Term
| What are the five major mechanisms that eukaryotes use to regulate gene expression? |
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Definition
Transcription initiation
Transcription termination
Processing/transport of mRNA, tRNA
Message stabilization
Translation |
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Term
| At the eukaryotic transcriptional level, what are the elements that can be cis-acting? |
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Definition
1. Core Promoter Sequence
2. Proximal Promoter Sequence
3. Enhancer/Silencer/Insulator
4. Response Element Sequence |
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Term
| What is the core promoter sequence? |
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Definition
The minimal portion of the promoter required to properly initiate transcription.
It contains the binding site for RNA polymerase and also the binding sites for general transcription factors.
Core promoter sequences can be cis-acting or trans-acting |
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Term
| What is the proximal promoter sequence? |
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Definition
the proximal sequence upstream of the gene that tends to contain primary regulatory elements
it contains specific transcription factor binding sites
it can regulate the TATA box or the CAP site
it can be cis-acting or trans-acting |
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Term
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Definition
An Insulator is a genetic boundary element that plays two distinct roles in gene expression, either as an enhancer-blocking element, or more rarely as a barrier against condensed chromatin proteins spreading onto active chromatin.
The need for them arises where two adjacent genes on a chromosome have very different transcription patterns because their function is to prevent a gene from being influenced by the activation (or repression) of its neighbors. |
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Term
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Definition
A silencer is a DNA sequence capable of binding transcription regulation factors termed repressors.
Upon binding, RNA polymerase is prevented from initiating transcription thus decreasing or fully suppressing RNA synthesis. |
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Term
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Definition
An enhancer is a short region of DNA that can bind proteins called an activator.
binding of activators to this enhancer region can initiate the transcription of a gene that may be some distance away from the enhancer, or can even be on a different chromosome.
The increase in transcription is due to the activators recruiting transcription factors, which enhances the binding of RNA polymerase. |
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Term
| What elements in eukaryotic transcription can be trans-acting? |
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Definition
1. Transcription Factors – “Core Promoter element”
2. Gene Regulatory Factors
Proximal Promoter element
Enhancer/Silencer/Insulator (CTCF)
3. Other Factors – Response elements |
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Term
| What is a response element? |
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Definition
| A response element is a short sequence of DNA within the promoter of a gene that is able to bind a specific hormone receptor complex and therefore regulate transcription |
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Term
| What is CTCF and why is it important? |
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Definition
CTCF is CCCTC-binding factor
a transcriptional regulator protein with 11 highly conserved zinc finger (ZF) domains. A nuclear protein able to use different combinations of the ZF domains to bind different DNA target sequences and proteins.
Depending upon the context of the site, it can function as a transcriptional activator or a transcriptional repressor. It can also bind to an insulator element and block communication between enhancers and upstream promoters. |
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Term
| What are the different possible effects of CTCF binding to the insulator? |
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Definition
It can inhibit elongation of the RNA
It can silence the initiation of transcription
It can function as a boundary and prevent the enhancer from enhancing the promoter it is supposed to |
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Term
| What is CTCF binding dependent on? |
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Definition
| Since it can bind to the insulator, silencer, or enhancer, where it binds is dependent on which site is more accessible. |
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Term
| How do proteins bind to DNA sequences? |
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Definition
Proteins recognize a specific DNA sequence because the surface of the protein is extensively complementary to the special surface features of the double helix in that region.
The protein makes a large number of contacts with the helix involving hydrogen bonds, ionic bonds and hydrophobic interactions.
Each contact is weak, but the combined total ensure the interaction is the tightest and most specific molecular interactions in biology.
Many of these proteins contain a small set of DNA binding structural motifs. |
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Term
| What are some examples of DNA binding structural motifs? |
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Definition
homeodomain proteins contain Three a-helicies where helix 2 and 3 are in a “helix-turn-helix” motif
CAP-binding protein is an example
the zinc finger (as in CTCF) where a Zinc cation cross-links 2 Cys and 2 His or 4 Cys and creates a loop
the leucine zipper motif - amphipathic and dimeric
the helix-loop-helix motif - amphipathic and dimeric |
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Term
| What is the role of antisense RNA in regulation of transcription? |
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Definition
There are some genes where only one copy of the gene needs to be transcribed (and eventually expressed)
An example is the gene for the Insulin-like growth factor receptor 2
In the maternal gene the upstream promoter is active and the downstream set of CpG islands are methylated (thus inactive) and regular mRNA transcription occurs
In the paternal gene the upstream promoter is inactive (methylated) but the downstream promoter is active and transcribes an antisense RNA.
The antisense RNA is theorized to participate in shutting the gene down (only the paternal gene) |
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Term
| What are the different inheritance possibilities of insulin-like growth receptor 2 (Igf2r) that could cause problems related to antisense RNA? |
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Definition
Accidental (in humans) inheritance of two copies of a particular chromosome from one parent and none from the other parent is usually fatal (even though a complete genome is present).
