarcogycan/sarcospan subcomplex of integral membrane proteins, and the cytosolic adapter subcomplex comprising dystrophin, other adapter proteins, and signaling molecules. |
|
|
Term
| [ECM][Adhesive Interactions] Describe the binding of the DGC in skeletal muscle cells. |
|
Definition
| beta-dystroglycan is O-linked to components of the basal lamina (laminin and perlecan) as well as cell surface proteins (neurexin on neurons). Dystrophin links betaDystroglycan to to the actin cytoskeleton. alpha-dystrobrevin links dystrophin to the sarcoglycan/sarcospan subcomplex. |
|
|
Term
| [ECM][Adhesive Interactions] Describe the function of the DGC on skeletal muscle cells |
|
Definition
| NO synthase produces NO when Ca levels are high during muscle contraction, NO is a signaling molecule that diffuses to promote smooth muscle relaxation, leading to a rise in the flow of blood supplying nutrients and oxygen to skeletal muscle. Involved with the GRB2 pathway. |
|
|
Term
| [ECM][Adhesive Interactions] Describe cell-cell adhesion in leukocyte extravasation |
|
Definition
| In the absence of inflammation/infection, leukocytes and endothelial cells are in resting state. inflammatory signals released that activate P-selectin exocytosis from epithelials. exposed selectins lead to loose binding of leukocytes with carbohydrate ligands on the leukocytes; also, epithelials release platelet activating factor and ICAM1. PAF induce changes in the leukocyte shape and activation of leukocyte integrins such as alphaLbeta2. activated integrins on leukocytes tightly bind with CAMs on the epithelium, so there is firm adhesion and movement into the underlying tissue. |
|
|
Term
| [ECM][Adhesive Interactions] What does ICAM stand for? |
|
Definition
| intercellular adhesion molecule |
|
|
Term
| [Cell Cycle] What does cell division function in? |
|
Definition
| reproduction, growth, repair |
|
|
Term
| [Cell Cycle] Describe staining methods for cell division |
|
Definition
| red, antibodies against tubulin. green, antibodies against CeBUB-1, a spindle checkpoint protein (located on chromosomes and kinetochore-attached spindle microtubules during metaphase). blue, DAPI for DNA. |
|
|
Term
| [Cell Cycle] What are the major steps of the mammalian cell cycle? |
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Definition
|
|
Term
| [Cell Cycle] What occurs during prophase? |
|
Definition
| centrosomes migrate to the poles, chromosomes begin to condense and so appear as long threads |
|
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Term
| [Cell Cycle] What happens during prometapahse? |
|
Definition
| the spindle forms, the nuclear envelope degrades, the chromosomes finish condensing, the chromosomes are held together at their centromeres by spindle microtubules |
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Term
| [Cell Cycle] What happens during metaphase? |
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Definition
| chromosomes align at the equatorial plane |
|
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Term
| [Cell Cycle] What happens during anaphase? |
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Definition
|
|
Term
| [Cell Cycle] What happens during telophase/cytokinesis? |
|
Definition
| the nuclear envelope reforms, chromosomes decondense, spindle disassembles |
|
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Term
| [Cell Cycle] What happens during cytosenesis? |
|
Definition
| the contractile ring leads the cytoplasm to divide |
|
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Term
| [Cell Cycle] What happens during G1 |
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Definition
| growth, centrosomes replicate during late G1 |
|
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Term
| [Cell Cycle] what happens during S phase? |
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Definition
|
|
Term
| [Cell Cycle] What happens during G2? |
|
Definition
| DNA exists as a chromatid (replicated), the centrosomes separate |
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Term
|
Definition
| perpetually staying in G1 |
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|
Term
| [Cell Cycle] What are the machines that drive cell division? |
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Definition
| centriole replication, mitotic spindle formation, chromosome movement, contractile ring |
|
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Term
| [Cell Cycle][Machines that drive cell division] Describe how centriole replication occurs. |
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Definition
| 1 centrosome = 2 centrioles. These two centrioles separate at the end of G1. During S, a new centriole grows next to each old centriole, resulting in a total of 4 centrioles. During G2, the new centrioles continue to grow, and the chromosomes mature. During prophase, the two centrosomes migrate to the two poles to form the mitotic spindle |
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Term
| [Cell Cycle][Machines that drive cell division] During what stage do the centrosomes migrate to the two poles? |
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Definition
|
|
Term
| [Cell Cycle][Machines that drive cell division] When does centriole replication occur? |
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Definition
|
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Term
| [Cell Cycle][Machines that drive cell division] What are the components of mitotic spindle formation? |
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Definition
| centrosomes, asters (astral microtubules... look like stars as they radiate outward from centrosomes), spindle body (kinetochore MT, polar MT=overlap MT), motor proteins |
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Term
| [Cell Cycle][Machines that drive cell division] During mitotic spindle formation, what are the three types of microtubules that function? |
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Definition
| astral microtubules, which extend from the spindle poles to the cell cortex, and orient the spindle with the axis of cell division. kinetochore MT's, which link the spindle poles to the kinetochores of the chromosomes; first find the chromosomes, then attaches them through the two kinetochores to both spindle poles then transports them to the poles during anaphase A. polar MTs extend from each spindle pole body toward the opposite one and interact in an antiparallel manner, initially push the duplicated centrosomes apart during prophase, then maintain the spindle's structure, then pushing the spindle poles apart in anaphase B |
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Term
| [Cell Cycle] What do the astral MT's do during mitotic spindle formation? |
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Definition
| astral microtubules, which extend from the spindle poles to the cell cortex, and orient the spindle with the axis of cell division. |
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Term
| [Cell Cycle] What do the kinetochore MT's do during mitotic spindle formation? |
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Definition
| kinetochore MT's, which link the spindle poles to the kinetochores of the chromosomes; first find the chromosomes, then attaches them through the two kinetochores to both spindle poles then transports them to the poles during anaphase A. So, responsible for chromosome movement. |
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Term
| [Cell Clycle] What do polar MT's do during mitotic spindle formation? |
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Definition
| Polar MTs extend from each spindle pole body toward the opposite one and interact in an antiparallel manner, initially push the duplicated centrosomes apart during prophase, then maintain the spindle's structure, then pushing the spindle poles apart in anaphase B. So, responsible for symmetry and shape of the spindle. |
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Term
| [Cell Cycle] What happens to motor proteins during mitosis? |
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Definition
| motor proteins become a lot more dynamic because of the loss of stabilizing MAP function. +end directed kinesin related motor proteins become enriched in the interdigitation region and start treadmilling by adding to +end and losing from -end. The kinesin istimulates MT disassembly. +end directed Dynein motor protein moves the chromosome toward the pole. |
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Term
| [Cell Cycle] What controls passage through the cell cycle? |
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Definition
| heterodimeric protein kinases that contain a regulatory subunit (cyclin) and a catalytic subunit (cyclin-dependent kinase, CDK). Regulate the activities of multiple proteins involved in DNA replication and mitosis by phosphorylating them at specific regulatory sites, activating some and inhibiting others. |
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Term
| [Cell Cycle][Regulation] Describe regulation during G1 |
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Definition
| early G1: no cyclin-CDK's are active. midG1: G1 cyclinCDKs activate transcription of genes required for DNA replication. SCF ubiquitin-protein ligase polyubiquitinylates inhibitors of Sphase cyclin CDK's, marking them for degradation, initiating S phase |
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Term
| [Cell Cycle][Regulation] Describe regulation of S phase |
|
Definition
| initiated by SCF ubiquitin protein ligase's polyubiquinylation of inhibitors of S-phase cyclinCDK's, marking the inhibitors for degradation... thus activating S phase cyclin CDK's. S phase cyclinCDK's activates DNA replication. Once replication is done, cell enters G2. |
