Human homologs of yeast Prp16 and Prp17 reveal conservation of the mechanism for catalytic step II of pre-mRNA splicing

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Abstract
Pre‐mRNA splicing takes place in two catalytic steps. The second step is poorly understood, especially in mammals. In yeast, the splicing factors, Prps 16, 17, 18 and Slu7 function exclusively in step II. Here we report the isolation of cDNAs encoding human Prps 16 and 17 which are 41 and 36% identical to their yeast counterparts. The Prp16 gene is essential in yeast, and we show that a chimeric yeast–human Prp16 protein rescues a yeast Prp16 knockout strain. Immunodepletion of hPrp16 from splicing extracts specifically blocks step II, and the activity can be fully restored with recombinant hPrp16. Moreover, both hPrps 16 and 17 associate with the spliceosome late in the splicing pathway. Mutations at the 3′ splice site that specifically block step II do not affect the association of hPrps 16 and 17 with the spliceosome, indicating that these factors may function at a stage of step II prior to recognition of the 3′ splice site. Recently, the human homologs of Prp18 and Slu7 were identified. The observation that humans contain homologs of all four known step II proteins in yeast indicates that the mechanism for catalytic step II is highly conserved.