PPS, a Large Multidomain Protein, Functions with Sex-Lethal to Regulate Alternative Splicing in Drosophila 英文参考文献.docVIP

PPS,aLargeMultidomainProtein,FunctionswithSex- LethaltoRegulateAlternativeSplicinginDrosophila MatthewL.Johnson¤a,AlexisA.Nagengast¤b,HelenK.Salz* DepartmentofGenetics,CaseWesternReserveUniversity,SchoolofMedicine,Cleveland,Ohio,UnitedStatesofAmerica Abstract Alternativesplicingcontrolstheexpressionofmanygenes,includingtheDrosophilasexdeterminationgeneSex-lethal(Sxl). Sxlexpressioniscontrolledviaanegativeregulatorymechanismwhereinclusionofthetranslation-terminatingmaleexonis blocked in females. Previous studies have shown that the mechanism leading to exon skipping is autoregulatory and requires the SXL protein to antagonize exon inclusion by interacting with core spliceosomal proteins, including the U1 snRNP protein Sans-fille (SNF). In studies begun by screening for proteins that interact with SNF, we identified PPS, a previously uncharacterized protein, as a novel component of the machinery required for Sxl male exon skipping. PPS encodes a large protein with four signature motifs, PHD, BRK, TFS2M, and SPOC, typically found in proteins involved in transcription.WedemonstratethatPPShasadirectroleinSxlmaleexonskippingbyshowingfirstthatlossoffunction mutationshavephenotypesindicativeofSxlmisregulationandsecondthatthePPSproteinformsacomplexwithSXLand theunsplicedSxlRNA.Inaddition,wemappedtherecruitmentofPPS,SXL,andSNFalongtheSxlgeneusingchromatin immunoprecipitation (ChIP), which revealed that, like many other splicing factors, these proteins bind their RNA targets whileincloseproximitytotheDNA.Interestingly,whileSNFandSXLarespecificallyrecruitedtotheirpredictedbinding sites, PPS has a distinct pattern of accumulation along the Sxl gene, associating with a region that includes, but is not limitedto,theSxlPmpromoter.Together,thesedataindicatethatPPSisdifferentfromothersplicingfactorsinvolvedin male-exonskippingandsuggest,forthefirsttime,afunctionallinkbetweentranscriptionandSXL–mediatedalternative splicing.LossofzygoticPPSfunction,however,islethaltobothsexes,indicat