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Term
| [Cell Cycle][Regulation] Describe regulation of G2 phase |
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Definition
| completion of DNA regulation during S phase commences G2. In late G2, mitotic cyclinCDK's activate early steps of mitosis (nuclear envelope breakdown, MT remodeling, chromosome condensation...) |
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Term
| [Cell Cycle][Regulation] Describe regulation of mitosis |
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Definition
| during late G2, mitotic cyclinCDK's become active, which activate early steps of mitosis: nuclear envelope breakdown, MT remodeling to form mitotic spindle, chromosome condensation during prophase, chromosome kinetochore attachment to MT's of the spindle in metaphase. MT motors and spindle shorteing pullthe daughter chromatids toward opposite poles, but CANNOT separate until anaphase-promoting complex (APC/C), a ubiquitin-protein ligase, polyubiquitinylates securin, marking it for degvradation, resulting in degrdation of protein complexes linking sister chromatids at the centromere, and the onset of anaphase. After chromosome movement to the poles, APC/C polybiquitinylates mitotic cyclins, causing their degradation. drop in mitotic cyclinCDK activity and the action of cdc14 phosphatase, causes chromosome decondensation, reassembly of nuclear membranes, MT remodeling into cytoskeleton, and cytokinesis. |
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|
Term
| [Cell Cycle][Regulation] What are the major groups of cell cycle regulators? |
|
Definition
| protein synthesis, protein degradation, kinases, phosphatases, regulatory proteins |
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|
Term
| [Cell Cycle][Experiment] Describe hybridoma, and discuss the conclusion drawn from it. What organism was this conducted in? |
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Definition
| a mitotic cell, with condensed DNA, was fused with a G1 cell, with noncondensed DNA. Upon fusion, the DNA from the G1 cell started to condense, though not to the extent of the mitotic DNA. This shows that there is a mitotic factor. Sea urchin egg/embryo |
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|
Term
| [Cell Cycle][Experiment] Describe the experiment of the use of budding yeast to isolate cdc28 |
|
Definition
| Mutate cells with a temperature sensitive mutation in a CDC gene (cell division cycle). These mutants are then transformed with a genomic library from wild type cells and plated at a nonpermissive temperature. Each transformed cell takes up a single plasmid containing one genomic DNA fragment. Most of these genes do not encode for the defective protein, and so the cells that take up these genes won't form a colony. the rare cell that takes up a plasmid containing the WT version of the mutant gene is complemented, which allows the cell to replicate and form a colony at the nonpermissive temperature. Since many of the proteins that regulate the cell cycle are highly conserved, yeast can be used to isolate human genes encoding cell cycle proteins. |
|
|
Term
| [Cell Cycle] mammalian cell cycle is similar to that of yeast fission |
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Definition
|
|
Term
| [Cell Cycle][Experiment] What happens when there is a recessive cdc mutation? |
|
Definition
| cells become too long... this is because cdc genes promote cell division, so without this function, cells keep growing without being able to divide |
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|
Term
| [Cell Cycle][Experiment] What happens with a dominant cdc mutation? |
|
Definition
| cells too small... since cdc promotes cell division, a dominant mutation leads to too much division, before the cells can grow to their proper size. |
|
|
Term
| [Cell Cycle][Experiment] What happens in wee1 mutants? |
|
Definition
| too short... since wee1 genes inhibit cell division.. their mutation leads to uncontrolled cell divison |
|
|
Term
| [Cell Cycle][Experiment] What are the advantages of using xenopus oocytes? |
|
Definition
| oocytes are large, and so are easy to manipulate. can easily tell which phase they're at (spindle is visible to the naked eye); can track MPF activity, can track cyclin B concentration, fertilized egg cycles rapidly and synchronously between S phase and mitosis (15 minutes) |
|
|
Term
| [Cell Cycle][Experiment] What does xenopus egg cytosol contain? |
|
Definition
| materials for DNA replication, chromosome assembly, nuclear envelope assembly, translation |
|
|
Term
| [Cell Cycle][Experiment] What happens if you put xenopus egg cytosol + sperm chromatin? |
|
Definition
| several rounds of mitosis |
|
|
Term
| [Cell Cycle][Experiment] Take an oocyte arrested in G2 and add progesterone...what happens? |
|
Definition
| progesterone stimulates the oocyte to undergo meiosis 1 and 2 to produce the first polar body and an egg. The egg is arrested in metaphase of meiosis 2. it can then be fertilized by sperm cells |
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|
Term
| [Cell Cycle][Experiment] How can you collect mitotic cells? |
|
Definition
| Stimulate oocytes to undergo meiosis with progesterone, then fertilize the egg in order to produce mitosis ready cells |
|
|
Term
| [Cell Cycle][Experiment] What happens if you transfer cytoplasm from a cell that is arrested in metaphase of meiosis 2 to a cell that is arrested in G2? |
|
Definition
| The injected cell undergoes meiosis 1 and is arrested in metaphase of meiosis 2, like the donor cell... this showed the existed of MPF, maturation promoting factor (also mitosis promoting factor) |
|
|
Term
| [Cell Cycle] What is MPF? |
|
Definition
| maturation promoting factor, key in triggering mitosis, is cyclic in that it rises and falls with cyclin concentration. At highest MPF peak, entry into mitosis occurs. |
|
|
Term
| [Cell Cycle][Experiment] Describe the cycling of MPF |
|
Definition
| low during G2... rises during meiosis one but then declines...rises again during meiosis II and remains high during metaphase of meiosis 2 arrest.. then declines upon fertilization... then rises again during first embyonic mitosis, decreases... then rises again for second mitosis... |
|
|
Term
| [Cell Cycle] When does entry into mitosis occur? |
|
Definition
| During the highest MPF peak. |
|
|
Term
| [Cell Cycle] What requires protein synthesis in order to form? |
|
Definition
| MPF... cannot induce meiosis and mitosis if don't have protein synth to synth MPF |
|
|
Term
| [Cell Cycle][Experiment] What did the western blot using 35S methionin labeling show minutes after postfertilization in early sea urchin embryos? |
|
Definition
| proteins B and C increased more and more. cyclin increased, but suddenly decreased 76 min after fertilization, then increased again at 86 min. peaked again at 106 min and decreased again later. |
|
|
Term
| [Cell Cycle][Experiment] What stain was used to detect cyclin, protein B, protein C? and in what organism? |
|
Definition
| 35S methionine, early sea urchin embryos |
|
|
Term
| [Cell Cycle][Experiment] What did the plot of intensity of cyclin band and %cells that had divided during previous 10 min interval vs. min postfertilization show? |
|
Definition
| Cyclin B peaks early in mitosis, falls abruply just before cell cleavage, then accumulates again during the next interphase to peak during early mitosis...falls abruptly before second cleavage |
|
|
Term
| [Cell Cycle] What happens to cyclin B levels during the cell cycle? |
|
Definition
| synthesized continuously during the cell cycle, peaks early in mitosis, then is abruptly destroyed following each anaphase... since its concentration peaks in mitosis, it functions as a mitotic cyclin |
|
|
Term
| [Cell Cycle][Experiment] What occured in the xenopus extracts after addition of sperm nuclei UNTREATED extract |
|
Definition
| MPF activity and cyclin B concentration increase together during early mitotis, then plummet together during late mitotic events... |
|
|
Term
| [Cell Cycle][Experiment] What ocurred in the xenopus extracts after the addition of sperm nuclei, RNAase treated extract |
|
Definition
| 0 levels of MPF activity and cyclin B ... so no mitosis |
|
|
Term
| [Cell Cycle][Experiment] What happens to MPF activity level and cyclin B concentration in an RNAase treated extract + WT cyclin B mRNA? |
|
Definition
| MPF acitivty and cyclin B concentration increase durng early mitosis and plummet during late mitosis, like normal... so cyclin B mRNA is rescued, and is necessary for mitosis |
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|
Term
| [Cell Cycle][Experiment] What happens to MPF activity and cyclin B concentration in RNase-treated extract + mRNA encoding nondegradable cyclin B? |
|
Definition
| MPF activity and cyclin B increases and remains at peak levels...arresting the cell in early mitosis, so that chromosomes remain condense, and the nuclear envelop remains disassembled. Lack the late mitosis events of chromos decondensation and nuclear envelop assembly |
|
|
Term
| [Cell Cycle] cyclin syntehsis is NECESSARY and SUFFICIENT for continued cell cycle |
|
Definition
|
|
Term
| [Cell Cycle] What is required to finish mitosis and continue cell cycle? |
|
Definition
|
|
Term
| [Cell Cycle] What is cyclin degradation necessary for? |
|
Definition
| finishing mitosis and continuation of the cell cycle |
|
|
Term
| [Cell Cycle] What is the CDK and the mitotic cyclin in S. Pombe? |
|
Definition
|
|
Term
| [Cell Cycle] What is the CDK and the important cyclins in S. Cerevisiae? |
|
Definition
| Cdc28, Cln3 (midG1) and Clb1 and Clb2(late mitosis) |
|
|
Term
| [Cell Cycle] Name the important components of vertebrate mitosis |
|
Definition
