BIOL 302 1nd EditionExam # 3 Study Guide Lectures: 14 - 18Lecture 14Role of Chromatin/histone modifications in transcriptionYou cannot get DNA because there is limited access to the nucleosomes and chromatin due to itrepressing transcription; DNA cannot reach it. SWI/SWF: have enzymatic activities that modify to allow recognition by other factions and access to DNA sequences – there are 10 proteins in a complex. Hydrolysis of ATP is used to change the organization of the nucleosomes. Histones arechemically modified to allow recognition – Histone acetyl-tranferase (HAT) and Histone de-acetyl transferase (DHAT). HAT: adds an acetyl group that tends to make it active while DHAT removes it and inactivates it. The acetyl group makes it negatively charged due to the histones losing their grip on the DNA and the genes are turned on. Carboxyl terminal with the NH terminal are with all the positive charges to interact with the negative at neutral pH. Transcription TerminationBacteria cells have a termination signal in termination while eukaryotic cells do not. E Coli: can end the process by using the termination signal in the DNA that’s transmitted in the DNA by RNA; it releases polymerase and mRNA from template (ACTIVE). Properties: termination signal is in DNA and signal transmitted by RNA transcript (3 prime end – where there are a bunch of U’s in the sequence that undergoes internal base pairing). E. Coli Termination Sequence: it makes a stem loop structure with A and U’s at the end. RNA polymerase makes this transcript and synthesizes and forms a double stranded structure. It should be G-C rich stem loop of 8 or so nucleotides which makes it stable and all the U’s have to be at the bottom of the RNA. Eukaryotic “Termination”: transcription occurs and makes RNA molecule not functional messenger RNA. Sequence is required and presence in Eukaryotic cells. mRNA at 20-10pt 3” prime. CF: cleavage factors; CPSF: cleavage and polyadenylation specificity factor. Polydenylation signals a reaction that cuts 3’ end, but keeps polymerase keeps going but RNA falls off. Capping & polydenylation II Added by guanlytransferase which add an unusual nucleotide to 5 prime end part of elongation complex, and its added to the 5’ end soon after initiation and is capped before termination. Usually its AUG (the start codon). The first nucelotided has been phosphorylated. The guaninew/sugar added through the 5 prime prime carbon. 5 to 5 prime triphosphate bond. Guanine is modified w/methyl group. Every messenger has this cap added. It requires an AAUAA sequence at the 3’ end. As it cleaves poly A polyermase adds (50-250 nucleotides) and doesn’t require a template. It exports Lecture 15RNA SplicingRNA splicing removes introns from primary transcript which joins exons to generate mature mRNA. It’s split by introns and the exons are needed to be joined together. Within RNA molecule specific groups of sequence. There are three different components: 1. Specific Sequences: on the RNA that extinguishes introns and exonsExon 1: very highly conserved sequences (CAG – end of exon) and other sequence (GUAAGU). Exon 2: nucleotide from end of intron which is another conserved sequence (ACUAC) adenine is extremely important in the process2. Spliciosomes: recognizes and identifies the sequencesSplices RNA and doesn’t make protiens. 5 distinct RNP’s and 1 small RNA; large multi-protien sequence. the U1 RNA base pairs with the sequcnes at the end of the intron. U2 – small RNA that base pairs with the adenine group in the intron. 3. Auto – hydrolysis: RNA is active; self-contained Splicing defects: which destroys the splice set. Instead of the CAG, it’s CAU and it doesn’t recognize as an intron and completely skips. It causes numerous diseases due to inherited splicing defects. Alternative Splicing: differential and alternative use of exons which generates different mRNA. Particular transcript has more exons than 1 time and under circumstances. Lecture 16Nuclear membrane/nuclear pore complexVery sophisticated complex, in and out of the nucleus and highly regulated. mRNA laves the channel and into the cytoplasm. All the proteins take place in the cytoplasm because that is where transcription is taken place. The nucleus is double membrane (outer and inner – the space between is called perinuclear space). The ER: involved in transport, very complicated and goes through the whole complex. Annular subunits: can open & close the channel. Luminal subunits: hold between 2 membranes: hold into the 2 membranes; wing like structures sticking out perpendicular & maintain between the 2 membranes. Receptors (nucleoporines) whichrequires ATP to open up and allow mRNA into the cytoplasm and make there way to the ribosome. Post-Transcriptional Gene RegulationProtein stability: proteins exist for 12 hours it accumulates but 1 hour less accumulation. The amount of protein made is determined on transcription. Post-transcriptional determines amount of proteins usage of mRNA. mRNA stability: the longer it last the more proteins made. Protein Binding: control of translation of ribosomal protein. Proteins bind to rRNA to build ribosome. Cell is constructing ribosomes – proteins that self-assemble to make ribosomes; bindsto the stem loop. If there is no available rRNA the cell will stop making rRNA and shuts down translation. S7: binds to its own mRNA for it to shutdown translation.Lecture 17ER signal sequence: synthesis of a protein destined to be secreted from a cell. Ribosomes attached to ER signal sequence – synthesis of a protein destined to be secreted from a cell. Ribosomes attached to ER – goes through a sequence of events that moves from the ER into the golgi body carried by vesicles. They get glycosylated and packaged into secretory vesicles and fuses with cell membrane then gets sent out. Lecture 18Membranes and membrane proteins Ion channels and cell adhesion. Ions are voltage neurons because after stimulus electrical current on the membrane will open or close a particular channel. When it opens, millions are diffuse into or out of the cell. Calmodulin is calcium binding protein it has a regulatory but once it binds to calcium it changes conformation and takes on activity. Signal transduction pathway: receptor on cell surface binds to factor (channel opens up) where calcium floods in and change genetic features. Membrane proteins: barrier prevents that charged molecules from