Spring 09 lecture 27

Spring 09 lecture 27 - 11 mRNA processing: 5' Capping Soon...

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22 mRNA processing: 5’ Capping Soon after RNAPII starts making transcript (chain size 20-30 nucleotides)- co-transcriptional modification(s) 5’G 1st base in original transcript (Lewin, 2006. Fig.7.18) 5’ 3 modifications 7-met-guanosine coupled to 5´-end; binding through 5´-5´triphosphate bridge O2´-methylation of ribose in Cap1 and Cap2 N6-methylation of adenine possible in Cap1 and Cap2 (if adenine if first two positions)
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33 3 enzymatic reactions: 1. RNA 5´-Triphosphatase (RTP) removes the 5’ phosphate 2. Guanylyl transferase (GT) attaches GMP 1 and 2 are parts of the same protein in higher eukaryotes 3. 7-methyltransferase (MT) modifies the terminal guanosine 1. 2. 3. Phosphorylated CTD recruits capping enzymes (they bind directly to phosphorylated CTD) Remember: function of 5’methyl cap - protection from degradation but important role in splicing , nuclear transport and translation as well From: Cell, Vol. 108, 501–512, February 22, 2002 Integrating mRNA Processing with Transcription Nick J. Proudfoot, Andre Furger and Michael J. Dye
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44 mRNA processing: Polyadenylation Polyadenylation signal = polyadenylation consensus element (except in yeast) 5’-AAUAAA-3’ Alternative: 5’-AUUAAA-3’ Followed by 5’-YA-3’ (Y=pyrimidine, mostly C) in the next 11-20 nucleotides cleavage and poly(A) addition site
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55 3’ UTR: genomic and mRNA sequence Cytoplasmic adenylation element, T/U- rich; for polyadenylation of maternal messages G-T rich involved in polyadenylation remember: this sequence “stays” in mRNA
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66 Polyadenylation cont. CPSF - cleavage and polyadenylation specificity factor – binds to AAUAAA CStF – cleavage stimulatory factor- binds to G/U region CFI & II – cleavage factor I & II-bending (not shown) ; mRNA is cleaved PAP - poly (A) polymerase binds Following cleavage: 1. slow phase (first 6 12 nucleotides) 2. rapid phase - requires nuclear poly (A) binding protein II ( PABII )- 200- 250 nucleotides Role of CTD tail: to recruit enzymes necessary for polymerization
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77 Learning Objectives Describe the mechanism of RNA splicing including the role of RNA-RNA and RNA- protein interactions Define trans-splicing and its occurrence in different organisms Splicing of primary transcripts RNA processing & post-transcriptional control after transcription by RNAPII
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Processing of mRNA: Splicing Initially seen using electron microscopy (1977) R-looping technique Splice sites can be determined by comparing genomic DNA with cDNA or mRNA. Hybridization between globin precursor RNA and globin gene
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This note was uploaded on 08/03/2009 for the course BIOL BIOL308 taught by Professor Miskovic during the Spring '09 term at Waterloo.

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Spring 09 lecture 27 - 11 mRNA processing: 5' Capping Soon...

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