| late G1 and Sphase CDK: CDK2........mitotic CDK's: CDK1 and CDK2..........Sphase and mitotic cyclin: cyclin A...........mitotic cyclins: cyclin A and cyclin B |
|
|
Term
| [Cell Cycle] What is MPF? |
|
Definition
| mitotic cyclin (Cyclin B) + mitotic cdk (cdk1=cdc2) |
|
|
Term
| [Cell Cycle] What does MPF target? |
|
Definition
| histone H1 (condensation), nuclear lamin (disassembly), APC, itself, microtubules, GRASP65 |
|
|
Term
| [Cell Cycle] What does the MPF kinase portion phosphorylate? |
|
Definition
|
|
Term
| [Cell Cycle] What does cdc2 need to function? |
|
Definition
| cyclin... MPK = cdc2+cyclin |
|
|
Term
| [Cell Cycle] What are different phases promoted by? |
|
Definition
|
|
Term
| [Cell Cycle] Give the ways in which Cdk's are regulated |
|
Definition
| association/destruction of cyclins, the activation of phosphorylation of Cdk, inhibition of phosphorylation of Cdk, Cdk inhibitors (CKI)... however, remember that Cdk levels remain pretty constant throughout cell cycle |
|
|
Term
| [Cell Cycle] What mediates cyclin association/destruction of M-cyclin and when? |
|
Definition
| APC degrades M-cyclin in late mitosis |
|
|
Term
| [Cell Cycle] Describe the activating phosphorylation events on Cdk. |
|
Definition
| CAK phosphorylates cdc2-T161 to activate it |
|
|
Term
| [Cell Cycle] Describe the inhibitory phosphorylation events on Cdk. |
|
Definition
| Wee1 (a Tyr kinase) phosphorylates cdc2 on Y15 to deactivate it; cdc25 dephosphorylates Y15 on cdc2 to activate it again. |
|
|
Term
| [Cell Cycle] What does CAK do? |
|
Definition
| it phosphorylates cdc2-T161 to activate it (required) |
|
|
Term
| [Cell Cycle] What does APC do? |
|
Definition
| APC degrades Mcyclin during late mitosis |
|
|
Term
| [Cell Cycle] What does Wee1 do? |
|
Definition
| Wee1 phosphorylates cdc2-Y15 to inactivate it. |
|
|
Term
| [Cell Cycle] What does cdc25 do? |
|
Definition
| it desphosphoryaltes cdc2-Y15 (which was phosphorylated by Wee1) in order to reactivate cdc2 |
|
|
Term
| [Cell Cycle] What is a positive feedback loop control in the cell cycle? |
|
Definition
| cdc25 dephosphorylation of cdk1 |
|
|
Term
| [Cell Cycle] Describe the activation of MPF |
|
Definition
| inactive MPF --- phosphorylated at Y15 by Wee1 ---- inactive MPF ----- phosphorylated at T161 by CAK ------- inactive MPF -------- dephosphorylation at Y15 by cdc25 ------ACTIVE MPF (figure 20-14) |
|
|
Term
| [Cell Cycle] Describe how MPF is deactivated and during what stage does this occur? |
|
Definition
| Late Anaphase. There is Inactive APC/C and phosphorylated Cdh1. Cdc14 dephosphorylates Cdh1, so that Cdh1 can complex with APC/C ----APC/C-Cdh1 = Active APC/C. Active APC/C causes polyubiquination of Mcyclin so that it goes to proteasome to be degraded. G1 cyclin CDK then phosphorylates Cdh1, so that it decomplexes from APC/C, deactivating it. |
|
|
Term
| [Cell Cycle] When is APC/C active? |
|
Definition
| ONLY during late anaphase, to polyubiquitinate Mcyclin |
|
|
Term
| [Cell Cycle] What is the interaction between cdc25 and Wee1 with respect to MPF activity? |
|
Definition
| They have opposing effects. Deficit of cdc25 or excess of Wee1 --- increased G2 ---elongated cells. Deficit of Wee1 or excess of Cdc25 ---- decreased G2 ---- Small cells |
|
|
Term
| [Cell Cycle] What does nuclear lamin phosphorylation lead to? |
|
Definition
| nuclear lamina disassembly |
|
|
Term
| [Cell Cycle] Describe the mechanism for nuclear lamina disassembly |
|
Definition
| The nuclear lamina (made of intermediate filaments) connects the inner nuclear membrane and DNA. MPF phosphorylates lamin tetramers, initiating lamina disassembly. |
|
|
Term
| [Cell Cycle] Describe the ATP analog-dependent CDK mutant |
|
Definition
| Cdc28 has a ATPbinding site that cannot bind bulky ATP analogs with groups attached. In the mutant, F88 is changed to glycine, which lacks a large side chain, so now bulky ATP analogs can fit, and thus bind, activating the CDK |
|
|
Term
| [Cell Cycle] What does the nuclear lamina consist of? |
|
Definition
| lamin tetramers aka. two dimers. |
|
|
Term
| [Cell Cycle] What does MPK do to the nuclear lamina? |
|
Definition
| MPF phosphorylates lamins, leading to the lamina's disassembly into lamin dimers |
|
|
Term
| [Cell Cycle] Describe the state of the nuclear lamina throughout the cell cycle |
|
Definition
| During interphase, lamin dimers are in the form of tetramers, so the nuclear envelope is intact ---at the beginning of mitosis, MPF phosphorylates lamins, so nuclear lamina disassembles into dimers, so nuclear envelope is no longer held together (prophase) ----- lamins are dephosphorylated during early telophase, so that the nuclear lamina reforms, and the nuclear envelope starts to come together again ----- in late telophase, nuclear envelope fragments fuse |
|
|
Term
| [Cell Cycle][Experiment] Site Directed Mutagenesis (SDM) was used to prepare mutant human lamin A, whose serines were replaced with alanines, so that it was no longer phosphorylatable by MPF. WT and mutant human lamin A was trasnfected into hamster, and human is much more expressed than endogenous lamin. cells at various stages of the cell cycle were stained with fluorescently labeled monoclonal antibody specific for human lamin A with a fluorescent dye that binds DNA. What happened? |
|
Definition
| In WT transfected cells, lamin A formed a bright halo around DNA in interphase, then disintegrated during prophase, and was fuzzy during metaphase. The DNA was only inside nucleus during interphase, then globular during prophase, and some DNA left the nucleus. In mutant, lamin A remains a bright band throughout the cycle, and DNA becomes globular during prophase. |
|
|
Term
| [Cell Cycle][Experiment] Site Directed Mutagenesis (SDM) was used to prepare mutant human lamin A, whose serines were replaced with alanines, so that it was no longer phosphorylatable by MPF. WT and mutant human lamin A was trasnfected into hamster, and human is much more expressed than endogenous lamin. cells at various stages of the cell cycle were stained with fluorescently labeled monoclonal antibody specific for human lamin A with a fluorescent dye that binds DNA. What happened? |
|
Definition
| In WT transfected cells, lamin A formed a bright halo around DNA in interphase, then disintegrated during prophase, and was fuzzy during metaphase. The DNA was only inside nucleus during interphase, then globular during prophase, and some DNA left the nucleus. In mutant, lamin A remains a bright band throughout the cycle, and DNA becomes globular during prophase. |
|
|
Term
| [Cell Cycle] What does phosphorylation of the nuclear envelope lead to? |
|
Definition
| phosphorylation of nuclear pore proteins leads to their separation from the nuclear envelope, during prophase. phosphorylated integral membrane proteins decrease their affinity for chromatin and so further contribute to nuclear envelope disassembly. Lamin phosphorylation leads to their depolymerization, and thus dissolution of the nuclear lamina. MPF then phosphorylates chromatin proteins, which induces chromatin condensation and inhibits interactions between chromatin and the nuclear envelope. |
|
|
Term
| [Cell Cycle] What causes the nuclear envelope to reassemble? |
|
Definition
| In the absence of MPF because cyclin has been degraded, lamin phosphatase is unopposed, and so it dephosphorylates everything. extensions of the ER associated with the decondensing chromosomes, and then fuse together to form a double membrane around the chromosome. the nuclear pores form again |
|
|
Term
| [Cell Cycle] What happens to the Golgi Apparatus during mitosis |
|
Definition
| During prometaphase, MPF phosphorylates Golgi matrix proteins GRASP65/55, which causes the cisternae to unstack. unstacked Golgi cause the acceleration of vesicle budding. During Golgi disassembly, monoubiquitination occurs. Golgi vesicles are then equally distributed into the daughter cells during mitosis. during telophase, p97, which is ubiquitin dependent, and NSF, which is ubiquitin independent, cause the Golgi membranes to fuse to form a single cisterna. Dephosphorylation of GRASP65/55 causes restacking and trans oligomerization. |
|
|
Term
| [Cell Cycle] What does NSF do to the Golgi and when? |
|
Definition
| NSF, which is ubiquitin INdependent, causes the fusion of Golgi membranes to form a single cisternae. Works along side p97. |
|
|
Term
| [Cell Cycle] What does p97 do to the Golgi? |
|
Definition
| during Golgi disassembly, it is mono-ubiquitinated. during telophase, ubiquitin dependent p97, along with ub-independent NSF, cause the fusion of Golgi membrane into one single cisterna. |
|
|
Term
| [Cell Cycle] Sister Chromatids are linked together by Cohesin. How are the sister chromatids able to separate? |
|
Definition
| The sister chromatids are held together by two bands that form a lasso (Smc3 and Smc1) and are held together at their ends by kleisin. before anaphase, Securin binds Separase and inhibits it. After the kinetochores have attached to spindle MT's, and the spindle apparatus is assembled and oriented, cdc20 specific factor attaches to APC and directs it towards the securin-separase complex and polyubiquitinylates securin. Securin is degraded, so Separase is now free to cleave the kleisin that holds the two Smc's together, allowing sister chromatids to be pulled apart. |
|
|
Term
| [Cell Cycle] Name the substrates of APC/C |
|
Definition
| cyclin (cyclin degradation), securin (polyubiquitinylation too) |
|
|
Term
| [Cell Cycle] What is the term for the ring that forms around sister chromatids, and what are its components? |
|
Definition
| Cohesin: Smc3, Smc1, kleisin.... kleisin broken by separase... |
|
|
Term
| [Cell Cycle] What does PP2A do? |
|
Definition
