2 - Gene Regulation Flashcards
Explain the central dogma of molecular biology succinctly.
Gene Regulation
The central dogma of molecular biology stipulates that DNA is transcribed to RNA which in translated into amino acids/proteins.
What is a control gene?
Gene Regulation
a control gene regulated the expression of other genes
How are the steps of the central dogma regulated?
Gene Regulation
Transcription:
1. chromatin structure modification through histone acetylation or DNA methylation (heterochromatin vs euchromatin)
2. transcription factors (activation or repression)
3. premature termination of transcription (synthesis issues)
4. splicing and alternative splicing (splicesome srRNA)
5. nuclear export (NES,exportin)
Translation:
1. mRNA degradation in cytosol
2. translational controls such as inhibitors/activators/enzymes/cofactors
What is a promoter? Why is the concept significant?
Gene Regulation
Transcriptional factors can bind to promoters upstream of the gene locus sequence that the promoter/TF combo affects. DNA Pol 2 and other TF can bind here.
promoter mutation results in low levels of gene expression
What is a Kozak Sequence? Why is it significant?
Gene Regulation
A kozak sequence is an initiation site in eurkaryotic mRNA (RCCATG, R is a purine - (Pur)e (A)s (G)old ). Kozak sequences support binding of small units of ribosomes to mRNA for translation.
mutations in sequence impairs initiation of translation, lowers protein
What are transcription factors? Where do they bind? What kinds are there?
Gene Regulation
Transcription factors are proteins that regulate gene transcription. The binding sites for TF’s are called regulatory elements. Enhancers or Positive regulatory elements (PRE) are DNA sequences where activating proteins bind to increase expression of a gene on the same chromosome. Silencers or Negative regulatory elements (NRE) are DNA sequences where repressor proteins bind to decrease gene expression on the same chromosome.
Enhancers/silencers ca be located proximal, far, or within (intron) the gene whose expression they regulate
How do transcription factors bind to DNA?
- Zinc Finger motifs: 4 cysteine or histidine residues bind to a zinc ion in major groove of DNA
- Leucine Zipper motifs: 2 distinct polypeptide chains that form hydrophobic interactions to dimerize
- Helix-turn-helix motifs: 3 or 4 helical regions that stabilize each other
- Helix-loop-helix motifs: helical regions that bind to DNA like leucine zippers. Dimerization is connected to DNA by a loop.
Gene Regulation
What is basal transcription machinery? What is it composed of? How does those components function?
Gene Regulation
Basal transcription machinery includes RNA polymerase and 6 general transcription factors (TF2A, TF2B, TF2D, TF2E, TF2F, TF2H). This is the minimum general requirement for transcription in all eukaryotic cells. There are 3 RNA polymerases (Pol 1, 2, 3) which are numbered in the order that theirproducts are used in protein synthesis: rRNA, mRNA, and tRNA. The most inportant basal transcription factors are TF2D and TF2H. TF2D recognizes the TATA box and regulated DNA binding. TF2H unwinds DNA at the transcription start point and phosphorylated a serine of the RNA polymerase to release it from the promoter.
Coactivators (mediator proteins) bind to the transactivation domains of transcription factors to enhance the assembly of the basal transcription complex.
How is RNA processed?
Gene Regulation
- primary transcript of RNA is heterogenous nuclear RNA (hnRNA)
- the hnRNA received a 7-methylguanosine cap on the 5’ end
- the 3’ end is polyadenylated to make a poly A tail. This is for stability and mutation in the polyadenylation signal leads to early degradation of mRNA before translation.
- introns are spliced out
mRNA is capped, tailed, and spliced.
mRNA quality control occurs at cytoplasmic processing bodies (p- bodies), which contain exonucleases; mRNAs may be degraded or storied in p-bodies for future translation.
What is RNA splicing? What facilitates it? What happens when splicing goes errently?
Gene Regulation
RNA splicing is the process by which precursor mRNA is transformed in mature mRNA. Splicing is mediated by the spliceosome which is made of small nuclear ribonucleoprotein particles (snRNP). If snRNP assembly is affected, vital proteins such as survival motor neuron proteins in spinal muscular atrophy, cannot be made and lead to weakness.
Why is splicing important?
Gene Regulation
With only a certain amount of amino acids and one hnRNA transcript, alternative splicing can produce a veriety of protein products from that one transcript.
Alternative splicing is carefully regulated and shows tissue-specificity
Why is mRNA stability during transport important? How is mRNA transported? How is it protected and regulated?
Gene Regulation
- mRNA stability is critical in regulationg gene expression ->mRNA that lasts longer can generate more protein
- mRNA is bound to proteins to help prevent is degradation (polyA tail). As mRNA ages, the polyA tails get shorter
- mRNA export proteins are docked to nascent mRNAs during elongation to account for transport
- Nuclear surveillance mechanisms ensure that only mature and function mRNP reach the cytoplasm
- nuclear mRNA transport depends on formation of transport competent mRNP’s through nuclear pore complexes
Can gene expression be regulated at the translational level? Give an example or explain the process.
Gene Regulation
Yes. For example, Ferritin (stores iron) is synthesized when iron levels increase. The mRNA for ferritin has an iron response element that can bind a regulatory protein. When this regulatory protein does not bind iron, it prevents initiation of ferritin translation. When the regulatory protein binds iron, it changes conformation, releasing the IRE and allowing mRNA translation.
What is Micro RNA (miRNA) and why is it important?
Gene Regulation
MicroRNAs (miRNAs) are small RNA molecules that regulate protein expression at a posttranscriptional level. miRNA can induce the degradation or block the translation of a target mRNA. miRNA is transcribed by RNA polymerase 2. pre-miRNA is modified in the nucleus by an RNA-specific endonuclease (Drosha) and exported to the cytoplasm where it is cleaved by the endonuclease Dicer. One of the strands of miRNA is incorportated in the RNA induced silencing complex (RISC) while the other strand is degraded. RISC will block translation of the target mRNA.
What is chromatin remodeling?
Gene regulation
At the DNA structural level, chromatin must be remodeled to allow access for RNA polymerase, which is accomplished, in part, by proteins with histone acetyltransferase (HAT) activity.
How can enhancer positioning or mutation cause disease?
Gene Regulation
Follicular B-Cell Lymphoma is caused by a 14:18 chromosome translocation that results in the over-expression of BCL-2. The BCL-2 gene normally exists on chromosome 18, but instead it gets placed into the immunoglobulin heavy chain locus on chromosome 14. The 14:18 translocation is present in the majority of follicular lymphomas and results in the uncontrolled expression of BCL-2. Increased levels of Bcl-2 protect the cells from apoptosis and leads to uncontrolled cell division and enhanced resistance to chemotherapeutic agents.
Disruption of an AP-2α binding site in an IRF6 enhancer is associated with cleft lip. Non-syndromic cleft lip with/without cleft palate (NSCL/P) is a common complex birth defect. Single nucleotide polymorphisms (SNPs) in IRF6 are association with NSCL/P.
How does abberant splicing cause disease?
Gene Regulation
Point mutations in the cis-acting splicing regulatory signals can lead to aberrant mRNA generation and disease.
