THE YEAST PRP3 PROTEIN IS A U4 U6 SNRNP PROTEIN NECESSARY FOR INTEGRITY OF THE U4/U6 SNRNP AND THE U4/U6.U5 TRI-SNRNP/

Previously, yeast prp3 mutants were found to be blocked prior to the f irst catalytic step of pre-mRNA splicing. No splicing intermediates or products are formed from pre-mRNA in heat-inactivated prp3 mutants or prp3 mutant extracts. Here we show that Prp3p is a component of the U 4/U6 snRNP and is also present in the U4/U6.U5 tri-snRNP. Heat inactiv ation of prp3 extracts results in depletion of free U6 snRNPs and U4/U 6.U5 tri-snRNPs, but not U4/U6 snRNPs or U5 snRNPs. Free U4 snRNP, nor mally not present in wild-type extracts, accumulates under these condi tions. Assays of in vivo levels of snRNAs in a prp3 mutant revealed th at amounts of free U6 snRNA decreased, free U4 snRNA increased, and U4 /U6 hybrids decreased slightly. These results suggest that Prp3p is re quired for formation of stable U4/U6 snRNPs and for assembly of the U4 /U6.U5 tri-snRNP from its component snRNPs. Upon inactivation of Prp3p , spliceosomes cannot assemble from prespliceosomes due to the absence of intact U4/U6.U5 tri-snRNPs. Prp3p is homologous to a human protein that is a component of U4/U6 snRNPs, exemplifying the conservation of splicing factors between yeast and metazoans.