Gen511-3-12-08F

Gen511-3-12-08F - Gen511: Wednesday 12 March 2008 Chapter...

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Gen511: Wednesday 12 March 2008 Chapter 14: More on Splicing! Role of RNA PolII CTD How are multiple introns spliced? Alternative splicing Self-splicing introns Group I introns Group II introns
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14-2 Role of the RNA Polymerase II CTD C-terminal domain of the Rpb1 subunit of RNA polymerase II stimulates splicing of substrates that use exon definition This does not apply to those that use intron definition to prepare for splicing CTD binds to splicing factors and could assemble the factors at the end of exons to set them off for splicing
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14-3 Fig. 14.35 Experiment: P-32 labeled pre-mRNA was incubated with a splicing extract containing GST (Glutathione-S-Transferase, Control) or CTD (RNA PolII C- terminal domain linked to GST). Wedges indicate increased time. Conclusion: CTD stimulates splicing 3 - 5 fold (Compared with GST alone). Do you notice anything strange about these results?
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14-4 Fig. 14.36 Experiment: in vitro splicing reactions; two substrates on left have complete exons. Substrate on right has an incomplete exon. Note that CTD stimulates splicing only of the substrates with complete exons. Therefore CTD stimulates splicing by EXON DEFINITION (if the complete exon is not there, it cannot be defined.)
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14-5 Fig. 14.37
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Are introns removed in a specific order from a particular RNA? (e.g., from one end of the mRNA to the other?) Ovomucoid precursor RNA: 5.5 kb, 7 introns. How many possible pathways to mature mRNA are there? Northern blotting of nuclear RNA gives predominant bands suggesting that introns are removed in the following order: 5 and 6; then 4 and 7; then 2; then 1 and 3.
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Gen511-3-12-08F - Gen511: Wednesday 12 March 2008 Chapter...

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