| controls cohesin concentration. normally associates with centromeres. is a protein phosphatase. rapidly dephosphorylates cohesin complexes phosphorylated b in late prophase by kinases, but only occurs in the vicinity of the centromere, so that centromere-associated cohesins do not dissociate during late prophase like cohesin complexes in the chromosome arms, but rather continue to link at centromere. |
|
|
Term
| [Cell Cycle] How are cohesins regulated? |
|
Definition
| phosphorylation leads to their dissociation. dephosphorylation (by PP2A) leads to their association. |
|
|
Term
| [Cell Cycle] What are APC/C's main substrates, and what is it activated by for each? |
|
Definition
| polyubiquitinates Mcyclin after being activated by Cdh1(after it is dephosphorylated by cdc14 phosphatase during late Anaphase). polyubiquitinylates securin after being activated by cdc20 |
|
|
Term
| [Cell Cycle] How are cyclins destroyed? |
|
Definition
| APC/C activates their destruction via polyubiquitination of their destruction boxes |
|
|
Term
| [Cell Cycle] Describe the cyclin destruction box |
|
Definition
| cyclin D-box has 9aa in cyclin B, Lys is polyubiquitinated and thus targets the cyclin for destruction, during anaphase. |
|
|
Term
| [Cell Cycle] Why is the Dbox in cyclins necessary? |
|
Definition
| entry into mitosis requires high cyclin B levels; exit from mitosis requires cyclin B degradation |
|
|
Term
| [Cell Cycle] How is progression through anaphase ensured? |
|
Definition
| sequential degradation of securin (cohesins) and cyclin B. In late anaphase, cdc20 directs APC/C to polyubiquitinate securin for degradation. Then, Cdh1 directs APC/C to mitotic cyclin polyubiquitination. |
|
|
Term
| [Cell Cycle] What inhibits cdc20 from directing APC/C to polyubiquitnate securin? |
|
Definition
| inhibited until all chromosomes are attached and aligned. |
|
|
Term
| [Cell Cycle] What inhibits cdh1 activity in order to prevent premature Mcyclin degradation? |
|
Definition
| Cdh1 is inhibited until the sister chromatids are separated. |
|
|
Term
| [Cell Cycle] Name cell cycle checkpoints. |
|
Definition
| Cdc20 cannot act until all chromosomes are attached and aligned. Cdh1 cannot act until sister chromatids are separated. |
|
|
Term
| [Cell Cycle][Prograssion in Interphase] What model organism was used to identify S-phase promoting factors (SPF) during interphase? Name differences in budding yeast vs fission yeast vs mammalian |
|
Definition
| Yeast. Cdc28 is the ONLY cdk in buddying yeast, Cdc2 in fission yeast... cdk1 in mammalian for mitotic prograssion.... but the mammalian Sphase cdk is cdk2 |
|
|
Term
| [Cell Cycle][Progression in Interphase] What promotes synthesis phase? |
|
Definition
|
|
Term
| [Cell Cycle][Progression in Interphase] In yeast, what comprises SPF? |
|
Definition
|
|
Term
| [Cell Cycle][Progression in Interphase] What happens if SPF is somehow nonfunctional? |
|
Definition
|
|
Term
| [Cell Cycle][Progression in Interphase] Experiment: What happened when wild type cells were subjected to 25C and 36C? |
|
Definition
| WT CDK able to bind G1cyclin to form SPF, and so in both cases, colonies form. |
|
|
Term
| [Cell Cycle][Progression in Interphase] Experiment: What happened when cdc28ts (temperature sensitive) cells were subjected to 25C and 36C? |
|
Definition
| mutated CDK had low affinity for G1cyclin; SPF formed in 25C conditions, so colonies formed; in 36C, could not form SPF complex, and so colonies could not form, were arrested in G1 |
|
|
Term
| [Cell Cycle][Progression in Interphase] Experiment: what happens when cdc28ts cells transformed with high-copy G1cyclin plasmids (CLN1 or CLN2) are subjected to 25C and 36C? |
|
Definition
| highcopy G1cyclin leads to a lot of G1cyclin production, which dramatically shifts equilibrium, so that colonies form under both temperatures despite low affinity |
|
|
Term
| [Cell Cycle][Progression in Interphase] Experiment: what is CLN1 or 2? |
|
Definition
| plasmid transformed into cells that causes high levels of G1cyclin... this was able to rescue the low affinity that resulted in 36C from cdc28 temperature sensitive mutations |
|
|
Term
| [Cell Cycle][Progression in Interphase] What regulates G1cyclin? |
|
Definition
| SCF, a E3 ubiquitin ligase, (Skp1/Cul1/Fbox protein), mediates the breakdown of Sic1, a G1cyclin inhibitor |
|
|
Term
| [Cell Cycle][Progression in Interphase] Describe the 4th mechanism for regulating cdk activity, whereby DNA replication is allowed to take place |
|
Definition
| Sphase cyclin is bound to CDK, but is also bound by its inhibitor Sic1, during midlateG1. High levels of G1cyclin-CDK phosphorylate Sic1. SCF, a UB ligase, polyUB's the phosphorylated Sic1, so that it is degraded. Sic1 unbound Scycling-CDK can now initiate transition to S phase, leading to DNA replication |
|
|
Term
| [Cell Cycle][Progression in Interphase] Give some mammalian analogs of Sic1 |
|
Definition
|
|
Term
| [Cell Cycle][Progression in Interphase] Describe the assembly and regulation of prereplication complexes during G2 and M |
|
Definition
| During G2 and M phases, dsDNA is bound by an ORC, and replication is inhibited by Btype cyclin CDK (which is low telophase-G1, high in S, G2, and M through anaphase. Btype Cyclin CDK serves to restrict DNA replication to ONCE per cycle |
|
|
Term
| [Cell Cycle][Progression in Interphase] Describe the assembly and regulation of prereplication complexes during G1 phase |
|
Definition
| Initiation factors MCM(DNA helicase), Cdc6(DNA helicase loading factor), and Cdt1 increase in concentration and bind to an ORC, forming the prereplication complex. cdc6 prevents MCM from moving |
|
|
Term
| [Cell Cycle][Progression of Interphase] Describe what happens during S |
|
Definition
| At the beginning of S phase, Sic1 is degraded (remember how?) and so Sphase cyclin CDK is uninhibited, and it phosphorylates cdc6, cdt1, and MCM... phosphorylated cdc6 is DEGRADED, and so now MCM can move ... this opens up the replication fork, separating the two DNA strands... RPA then binds to the ssDNA and DNA polymerase replicates the DNA |
|
|
Term
| [Cell Cycle][Progression of Interphase] How is cdc6 regulated, and what does it regulate? |
|
Definition
| cdc6 is inhibited by phosphorylation, brought on by Scyclin after Sic1 degradation. active cdc6 acts to prevent MCM from moving down and opening the replication fork once the prereplication complex has been formed.Cdc6 is inhibited until M-cdk is inactivated at the end of mitosis |
|
|
Term
| [Cell Cycle][Progression of Interphase] Experiment: Describe the procedure for the microinjection of cyclin D antibodies |
|
Definition
| Used G0 cells and injected growth factors to make them bigger. Then divide them into two groups. Into group 1, inject control antibody, and into group 2, inject anti-cyclin D antibody. Add BrdU to both (synthetic nucleoside that acts as an analog of Thymidine, and so is incorporated into replicating DNA). Then, incorporation of BrdU is determined. Group 1 results in BrdU positive cells, meaning that these cells underwent DNA replication. Group 2 results in BrdU negative cells, so that cyclin D is bound to antibodies, leading the cell to be unable to cause DNA replication. |
|
|
Term
| [Cell Cycle][Progression of Interphase] What do mitogens do in order to induce cell division? |
|
Definition
| expression of E2F (transcription factor) and G1 cyclin |
|
|
Term
| [Cell Cycle][Progression of Interphase] What keeps E2F inactive? What does this cause? |
|
Definition
| Retinoblastoma. inhibition of cell division |
|
|
Term
| [Cell Cycle][Progression of Interphase] What does Rb do? |
|
Definition
| inhibits E2F, thereby preventing cell division. |
|
|
Term
| [Cell Cycle][Progression of Interphase] Describe how Rb is regulated, and what this causes |
|
Definition
| Rb is inactivated when it is phosphorylated by G1-CDK. This releases E2F, activating it. |
|
|
Term
| [Cell Cycle][Progression of Interphase] Describe the positive feedback loop associated with E2F. |
|
Definition
| G1-CDK phosphorylates Rb, thus liberating E2F and activating it. E2F acts to stimulate cell division. It induces expression of even more G1-CDK and itself as well as Sphase genes. G1-CDK further phosphorylates Rb. |
|
|
Term
| [Cell Cycle][Progression of Interphase] Give an example of a tumor suppressor gene. |
|
Definition
| Rb, because it inhibits E2F, thus inhibiting Sphase gene expression and cell divison. |
|
|
Term
| [Cell Cycle][Progression of Interphase] In the Rb pathway, what induces G1-CDK? |
|
Definition
|
|
Term
| [Cell Cycle][Progression of Interphase] What are the three functions of E2F |
|
Definition
| 1. induces DNA replication, thus cell division 2. induces more production of G2-CDk, which is what activated E2F 3. induces production of more E2F |
|
|
Term
| [Cell Cycle][Checkpoints] Describe where the first checkpoint takes place |
|
Definition
|
|
Term
| [Cell Cycle][Checkpoints] Describe the purpose of the first checkpoint |
|
Definition
| Ensures that all DNA is replicated completely, ensures that environmental conditions are favorable |
|
|
Term
| [Cell Cycle][Checkpoints] What is the sensor for the first checkpoint |
|
Definition
| detection of replication forks by ATR |
|
|
Term
| [Cell Cycle][Checkpoints] How does the first checkpoint act? |
|
Definition
| it inhibits Cdc25C to prevent McyclinCDK activation, thus blocking early mitosis. Carried out by MPF |
|
|
Term
| [Cell Cycle][Checkpoints] What does the second checkpoint ensure? |
|
Definition
| Spindle Assembly Checkpoint: prevents Anaphase unless all kinetochores are properly associated with spindle MT's |
|
|
Term
| [Cell Cycle][Checkpoints] What are the two main mechanisms by which the second checkpoint is mediated? |
|
Definition
| kinetochore mediated, by cdc20; MTOC mediated, by Mad2 |
|
|
Term
| [Cell Cycle][Checkpoints] In the second checkpoint, what does cdc20 do? |
|
Definition
| kinetochore mediated: senses when all chromosomes have been attached and proper alignment has occured |
