期刊
RNA
卷 13, 期 8, 页码 1287-1300出版社
COLD SPRING HARBOR LAB PRESS, PUBLICATIONS DEPT
DOI: 10.1261/rna.403607
关键词
sRp30c; hnRNP I/ PTB; 39 splice site selection; alternative splicing; anti-repression
The control of alternative pre-mRNA splicing often requires the participation of factors displaying synergistic or antagonistic activities. In the hnRNP A1 pre- mRNA, three elements promote the exclusion of alternative exon 7B, while a fourth intron element (CE9) represses splicing of exon 7B to the downstream exon. We have shown previously that the 59 portion of the 38-nucleotide-long CE9 element is bound by SRp30c, and that this interaction is important for repression in vitro. To determine whether SRp30c alone can impose repression, we tested a high-affinity SRp30c binding site that we identified using the SELEX protocol. We find that multiple high-affinity SRp30c sites are required to replicate the level of repression obtained with CE9, and that both the 59 and the 39 portions of CE9 contribute to SRp30c binding. Performing RNA affinity chromatography with the complete CE9 element recovered hnRNP I/PTB. Surprisingly however, His-tagged PTB reduced the binding of SRp30c to CE9 in a nuclear extract, stimulated splicing to a downstream 3' splice site, and relieved the CE9-mediated splicing repression in vitro. Our in vivo results are consistent with the notion that increasing PTB levels alleviates the repression imposed by CE9 to a downstream 3' splice site. Thus, PTB can function as an anti-repressor molecule to counteract the splicing inhibitory activity of SRp30c.
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