title: A genetic screen implicates a CWC16/Yju2/CCDC130 protein and SMU1 in alternative splicing inArabidopsis thaliana.
abstract: To identify regulators of pre-mRNA splicing in plants, we developed a forward genetic screen based on an alternatively splicedGFPreporter gene inArabidopsis thalianaIn wild-type plants, three major splice variants issue from theGFPgene but only one represents a translatableGFPmRNA. Compared to wild-type seedlings, which exhibit an intermediate level ofGFPexpression, mutants identified in the screen feature either a "GFP-weak" or "Hyper-GFP" phenotype depending on the ratio of the three splice variants. GFP-weak mutants, including previously identifiedprp8andrtf2, contain a higher proportion of unspliced transcript or canonically spliced transcript, neither of which is translatable into GFP protein. In contrast, the coilin-deficienthyper-gfp1(hgf1) mutant displays a higher proportion of translatableGFPmRNA, which arises from enhanced splicing of a U2-type intron with noncanonical AT-AC splice sites. Here we report three newhgfmutants that are defective, respectively, in spliceosome-associated proteins SMU1, SmF, and CWC16, an Yju2/CCDC130-related protein that has not yet been described in plants. Thesmu1andcwc16mutants have substantially increased levels of translatableGFPtranscript owing to preferential splicing of the U2-type AT-AC intron, suggesting that SMU1 and CWC16 influence splice site selection inGFPpre-mRNA. Genome-wide analyses of splicing insmu1andcwc16mutants revealed a number of introns that were variably spliced from endogenous pre-mRNAs. These results indicate that SMU1 and CWC16, which are predicted to act directly prior to and during the first catalytic step of splicing, respectively, function more generally to modulate splicing patterns in plants.
authors.affiliation: Institute of Plant and Microbial Biology, Academia Sinica, Taipei 115, Taiwan.
authors.forename: Tatsuo
authors.initials: T
authors.lastname: Kanno
authors.affiliation: Institute of Plant and Microbial Biology, Academia Sinica, Taipei 115, Taiwan.
authors.forename: Wen-Dar
authors.initials: WD
authors.lastname: Lin
authors.affiliation: Institute of Plant and Microbial Biology, Academia Sinica, Taipei 115, Taiwan.
authors.forename: Jason L
authors.initials: JL
authors.lastname: Fu
authors.affiliation: Institute of Plant and Microbial Biology, Academia Sinica, Taipei 115, Taiwan.
authors.forename: Antonius J M
authors.initials: AJM
authors.lastname: Matzke
authors.affiliation: Institute of Plant and Microbial Biology, Academia Sinica, Taipei 115, Taiwan marjorimatzke@gate.sinica.edu.tw antoniusmatzke@gate.sinica.edu.tw.
authors.forename: Marjori
authors.identifier: 0000-0002-8956-4472
authors.initials: M
authors.lastname: Matzke
mesh_terms: D017398:Alternative Splicing; D017360:Arabidopsis / Q000235:genetics* / Q000378:metabolism; D029681:Arabidopsis Proteins / Q000235:genetics / Q000378:metabolism; D018506:Gene Expression Regulation, Plant; D017930:Genes, Reporter; D009154:Mutation; D009687:Nuclear Proteins / Q000235:genetics / Q000378:metabolism*; D018749:RNA, Plant / Q000235:genetics*
chemical_list: D029681:Arabidopsis Proteins; D009687:Nuclear Proteins; D018749:RNA, Plant
keywords: CWC16; SMU1; SmF; Yju2; alternative splicing