|
|
Term
| [Cell Cycle][Checkpoints] How does the second checkpoint work? Give the full mechanism |
|
Definition
| In order to enter into Anaphase, APC/C-Cdc20 mediated polyubiquitination of Securin is required in order to activate Separate. Mad2 associates with kinetochores that are NOT attached to MT's. nonMT bound - Kinetochore-bound Mad2 is converted to closed Mad2 conformation. This intial closed Mad2 causes many more Mad2 molecules in the cytosol to convert to the closed conformation. Closed-Mad2 binds and inactivates cdc20. Once all kinetochores are attached to MT's, Mad1/2 complexes detach and bid p31 instead. p31 also binds closed Mad2's, freeing cdc20. cdc20 directs securin degradation, initiating anaphase. |
|
|
Term
| [Cell Cycle][Checkpoints] Describe the function of p31. |
|
Definition
| When all kinetochores are bound to MT's, Mad1/2 decomplex from kinetochores and bind to p31. p31 also binds closed Mad2's, thus freeing cdc20 so that anaphase can occur. part of second checkpoint (mitotic spindle assembly) |
|
|
Term
| [Cell Cycle][Checkpoints] Where does the third checkpoint take place |
|
Definition
|
|
Term
| [Cell Cycle][Checkpoints] What does the third checkpoint ensure? |
|
Definition
| ensures that all chromosomes are properly segregated into the daughter cells before telophase and cytokinesis can take place |
|
|
Term
| [Cell Cycle][Checkpoints] As part of the third checkpoint, what is required to exit mitosis? |
|
Definition
|
|
Term
| [Cell Cycle][Checkpoints] where is cdc14 phosphatase localized? |
|
Definition
|
|
Term
| [Cell Cycle][Checkpoints] What does cdc14 do? |
|
Definition
| It promotes exit from mitosis, and thus is sequestered in the nucleolus during interphase and early mitosis. |
|
|
Term
| [Cell Cycle][Checkpoints] Describe the mechanism of the third checkpoint |
|
Definition
| Tem1, a GTPase switch, becomes associated with the spindle pole body SPB during anaphase. SPB=centrosome. At the SPB, Tem1 is in inactive GDP bound state, maintained so by GAP. The GEF that activates it is located in the cortex of the bud, and so is absent from the mother cell. Kin4, a kinase, is localized to the mother cell and is absent from the bud. When the MT elongation has correctly placed the daughter chromosomes into the bud, Tem1 comes into contact with GEF, so TEM1-GAP becomes phosphorylated, and thus inhibited. Tem1 is thus converted to its active GTPbound state, and it phosphorylates the nuclear anchor that binds/inhibits cdc14, releasing cdc14 phosphatase into the cytoplasm in both the mother and daughter cell. Active Cdc14 induces telophase and cytokinesis. |
|
|
Term
| [Cell Cycle][Checkpoints] What pathway does Tem1 activate? |
|
Definition
|
|
Term
| [Cell Cycle][Checkpoints] What does the fourth checkpoint do? |
|
Definition
| If there is DNA damage, it prevents progression through the cell cycle until the damage is repaired. |
|
|
Term
| [Cell Cycle][Checkpoints] What does arrest in G1 and S prevent? |
|
Definition
|
|
Term
| [Cell Cycle][Checkpoints] What does arrest in G2 prevent? |
|
Definition
| it allows DNA double stranded breaks to be repaired before mitosis |
|
|
Term
| [Cell Cycle][Checkpoints] What mediates the fourth checkpoint? |
|
Definition
|
|
Term
| [Cell Cycle][Checkpoints] What is p53? |
|
Definition
| it is a tumor suppressor, and is a transcription factor that activates the transcription of p21CIP when stabilized by DNA damage. p53 is unstable/inactive when DNA is NOT damaged. If DNA is damaged, p53 is stabilized/activated. |
|
|
Term
| [Cell Cycle][Checkpoints] What negatively regulates p53? |
|
Definition
| mdm2, a ubiquitin e3 ligase |
|
|
Term
| [Cell Cycle][Checkpoints] describe the mechanism of the fourth checkpoint |
|
Definition
| normal cell: healthy DNA, p53 is unstable/inactive, is ubiquitinated by Mdm2 cell continues with cell cycle. Damaged DNA: p53 is stabilized, activates the transcirption of p21CIP, a CDK inhibitor, thus arresting the cell in G1 and G2 |
|
|
Term
| [Cell Cycle][Checkpoints] What happens if there's extreme DNA damage? |
|
Definition
| p53 triggers cell apoptosis |
|
|
Term
| [Cell Cycle][Checkpoints] What is p21CIP? |
|
Definition
| inhibitor of cdk, causes cell arrest in G1 and G2, activated by p53 in the presence of damaged DNA |
|
|
Term
| [Apoptosis] What are the two types of cell death? |
|
Definition
|
|
Term
| [Apoptosis] What is apoptosis? |
|
Definition
| programmed cell death. genetically controlled. does not cause inflammation. cell itself activates death program |
|
|
Term
| [Apoptosis] What is necrosis? |
|
Definition
| accidental cell death. often causes inflammation. due to injury, radiation, chemicals, lack of nutrients... typically swell up and burst |
|
|
Term
| [Apoptosis] What is apoptosis good for? |
|
Definition
| removes unwanted cells or structures, it is for the good of the organism |
|
|
Term
| [Apoptosis] What are the four scenarios during which apoptosis takes place? |
|
Definition
| development, immune system, disease, ER stress caused by unfolded protein |
|
|
Term
| [Apoptosis] What has been the important organism for studying apoptosis during development? |
|
Definition
| C elegans, because it makes 1090 cells, then 131 undergo apoptosis |
|
|
Term
| [Apoptosis] Examples of Developmental Apoptosis? |
|
Definition
| C elegans loses cells, apoptosis in order to sculpt the limbs of developing mice (cell death markers are active in the space between forming digits), deleting unwanted structures such as the tadpole tail, removal of most neurons in order to have proper number of neurons, elimination of injured cells |
|
|
Term
| [Apoptosis][Experiment] Using chick embryo, emputate one of two developing limb buds. What happens to motor neuron generation? What happens to motor neuron survival? |
|
Definition
| generation, 100%, like normal. survival: 10%, lower than the normal limb's 50%. |
|
|
Term
| [Apoptosis]Experiment: Using chick embryo transplant limb bud from another embryo to this one, so that it has an extra bud on one side. WHat happens? |
|
Definition
| generation:100%, like normal. survival: 75%, higher than the normal of 50% |
|
|
Term
| [Apoptosis][Experiment] What was the conclusion drawn from the chick embryo amputation and transplantation study? |
|
Definition
| The size of the muscle neurons target determines the survival of the motor neurons |
|
|
Term
| [Apoptosis][Experiment] What was the result of NGF-/- or TrkA-/- mutations? |
|
Definition
| lack of NGF and TrkA, so loss of pain sensing neurons |
|
|
Term
| [Apoptosis][Apoptosis] What was the result of NT-3-/- or TrkC-/- mutations? |
|
Definition
| lack of neurotrophin 3 and TrkC, loss of balancing/proprioceptive neurons |
|
|
Term
| [Apoptosis][Experiment] What did the experiment of mutations affecting trophic/growth factors show? |
|
Definition
| Growth factors are important for cell survival |
|
|
Term
| [Apoptosis] Examples of immune system apoptosis? |
|
Definition
| removal of lymphocytes after the immune system has been challenged. Killing virus-infected cells. |
|
|
Term
| [Apoptosis] Examples of apoptosis in human disease? |
|
Definition
| uncontrolled cell growth triggers apoptosis...if not... cancer; neurons undergo apoptosis after stroke; neurodegenerative diseases such as retinitis pigmentosa (rhodopsin mutation that leads to photoreceptor apoptosis) |
|
|
Term
| [Apoptosis] ER stress because of an unfolded protein can trigger apoptosis |
|
Definition
|
|
Term
| [Apoptosis] What are the main features of apoptosis? |
|
Definition
| dense chromosome condensation along the periphery of the nucleus, cell shrinkage, formation of apoptotic bodies which are phagocytosed by magrophages, membrane blebbing |
|
|
Term
| [Apoptosis] What are apoptotic bodies? |
|
Definition
| fragmentation of the nucleus and cytoplasm, so that chromosomal DNA is hydrolyzed... these are consumed by macrophages |
|
|
Term
| [Apoptosis] What is pyknosis? |
|
Definition
| irreversible condensation of chromatin in the nucleus, resulting in large visible black circles under the microscope |
|
|
Term
| [Apoptosis] What is the purpose of apoptotic bodies? |
|
Definition
| The breakage of a cell without the spreading of harmful substances |
|
|
Term
| [Apoptosis] Experiment: What does Hoechst staining show? |
|
Definition
| It stains DNA, and so the stain is really bright and globular during apoptosis because of DNA condensation |
|
|
Term
| [Apoptosis] What is the basic chain of events during apotptosis? |
|
Definition
| Cell is signalled to die ---> caspase activation ---> cellular components are digested ---> cellular components are packed into apoptotic bodies ---> phagocytosis of apoptotic bodies |
|
|
Term
| [Apoptosis] How is caspase activated? |
|
Definition
| caspase is made as a proenzyme, and must be cleaved to be activated. Other caspases activate caspases by cleaving them at aspartic acids to generate subunits which form the mature caspase tetramer. There are initiator caspases (8and9) that activate caspases, and there are effector caspases (3) which cleave substrates that result in apoptosis. |
|
|
Term
| [Apoptosis] What do caspases do? |
|
Definition
| They are proteases, they cleave after aspartic acid residues (D), but must have Cys containing nucleophilic group in the active site, its signature motif |
|
|
Term
| [Apoptosis] Where did the name Caspase come from? |
|
Definition
| Cysteine-dependent ASPartate directed proteASES |
|
|
Term
| [Apoptosis] Caspases are regulated by Caspase inhibitors (IAP) |
|
Definition
|
|
Term
|
Definition
|
|
Term
| [Apoptosis] What is the sequence Caspases look for in order to cleave? |
|
Definition
| X-Glutamic Acid-X-Aspartic Acid |
|
|
Term
| [Apoptosis] Experiment: in C elegans, what do mutations in the apaf-1 homolog (CED4) and caspase (CED3) do in otherwise WT animals? |
