Plant-specific SR-related protein atSR45a interacts with spliceosomal proteins in plant nucleus.
Serine/arginine-rich (SR) protein and its homologues (SR-related proteins) are important regulators of constitutive and/or alternative splicing and other aspects of mRNA metabolism. To clarify the contribution of a plant-specific and stress-responsive SR-related protein, atSR45a, to splicing events, here we analyzed the interaction of atSR45a with the other splicing factors by conducting a yeast two-hybrid assay and a bimolecular fluorescence complementation analysis. The atSR45a-1a and -2 proteins, the presumed mature forms produced by alternative splicing of atSR45a, interacted with U1-70K and U2AF(35)b, splicing factors for the initial definition of 5' and 3' splice sites, respectively, in the early stage of spliceosome assembly. Both proteins also interacted with themselves, other SR proteins (atSR45 and atSCL28), and PRP38-like protein, a homologue of the splicing factor essential for cleavage of the 5' splice site. The mapping of deletion mutants of atSR45a proteins revealed that the C-terminal arginine/serine-rich (RS) domain of atSR45a proteins are required for the interaction with U1-70K, U2AF(35)b, atSR45, atSCL28, PRP38-like protein, and themselves, and the N-terminal RS domain enhances the interaction efficiency. Interestingly, the distinctive N-terminal extension in atSR45a-1a protein, but not atSR45a-2 protein, inhibited the interaction with these splicing factors. These findings suggest that the atSR45a proteins help to form the bridge between 5' and 3' splice sites in the spliceosome assembly and the efficiency of spliceosome formation is affected by the expression ratio of atSR45a-1a and atSR45a-2.
Tanabe
Noriaki
N
Department of Advanced Bioscience, Faculty of Agriculture, Kinki University, Nakamachi, Nara, Japan.
Kimura
Ayako
A
Yoshimura
Kazuya
K
Shigeoka
Shigeru
S
D017398:Alternative Splicing; D017360:Arabidopsis / Q000235:genetics / Q000378:metabolism*; D029681:Arabidopsis Proteins / Q000235:genetics / Q000378:metabolism*; D002352:Carrier Proteins / Q000235:genetics / Q000378:metabolism*; D002467:Cell Nucleus / Q000235:genetics / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D015723:Gene Library; D011485:Protein Binding; D022821:RNA Splice Sites; D012333:RNA, Messenger / Q000378:metabolism; D018749:RNA, Plant / Q000378:metabolism; D016601:RNA-Binding Proteins; D017381:Spliceosomes / Q000235:genetics / Q000378:metabolism*; D000072668:Splicing Factor U2AF; D020798:Two-Hybrid System Techniques
D029681:Arabidopsis Proteins; D002352:Carrier Proteins; D004268:DNA-Binding Proteins; D022821:RNA Splice Sites; D012333:RNA, Messenger; D018749:RNA, Plant; D016601:RNA-Binding Proteins; C483303:SR45 protein, Arabidopsis; D000072668:Splicing Factor U2AF; C000591218:U2AF(35)b protein, Arabidopsis