mid 27 - Modification in Eukaryotes: RNA processing mRNA...

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Post-transcriptional Modification in Eukaryotes: RNA processing • mRNA processing (pp 111-115; 493-505) 1. 5’ Capping 2. Poly(A) adenylation 3. splicing
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Splicing RNA Processing 1. 5’ Cap 2. 3’ Poly(A) tail addition 3. Splicing
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5 Cap functions: – transport out of the nucleus – initiation of translation – protect from exonucleases RNA Processing: 5’ Cap •Eukaryotic mRNAs have a 5’ cap formed by a 5’-5’ linkage of a 7-methylguanylate (GTP) residue. •Catalyzed by a dimeric capping enzyme, which associates with the phosphorylated CTD of RNA polymerase II. -Capping is specific for Pol II Since Pol I or III does not contain a CTD, the capping enzyme does not associate with either of these RNA polymerases
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Poly(A) polymerase (PAP) adds A residues to the 3’ end 200-250 Adenosine residues can be added to make the complete Poly(A) tail Poly(A) tail functions to protect the mRNA from premature degradation by 3’ 5’ exonuclease RNA Processing: Polyadenylation G/U
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RNA Processing: Splicing
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RNA Processing: Splicing • Four conserved sequences have been found to be necessary to remove the introns: - 5’ splice site at the exon/intron junction (GU) - Branch point (Adenosine): usually 25-50 bases from the 3’ splice site - Pyrimidine-rich region upstream of the 3’splice site -3’ splice site at the intron/exon junction (AG) • Generally, the central region of the intron is unnecessary for splicing to occur.
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Splicing: 2 Transesterification Rxns • In the first reaction, the ester bond between the 5 phosphorous of the intron and and the 3 oxygen of exon 1 is exchanged for an ester bond with the 2 oxygen of the the branch-site A residue . • In each transesterification reaction, one phosphodiester bond is exchanged for another. • In the second reaction, the ester bond between the 5 phosphorous of exon 2 and the 3 oxygen of the intron is exchanged for an ester bond with the 3 oxygen of exon 1 , releasing the intron as a lariat structure and joining the 2 exons.
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RNA Processing: Splicing • The splicing reaction is catalyzed by enzymes composed of a complex of small nuclear RNAs ( snRNA s) and proteins, referred to as small nuclear ribonucleoprotein particles or snRNP s (pronounced snurps ). • The RNA components of snRNPs are abundant, Uridine-rich snRNAs and are referred to as U1, U2, U4, U5 and U6 . They interact together to form the spliceosome . • Each snRNA is associated with 6-10 proteins
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• U2 base pairs to regions flanking the the “branch point” A -The branch point A itself, which is not base paired to U2 snRNA, “bulges out,” allowing its 2’ OH to participate in the first transesterification reaction. The 5’ region of U1 snRNA base pairs with the 5’ splice site of pre- mRNA.
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This note was uploaded on 03/05/2008 for the course LIFESCI 3 taught by Professor Staff during the Fall '06 term at UCLA.

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mid 27 - Modification in Eukaryotes: RNA processing mRNA...

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