|
Definition
|
|
Term
| [Apoptosis] If caspase3 is activated, apoptosis occurs. What are caspase3's substrates? |
|
Definition
| nuclear lamins --> loss of nucleus' integrity, inhibits caspase-activated Dnase/DNA fragmentation factor 45 --> factor 45 cuts up DNA, mitochondrial proteins --> no more electron transport, cytoskeletal and signaling proteins (actin, PKC..), GRASP65 and SNAREs --> fragmentation of intracellular membranes (Golgi, vesicles) |
|
|
Term
| [Apoptosis] What does factor 45 do? |
|
Definition
| cuts DNA into fragments, activated by caspase 3 |
|
|
Term
| [Apoptosis] What are the two death pathways? |
|
Definition
| Extrinsic and intrinsic, they intersect that the effector caspases |
|
|
Term
| [Apoptosis] What does extrinsic death pathway mean? |
|
Definition
| That cell's apoptosis is activated from outside the cell |
|
|
Term
| [Apoptosis] What is another name for the extrinsic death pathway? |
|
Definition
|
|
Term
| [Apoptosis] What is the mechanism for extrinsic apoptosis? |
|
Definition
| Killer lymphocytes have Fas ligands (tumor necrosis factor), that binds to a Fas receptor on the target cell. Three Fas bound Fas receptors come together to form a trimer. This Fas-receptor trimer binds procaspase8 via an adaptor protein FasL. Fas-Fasl-procaspase8 complex = DISC (death initiating signaling complex. This complexation activates caspase 8, which activcates caspase 3. |
|
|
Term
| [Apoptosis] What two purposes is Fas mediated apoptosis important for? |
|
Definition
| cytotoxic immune responses and elimination of self-reactive wbc's. Loss of Fas signalling leads to lymphoproliferative and autoimmune diseases |
|
|
Term
| [Apoptosis] What does loss of death ligand receptors (Fas receptors) lead to? |
|
Definition
|
|
Term
| [Apoptosis] What is required for cell survival? |
|
Definition
| The PKB pathway (PI4P is converted by pi3kinase to pi3,4 bisphosphate... which recruits and partially activates PKB.. then PDK's fully ativate PKB |
|
|
Term
| [Apoptosis] Where is cyt c? |
|
Definition
| it is located between the inner and outer membranes of the mitochondria...so when mitochondria are injured..they release cyt c |
|
|
Term
| [Apoptosis] What is the basic pathway of intrinsic death? |
|
Definition
| Injured mitochondria release cyt c from between their inner and outer membranes... free cyt c binds to adaptor protein Apaf-1.... cyt c-Apaf1 bind and activate procaspase 9, leading to the formation of the apoptosome (cyt c-Apaf1-procaspase 9).. apoptosome assembly is essential for the activation of procaspase 9 and the initiation of the intrinsic apoptotic pathway... now active caspase 9 activates a downstream effector caspase that triggers apoptosis |
|
|
Term
| [Apoptosis] What is Apaf1? |
|
Definition
| adaptor protein that cyt c binds to in order to bind procaspase 9 and thus form the apoptosome |
|
|
Term
| [Apoptosis] What is a suppressor of the intrinsic apoptotic pathway? |
|
Definition
|
|
Term
| [Apoptosis] What is Bcl-2? |
|
Definition
| It is an intrinsic apoptosis pathway suppressor... it is a TM protein found in the ER and mitochondria's outer membrane |
|
|
Term
|
Definition
| It is a pro-apoptosis regulator, and is found on the outer membrane of the mitochondria |
|
|
Term
| [Apoptosis] What does overexpression of Bcl2 cause? |
|
Definition
| enhanced cell survival which can lead to cancer |
|
|
Term
| [Apoptosis] What happens in Bcl2 (CED9) mutants? |
|
Definition
|
|
Term
| [Apoptosis] In CED9 mutants, what happens when transformed with mammalian Bcl2? |
|
Definition
| CED's antiapoptotic activity is rescued... so no apoptosis |
|
|
Term
| [Apoptosis] Bcl family members (Bcl2, CED9...) can form heterodimers |
|
Definition
|
|
Term
| [Apoptosis] How is Bcl thought to act to prevent apoptosis? |
|
Definition
| Bcl has been found to associate with PTP's, permeability transition pores (nonselective ion channels). opening of these channels leads to apoptosis. Bcl2, along with other antiapoptotic factors, closes these channels. Or, Bcl2 and others might just negatively regulate proapoptotic factors like Bax |
|
|
Term
| [Apoptosis] What does opening of PTP's cause? |
|
Definition
|
|
Term
| [Apoptosis] Bcl2 family members can be suppressors of apoptosis OR propapoptotic proteins. Give examples for both. |
|
Definition
| Bad: forms inactive heterodimers with Bcl2, thus causing apoptosis (Bcl2 Associated Death promoter). Bax: Forms dimer on outer membrane to form an ion channel, and can form larger pores as well to promote mitochondrial swelling |
|
|
Term
| [Apoptosis] What does a lack of a survival signal lead to? |
|
Definition
| interaction of Bad with Bcl2, leading to Bcl2 inactivation, and thus apoptosis because Bax is allowed to remain open and thus causes swelling |
|
|
Term
| [Apoptosis] What does a survival signal do? |
|
Definition
| survival receptor activates PI3 kinase, posphorylating PI to PIP2. PIP2 activates AKT (PKB). PKB=AKT phosphorylates the pro-apoptotic BAD, thus inactivating it, so that Bcl2 can do its magic and close PTP's. Phosphorylated is bound to 14-3-3 protein, thus keeping it away from the mitochondria. |
|
|
Term
| [Apoptosis] Discuss the apoptotic pathway across evolution. |
|
Definition
| It is evolutionarily conserved... but in C elegans, cyt c is not required though it is still positively and negatively regulated |
|
|
Term
| [Apoptosis] Describe how ER mediated apoptosis works |
|
Definition
| It is also through cyt c release. caused by ER stress. Transcription of Ire1 regulates this, as well as the balance between Bcl2 and Bax. Mitochondria releases caspase12 |
|
|
Term
| [Apoptosis] What causes ER stress which then induces apoptosis? |
|
Definition
| Ca++ imbalance, misfolded protein |
|
|
Term
| [Apoptosis] What are the three factors governing cell division, growth and apoptosis? |
|
Definition
| mitogens = stimulate cell division by releasing cell cycle blockers such as PDGF and EGF, erythropoeitin); Growth factors = stimulate cell growth by promoting protein synthesis and inhibiting degradation; survival factors: promote cell survival by inhibiting apoptosis (most growth factors can do all these things... PKB pathway important for survival! most mitogens can activate Ras and MAP kinase |
|
|
Term
| [Apoptosis] Compare the formation of the Death Initiation Signaling Complex of the extrinsic pathway to the formation of the apoptosome of the intrinsic pathway. What signal initiates the formation of each complex? Where are the complexes formed in the cell? How are the caspases activated? |
|
Definition
|
|
Term
| [Apoptosis] An example of trophic factor, and what it does? |
|
Definition
| NGF, member of neurotrophin family. neurotrophins bind receptor tyrosine kinases. |
|
|
Term
| [Apoptosis] [Experiment] What happens as a result of knowckouts of neurotrophins? |
|
Definition
| lack of specific types of neurons |
|
|
Term
| [Apoptosis] How does the MAPK pathway work? |
|
Definition
| mitogens activate MAPK pathway --- MAPK promotes transcription of c-fos and myc --- more cell division (cancer). myc increases txn of cyclin D, SCF ubiquitin ligase, both of which increase G1/S cdk, which phosphorylate the Rb protein. Myc also increases synthesis of E2F, causing entry into S phase. |
|
|
Term
| [Apoptosis] What does excessive activation of mitogen pathways lead to? How is this signalled? |
|
Definition
| cell cycle arrest, apoptosis, cancer... abnormally high levels of myc activate p19ARF, which binds mdm2 and so activates p53, p53 then causes cycle arrest/apoptosis |
|
|
Term
| [Apoptosis] Growth factors can promote cell growth in additional ways to the PKB pathway, which are these, and how do they work? |
|
Definition
| activation of el4E: txn factor that stimulates txn activation of s6kinase: phosphorylates the ribosomal protein S6, leading to increased ribosome activity. So, total protein synthesis is increased. |
|
|
Term
| [Apoptosis] In what two ways do survival signals suppress apoptosis? |
|
Definition
| PKB pathway: survival ligand plugs into survival receptor, activating PKB pathway, which phosphorylates BAD, activating Bcl2, thus suppressing apoptosis. MAPK pathway: inactivates Hid, an IAP(apoptosis inhibitor) inhibitor, thus inhibiting apoptosis? |
|
|
Term
| [Cancer] What are the two general classes of cancer causing genes? |
|
Definition
| tumor suppressors, oncogenes |
|
|
Term
| [Cancer] What are the three cancer types? |
|
Definition
| Carcinoma: from epithelials, sarcoma: from connective tissues or muscles, leukemia: from blood cells |
|
|
Term
| [Cancer] What are the six major characteristics of cancer cells? |
|
Definition
| 1. activation of proto-oncogenes, leading to self-sufficiency in growth signals 2. insensitivity to anti-growth signals, because of reduced checkpoint control and reduced tumor supressor genes. 3. evasion of apoptosis by inhibiting apoptosis and increased cell survival. 4. limitless replication because of increased telomerase activity. 5. invasion of tissue and metastasis because of cell adhesion breakdown. 6. sustained angiogenesis |
|
|
Term
| [Cancer] [Development] What is the difference between a primary and secondary tumor? |
|
Definition
| primary: tumor that is at the site where it first arose. secondary: tumor that has spread from the original site |
|
|
Term
| [Cancer] [Development] usually develops from a single abnormal cell |
|
Definition
|
|
Term
| [Cancer] [Development] What does cancer result from? |
|
Definition
| a somatic mutation that can be family related or carcinogen related |
|
|
Term
| [Cancer] [Development] Cancer cannot develop from a single mutation. many must occur (10^10 separate occasions of DNA mutations) |
|
Definition
|
|
Term
| [Cancer] [Development] Experiment: What differences were seen between a mouse with mutant myc, another with mutant ras, and another with both mutations? |
