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RNA Rearrangements in the Spliceosome

RNA Rearrangements in the Spliceosome. -extensive U4/U6 interaction is replaced with a U2/U6 structure -U1 displaced at 5’ splice site by U6. RNA Rearrangment at 5’ Splice Site. Why Splicing?. Why Splicing?. -ancient or new? -functional units (W. Gilbert)

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RNA Rearrangements in the Spliceosome

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  1. RNA Rearrangements in the Spliceosome -extensive U4/U6 interaction is replaced with a U2/U6 structure -U1 displaced at 5’ splice site by U6

  2. RNA Rearrangment at 5’ Splice Site

  3. Why Splicing?

  4. Why Splicing? -ancient or new? -functional units (W. Gilbert) -diversity of function, evolution -alternative splicing

  5. Exon Shuffling

  6. Sequence Conservation Within Introns 5’ Splice Site Yeast Human

  7. Sequence Conservation Within Introns Branch Region 3’ Splice Site Yeast Human

  8. Drosophila Doublesex (dsx) RNA

  9. Tissue and Developmental Regulation

  10. Alternative Splicing and Proteome Complexity Dscam 115 exons 20 constitutive, 95 alternatively spliced 38,016 possible proteins

  11. Mechanisms of Alternative Splicing -activation/repression -intron sequences: splice sites, pyrimidine tract, branch region -exon sequences (purine rich enhancers)

  12. Recall: Selection of 5’ Splice Site and Branch

  13. SR Proteins -S100 extract is deficient in pre-mRNA splicing -S100 + SR splices -SR proteins are phosphoproteins (SRPK kinase) -human SC35, 24 C terminal SR repeats

  14. RNA Recognition Motif (RRM) PTB RRM-3 Sxl RRM-1

  15. Activation -downstream activation of 5’ splice site -adenovirus E1A, also Tra/Tra2 Mol. Cell. Biol. (1999) 19, 7347

  16. “Enhancers”: -stimulate 5’ or 3’ splice site usage vs

  17. “Enhancers”: -stimulate 5’ or 3’ splice site usage vs

  18. “Enhancers”: -within exons -proximal to splice sites -conserved

  19. Science 297, 1007 (2002)

  20. Repression -formation of RNA secondary structure (blocking site access); relieved by ATP dependent helicases -binding of trans factors

  21. Drosophilasxl Gene

  22. sxl Regulation of Drosophila splicing -sxl competition with U2AF on dsx (also polypyrimidine tract binding protein -PTB; neuronal specific events)

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