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BIO230 Midterm Review.docx

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University of Toronto St. George
Desveaux/ Harris

BIO230 Midterm Review 1. Prokaryotic Transcriptional Regulation Genetic Switches Trp Operon Lac Operon Bacteriophage Lambda Synthetic Biology Transcription Attenuation 2. Eukaryotic Transcriptional Regulation DNA looping Co-activator and Co-repressors does not directly bind to DNA Eukaryotic transcriptional activation o Eukaryotic Activator Protein: Modular Design (BD & AD) Can bind directly alters chromatin structure Ways to alter chromatic structure o Nucleosome sliding (ATP-dependent) o Nucleosome removal (histone chaperones) o Histone exchange (histone chaperones) o Histone modifying enzyme o Attract, Position and modifying: General Transcription factor Mediator RNA polymerase II Eukaryotic transcriptional repression o Interfering with activator function (block BD) o Interfering with activator function (block AD, interaction with the general transcription factors) o Altering chromatin structure Recruitment of chromatin remodeling complexes Recruitment of histone deacetylases Recruitment of histone methyl transferase . Epigenetic inheritance 3. Regulation of the Transcriptome Post-transcriptional regulation o RNA processing RNA capping (additional of a modifies Guanine nucleotide to 5 end) Cap bound by Cap-bounding complex RNA splicing (removing introns of the RNA by splicosome) Site of proper splicing bound by Exon Junction Complex ( serve as a marker for properly splices RNA) Different cells can splice an RNA differently: Alternative splicing o Regulation of alternative splicing Positive and negative control Drosophila sex determination Sex-lethal: splicing repressor Transformer: splicing activator Doublesex: regulated sex gene expression 3 Polyadenylation RNA polymerase transfers protein complexes to RNA o CstF (cleavage stimulation factor) o CPSF (cleavage and polyadenylation specificity factor ) Poly-A polymerase(does not require template, not encode in the genome) add 200A nucleotide, bound by Poly-A binding protein Aid in RNA export and translation Coupling of transcription and RNA processing The C-terminal Domain of the RNA polymerase binds RNA processing proteins and transfer them to RNA at appropriate time Regulated by different phosphorylation on the polymerase allows the binding of different complex at different time. RNA nuclear export o The cell must have: 3 mature markers added (Cap binding complex, exon junction complex, poly-A-binding protein) those proteins travel with the mRNA to the cytosol. And 1 immature marker removed: proteins involved in RNA splicing (snRNPs) in order to export. o Improperly processed mRNA will eventually be degraded in the nucleus by exosome. o RNA transport from the nucleus can also be regulated: ex. human HIV HIV: consist of 2 RNA molecules, and a reverse transcriptase (creates double stranded DNA from RNA). The DNA is integrated in the host genome. To hijack the host cell, the HIV used the hosts polymerase to transcribe RNA, export, and make new viral protein. Challenge: need to export and overcome the check points. HIV transcribes into 30 different mRNA, and some of them retain introns. Have REV (mRNA with no introns, like a regular RNA) REV protein binds to REV response elements in unsliced RNAs, interacts nuclear export receptor and help it export, also allow HIV to divide. mRNA quality control o Eukaryotic initiation factors (eIFs): quality control of RNA, recognizing poly-A tail and 5 cap. eIF4E: complete off with CBC and bound to 5 Cap. eIF4G bound to poly-A bounding protein o Once the eIFs are bound, it recruit small ribosomal complex which will initiate translation at first AUG (exception: leaky scanning). o Nonsense-mediated mRNA decay: function of EJC in translation regulation, occurs as mRNA coming out the nucleus. When the mRNA first exiting the nucleus, it is met by a ribosome, EJC are displaced by the ribosome, the normal stop codon is reached when no EJC remains (stop codon must occur last). PASS! If the ribosome reach a premature stop codon (EJCs remains on mRNA when ribosome reaches stop), Upf will trigger the mRNA degradation. o Prokaryote mRNA quality control o Ribosome stalls on broken mRNA and do not release, it recruit a special tmRNA which carries an alanine to the A site (tRNA), and translated 10 more codons to the nucleotide. This tag is recognized by proteases that degrade the entire protein. mRNA stability o In prokaryotes, exonucleases rapidly degrade most mRNAs to adapt to the changing environment rapidly. o in eukaryotes, mRNA degradation is regulated by gradual shortening of poly-A- tail, process carried by deadenylase (act as a timer of mRNA lifetime) Once poly-A reaches a critical length 1) removal of 5 cap follow by a 5
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