|
Definition
| cancer a lot more prevalent in mice with both mutations than with either mutation alone |
|
|
Term
| [Cancer] [Development] Experiment: what differences were observed between Ras mutation in immortalized cells versus Ras mutation in normal cell lines? |
|
Definition
| Ras was able to transform immortalized cells into cancer, but not primary cells lines. |
|
|
Term
| [Cancer] [Development] colorectal cancer provides a great example of how cancer is a multistep process... describe this process |
|
Definition
| Loss of APC tumor suppressor gene --> polyp. activation of K-ras oncogene. loss of tumor suppressor gene in DCC (deleted in colon carcinoma). Loss of p53 tumor suppressor gene. |
|
|
Term
| [Cancer] [Development] tumor progression |
|
Definition
| single abnormal cell with multiple mutations -- mitogen encourages growth -- unlimited replication -- invasive -- metastasis -- does not respond to antigrowth signals or apoptotic signals -- loss of DNA repair (damaged DNA polymerase or mutations) |
|
|
Term
| [Cancer] [Development] What are possible origins of cancer stem cells? |
|
Definition
| Stem cells depend on their niche, which limits their expansion. 1. expansion of the stem cell niche can allow expansion of cancer cells that arose from normal stem cells. 2. cancer stem cells adapt to a different niche altogether, allowing them to expand. 3. cancer stem cells become niche independent, and so self renew and are autonomous. 4. cancer stems arising from a progenitor cell with decreased replication inhibition potential. |
|
|
Term
| [Cancer] [Development] Describe the pathways involved in instestinal stem cells |
|
Definition
| BMP (bone morphogenetic protein) inhibits stem cells. Noggin inhibits the inhibition caused by BMP, so it is a pro-cancer. Wnt activates the stem cell, and Dkk inhibits the inhibition caused by Wnt, thus activating the stem cell. |
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|
Term
| [Cancer] [Development] What is BMP? |
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Definition
| it is a member of the TGFbeta family, and inhibits stem cells. Noggin is a TGFbeta inhibitor |
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|
Term
| [Cancer] What is a neoplasm/benign tumor? |
|
Definition
| tumor mass whose growth regulation is out of control, but remains clustered together and contained... no metastasis |
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|
Term
| [Cancer] What is a malignant tumor? |
|
Definition
| tumor whose cells can break away to invade, by secreting proteases or protease activators |
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|
Term
| [Cancer] What is a metastasis tumor? |
|
Definition
| secondary tumor, located away from the original site |
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Term
| [Cancer] Adenoma vs adenocarcinoma |
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Definition
| adenoma is benign, inside a capsule... adenocarcinoma is malignant, can spread |
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Term
| [Cancer] In normal cells, what restricts their migration? |
|
Definition
| ECM, such as the basal lamina |
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Term
| [Cancer] Why do malignant cells secrete proteases/protease activators? |
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Definition
| this allows them to degrade the ECM and invade the bloodstream |
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|
Term
| [Cancer] How do malignant cells spread? |
|
Definition
| secrete proteases/protease activators that degrade the ECM |
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Term
| [Cancer] Give an example of "normal" metastasis |
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Definition
| white blood cells circulating the blood stream and then invading tissue to do its thing |
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Term
| [Cancer] What appendage do cancer cells use to degrade the basement membrane and invade through the ECM? |
|
Definition
| invadopodium, formed by actin, N-WASP, Cofilin |
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|
Term
| [Cancer] What are the components of invadopodia? |
|
Definition
| acting regulation, signaling/adpators, adhesion, proteases, membrane remodeling |
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|
Term
| [Cancer] in invadopodia, what are the components of actin regulation? |
|
Definition
| cortacting, N-Wasp, Arp2/3 complex, WIP, cofilin, tafin |
|
|
Term
| [Cancer] What are the components that invadopodia use for signaling? |
|
Definition
| cdc42, Nck1, p190RhoGAP, FAK... |
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|
Term
| [Cancer] What tools do invadopodia use for adhesion? |
|
Definition
| integrins, vinculin, paxillin, tensin |
|
|
Term
| [Cancer] What is angiogenesis? |
|
Definition
| the process of making new blood vessels to sustain new cells and growing cells |
|
|
Term
| [Cancer] What do tumors secrete that promotes angiogenesis? |
|
Definition
| HIF: hypoxia inducible factor (usually only present during low levels of O2). VEGF: vascular endothelial growth factor |
|
|
Term
| [Cancer] What can one use to inhibit angiongenesis? |
|
Definition
| endostatin and angiogenin |
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|
Term
| [Cancer][Cell Culture] What does transformation mean? |
|
Definition
| process by which culture cells lose normal characteristics and acquire cancer characteristics |
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Term
| [Cancer][Cell Culture] What are the characteristics of normal cells in culture, which resemble noncancerous cells in the body? |
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Definition
| have limited replication ability; are anchorage dependent because they cannot grow in suspension..if they are removed from their ECM, they apoptose; have contact inhibition, grow a monolayer and do not pile up on top of each other; are dependent on growth factors so cannot survive under low serum growth factor conditions..... TRANSFORMED CELLS LOSE THESE CHARACTERISTICS |
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|
Term
| [Cancer][Cell Culture] What is the difference between normal cells and immortal cells? |
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Definition
| normal cells cannot survive long in vitro because they undergo crisis. immortal cells, on the other hand, have unlimited replication ability... during crisis... small number of cells survive and begin proliferating... thus selected for to survive in culture... can escape crisis after experiencing genetic mutations |
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|
Term
| [Cancer][Cell Culture] What is required for immortality? |
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Definition
|
|
Term
| [Cancer][Cell Culture] In most cells, contact with other cells causes them to stop growing -- contact inhibition |
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Definition
|
|
Term
| [Cancer][Cell Culture] Describe the NIH3T3 model |
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Definition
| NIH3T3 cells are immortalized fibroblasts that have been derived from mouse embryos that survived "crisis" in vitro.... immortal... NOT transformed... since they are already immortal, they are very likely to transform |
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|
Term
| [Cancer][Cell Culture] Describe the experiment used to describe Ras |
|
Definition
| grow immortal cell line in vitro and give the dish DNA from human tumors. Transformed cells grow a focus. extract DNA from cells, transform new mouse cells...extract genomic DNA from these and insert into bacteriophages and grow them... the one with the transformed DNA will survive. |
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|
Term
| [Cancer][Cell Culture] Describe how oncogenicity is determined in vivo |
|
Definition
| Use nude mice: no thymus, no T-cells, and therefore no immunorejection of harmful cells. |
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|
Term
| [Cancer] cancer causing viruses have cellular counterparts... proto-oncogenes that are active in tumors... name some |
|
Definition
|
|
Term
| [Cancer] What is a common phenomenon in chromosomes in cancer? |
|
Definition
| loss of heterozygosity: you have two alleles for a gene that codes for a tumor suppressor... first hit kills one of the alleles... buts it's ok becaue you have the other copy to code for the tumor suppressor (this mutation can be passed on to children). The second hit is the mutation that deletes the other allele, thus no longer code for the tumor suppressor. |
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|
Term
| [Cancer] What is a microarray used for? |
|
Definition
| to identify differences in gene expression between normal cells and cancer cells. red=overexpression, green = underexpression. Transformed cells show a similar pattern to cancer cells. |
|
|
Term
| [Cancer] in a microarray, what is cluster analysis? |
|
Definition
| genes are grouped together based on some criteria, such as function or expression pattern |
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|
Term
| [Cancer] in a microarray, what does the x-axis and y-axis represent? |
|
Definition
| x = individual RNA samples. y = individual genes |
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|
Term
| [Cancer] What types of proteins are involved in cancer? |
|
Definition
| growth factors and their receptors, signal transduction proteins, transcription factors, proteins that control cell cycle(often act as tumor suppressors), DNA repair proteins, pro- or anti-apoptotic proteins |
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|
Term
| [Cancer] genes involved in cancer can be classified into two categories |