Inheritance of two copies of one of mother's genes and no copy of the father's (or vice versa) can produce serious developmental defects.
two copies of the mother's gene would cause too much mRNA to be produced.
two copies of the father's gene would cause too little mRNA to be produced. |
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Term
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Definition
siRNA = small interfering RNA or short interfering RNA or silencing RNA
It is first transcribed into preRNA that has ds hairpins.
A specific enzyme recognizes the ds RNA and cleaves it forming short fragments of 20-25 bp that are called siRNA |
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Term
| What is the function of siRNA in regulation of gene expression? |
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Definition
siRNA bind to the RNA-induced silencing complex (RISC) and activate it by unwinding from ds to ss.
The activated complex binds to the corresponding mRNA using the antisense RNA
The complex contains an enzyme which cleaves the mRNA thus suppressing gene expression
it has been noted that humans can use siRNA to silence viral RNA in the case of HIV-1 |
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Term
| What is miRNA and how does it compare to siRNA? |
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Definition
miRNA = microRNA
miRNA plays a similar role to siRNA (suppressing transcription) but also has some different characteristics.
mature miRNA is single stranded while mature siRNA is double stranded
siRNA has to unwind into a single strand before it can activate and bind to RISC but mature miRNA is already single stranded |
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Term
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Definition
Riboswitch is a form of RNA that act as a precision genetic switch.
Produced in many cases from noncoding DNA between known genes, a riboswitch folds into a complex shape. One part of the folded RNA (the aptamer) can bind to a specific target protein or chemical. Another part (the expression platform) contains the RNA code for a protein product.
The riboswitch turns "on" and produces the protein it encodes only when in the presence of its target. |
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Term
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Definition
slightly different proteins that are tissue specific,
one gene can code for many different isoforms that are expressed in a variety of differentiated cell types.
ex - the dystrophin gene (the longest gene in the human genome) is known to have 8 promoters each with its own initial exon and the gene may have as many as 79 exons downstream |
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Term
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Definition
the use of different, unique and often tissue-specific promoters
alternative splicing - a gene with multiple splice sites has the option to create several different exon combinations based on what is/is not spliced
the use of different polyA-addition signals - where the poly A tail is added will determine how long the mRNA transcript is |
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Term
| What are the basics behind cancer? |
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Definition
Stable, heritable changes in the DNA of cells.
Stepwise accumulation of these changes in regulatory genes leads to the capacity for malignant growth.
Requires several, independent events that are not corrected.
From 3 (in leukemias) to 7 (in carcinomas) events are required to turn a normal cell into a malignant cell.
Every gene will have about 1010 spontaneous mutations in a lifetime. Points out fidelity of the cellular correction machinery!!!!!!!!! |
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Term
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Definition
| normal gene whose product involved in stimulation of cell cycle. |
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Term
| What causes the change from a protooncogene to an oncogene? |
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Definition
A mutation that makes an inducer regulatory protein become hyper stimulatory.
The mutation only has to occur on one allele |
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Term
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Definition
Its a tumor Suppressor a gene whose product involved in suppression of the cell cycle.
ex's - p53 (damn Thomas) |
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Term
| How does an antioncogene lose its function? |
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Definition
| Unlike a protooncogene, an antioncogene must have mutations on both alleles in order to cause cancer |
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Term
| What is loss of heterogeneity? |
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Definition
LOH is loss of one of two alleles on a chromosome due to increasing aberrant behaviour of replication and recombination machinery.
Thus, one allele will have a mutation introduced into it (like APC). Through nondisjunct recombination or gene conversion or etc the second allele will be lost leading to only the mutated allele expressing a product or fixing homogeneity of that locus. |
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Term
| Why are people with inherited syndromes more susceptible to cancer? |
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Definition
| They start with one or more "hits" or separate genetic events that appear to be necessary to generate the neoplastic (cancer) phenotype. Some cancers only require 3 events while others 5-7 |
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Term
| Why is p53 so damn important? (Mutations in p53 occur in over 50% of human tumors) |
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Definition
It causes a pause in the cell cycle at G1/S when the cell senses DNA damage. Does this by inducing transcription of p21 (and other molecules like MDM-2 (HDM), and Bax) which binds and blocks phosphorylation of G1 cyclin/Cdk2 complexes.
This allows DNA repair to restore genome integrity, followed by release from the arrest. Alternatively, it induces a sensor molecule which, if cell cannot repair damage, promotes permanent exit from the cell cycle and apoptosis.
Mutations in p53 disable this “emergency brake” allowing the cell to replicate its defective genome, and stably introduce those mutations into the genome. |
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Term
| What is Li Fraumeni syndrome? |
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Definition
Inheritance of mutated allele of p53 (germline mutation).
Predisposed to increased risk of breast, brain, adrenocortical tumors, sarcomas, and leukemia.
It is an example of inherited syndrome predisposing one to cancer |
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Term
| How can the activity of p53 be disrupted? |
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Definition
Transcription activation by p53 can be disrupted by p53 mutations which:
1. prevent sequence specific DNA binding.
2. formation of active p53 oligomers.
3. interaction with the transcription machinery.
Also, overexpression of other gene products, like Mdm2 protein can prevent p53 interaction with initiation complex. |
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