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Definition
| oncogenes: cause cancer when activated/expressed at high levels -- gain of function mutations (proto-oncogenes, genes that when mutated become oncogenes). tumor suppressors: genes that cause cancer when their function is lost, caused by mutation or inactivation, usually recessive mutations |
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|
Term
| [Cancer] What kinds of genes that cause cancer were discovered using viruses and transformation experiments? |
|
Definition
|
|
Term
| [Cancer] What kind of genes that cause cancers were discovered from hereditary cancers? |
|
Definition
|
|
Term
| [Cancer] What are caretaker genes? |
|
Definition
| they are responsible for the repair or prevention of DNA damage... if mutated (lof), mutations can accumulate |
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|
Term
| [Cancer][oncogenes] Name the ways in which a proto-oncogene can be converted to an oncogene |
|
Definition
| 1. point mutation or deletion that activates the protein -- hyperactive protein made in normal amounts (protein different from normal) 2. DNA amplification that leads to overexpression of a proto-oncogene (same protein, higher levels). 3. chromosomal translocation that results in a fusion protein (normal protein, higher levels). 4. chromosomal translocation that places a proto-oncogene under the control of a strong promoter (fusion protein that is overproduced, or fusion protein is hyperactive) |
|
|
Term
| [Cancer] Mutated genes in cancer: |
|
Definition
| Src, Ras, APC, Bcl2, p53, Rb, betaCatenin, Myc |
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|
Term
| [Cancer][oncogenes] What are the two basic results from activation of oncogenes? |
|
Definition
| more activity per protein molecule, more proteins |
|
|
Term
| [Cancer][oncogenes] gene amplification can be caused in which two ways? |
|
Definition
| 1. the duplicated DNA is jointly organized at a single site on the chromosome (homogeneously staining region). 2. duplicated DNA exists as small, independent mini chromosomes, resulting in staining of "minute chromosomes". Stained using FISH (fluorescent in situ hybridization) |
|
|
Term
| [Cancer][oncogenes] What are the genes associated with colorectal cancer? |
|
Definition
| oncogenes: K-Ras, BetaCAtenin. tumor suppressors: p53, APC, Smad4, TGFbeta receptor II, MLH1 and other DNA mismatch repair genes |
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|
Term
| [Cancer][oncogenes] What is APC and what does it do? |
|
Definition
| causes polyps... it is a major gene involved in causing colorectal cancer, functions in the Wnt pathway |
|
|
Term
| [Cancer][oncogenes] In the Ras and MAPK pathway, what kinds of mutations lead to cancer? |
|
Definition
| 1. mutation at the 12th residue (G12V) that inhibits Ras from binding to GAP, inhibiting its GTPase activity, so it is always bound to GTP -- constitutively active. 2. mutation at the 61st residue (Q61R) leads to low GTPase activity, causing constitutive activation of Ras |
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|
Term
| [Cancer][oncogenes] Src pathway: how is it regulated normally, and how can it lead to cancer? |
|
Definition
| Src tyrosine kinase is phosphorylated and thus inactivated. if it is missing its C terminues, it cannot be phosphorylated/inactivated, so it is constitutively active -- activates Ras and MAPK pathways |
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|
Term
| [Cancer][oncogenes] How is Myc involved in cancer? |
|
Definition
| Myc usually regulates cell cycle via Rb, but is often amplified in cancer and is also often mutated in others. In Burkitt's lymphoma, cMyc is constitutively activated because of chromosomal translocation that places it next to a promoter. |
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|
Term
| [Cancer][oncogenes] Describe how Abl Tyrosine Kinase causes cancer |
|
Definition
| a chromosomal translocation that causes a fusion between Abl and Bcr, which causes constitutive activation of the Abl kinase. This activation causes chronic myelogenous leukemia... secondary mutation in p53 causes acute leukemia which is often fatal... treatment: Gleevec, an inhibitor of Abl |
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|
Term
| [Cancer][oncogenes] Give an example of an oncogenic ligand mutation (rare) |
|
Definition
| sis oncogene-platelet derived growth factor (PGDF) |
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|
Term
| [Cancer][oncogenes] Mutations that activate RTK's |
|
Definition
| 1. point mutation in Her2 receptor, which induces dimerization independent of the ligand. 2. EGF receptor truncation that leads to constitutive dimerization. 3. Her2 amplification found in many breast cancers, so treated with Her2 antibodies. 4. chromosomal translocation that fuses the coil-coil region of tropomyosin to the intracellular domain of the Trk receptor 5. oncogenesis that produces a ligand that dimerizes an Epo receptor, thus activating it (viral) |
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|
Term
| [Cancer][tumor suppressors] What are the 6 classes of tumor suppressors? |
|
Definition
| 1. intracellular proteins that regulate/inhibit progression through a point in the cell cycle (p16 and Rb) 2. receptors/signal proteins that inhibit cell proliferation (TGFbeta pathway) 3. negative regulators of pathways that promote proliferation (APC). 4. checkpoint control of proteins that stop the cell cycle if DNA is damaged or chromosomes are abnormal. 5. apoptosis promoting proteins. 6. enzymes that repair DNA |
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|
Term
| [Cancer][tumor suppressors] p53: what is it? |
|
Definition
| it is a DNA damage checkpoint during cell cycle (remember?). usually degraded, but is accumulated upon DNA damage. activated by ATM kinase. induces expression of Cdk inhibitor, p21, and apoptosis. Look at |
|
|
Term
| [Cancer][tumor suppressors] What activates p53? |
|
Definition
|
|
Term
| [Cancer][tumor suppressors] APC protein: adenomatous polyposis coli: What is it? |
|
Definition
| a protein involved in the Wnt pathway. Deletion or inactivation of APC causes cancer |
|
|
Term
| [Cancer][tumor suppressors] What is the TGFbeta pathway, and how can it cause cancer? |
|
Definition
| inhibits cell proliferation. interruption of this pathway leads to increased cell proliferation. |
|
|
Term
| [Cancer][tumor suppressors] What is Rb, and how can it cause cancer? |
|
Definition
| Rb is an inhibitor of the G1/S transition, so its mutation leads to uncontrolled transition to DNA synthesis and contination of the cell cycle. study figure 20.33 and 25.25 |
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|
Term
| [Cancer][tumor suppressors] Discuss heterditary vs sporadic mutations |
|
Definition
| Hereditary blastoma: inherit an Rb- allele, then later, mutation leads to the other allele being Rb- as well. Sporadic retinoblastoma: Rb+Rb+, then first mutation, then second mutation |
|
|
Term
| [Cancer][tumor suppressors] What are the two mechanisms that can result in loss of heterozygosity? |
|
Definition
| start out with a mutant allele and a normal allele. Mis-segregation and mitotic recombination (study figure 25-14) results in homozygosity for mutant alleles |
|
|
Term
| [Cancer][tumor suppressors] What pathways promote stem cell fate in adults? |
|
Definition
|
|
Term
| [Cancer][tumor suppressors] What are cdk inhbitors? |
|
Definition
| tumor suppressors (p15INK4b, p16INK4a, p19ARF)... deletion of these leads to cancer |
|
|
Term
| [Cancer][Limitless replication] What is senescence? |
|
Definition
|
|
Term
| [Cancer][Limitless replication] Is telomerase usually expressed in culture cells? |
|
Definition
|
|
Term
| [Cancer][Limitless replication] What is the purpose of the telomere? |
|
Definition
| since the DNA ends are hard to replicate, telomere is present, which are tandem repeats of GGGTTA for 10,000 nucleotides... telomerase recognizes the telomere and elongates in 5 to 3 direction... 3" is slightly longer than 5" end... so each time cell divides, it loses 50-100 nucleotides |
|
|
Term
| [Cancer][Limitless replication] What happens normally when a telomere is lost? |
|
Definition
| the cell will stop dividing due to the p53 cell cycle check point |
|
|
Term
| [Cancer][Limitless replication] cells with p53 loss and loss of cell cycle checkpoint with survive and proliferate |
|
Definition
|
|
Term
| [Cancer][Limitless replication] What end is protected from degradation? |
|
Definition
|
|
Term
| [Cancer][Limitless replication] Usually, human cells have low telomerase levels, but cancer cells have increased telomerase...and shortened telomeres leads to chromosome instability and cancer |
|
Definition
|
|
Term
| [Cancer][Cancer Therapy] What are the 4 main classes of drugs used to treat cancers? |
|
Definition
| small molecule drug inhibitors (inhibit oncoproteins), monoclonal antibody inhibitors, proteasome inhibitors, naturally occurring proteins |
|
|
Term
| [Cancer][Cancer Therapy] Describe small molecule drug inhibitors |
|
Definition
| Gleevec inhibits bcr-Abl oncoprotein in CML; Nolvadex, an antiestrogen for breast cancer; iressa inhibits EGF-R for lung cancer; RAD-OO1 and CCI... are rapamycin analogs that inhibit mTOR |
|
|
Term
| [Cancer][Cancer Therapy] Describe monoclonal antibody inhibitors |
|
Definition
| Herceptin inhibits EGF receptor 2 in breast cancer; Avastin inhibits angiogenesis by targeting VEGF in colorectal cancer |
|
|
Term
| [Cancer][Cancer Therapy] Describe proteasome inhibitors |
|
Definition
| Velcade inhibits a proteosome |
|
|
Term
| [Cancer][Cancer Therapy] Describe naturally occurring proteins |
|
Definition
| endostatin inhibits angiogenesis |
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