SAP155-mediated splicing of FUSE-binding protein-interacting repressor serves as a molecular switch for c-myc gene expression.

ABSTRACT: The Far UpStream Element (FUSE)-binding protein-interacting repressor (FIR), a c-myc transcriptional suppressor, is alternatively spliced removing the transcriptional repression domain within exon 2 (FIR?exon2) in colorectal cancers. SAP155 is a subunit of the essential splicing factor 3b (SF3b) subcomplex in the spliceosome. This study aims to study the significance of the FIR-SAP155 interaction for the coordination of c-myc transcription, pre-mRNA splicing, and c-Myc protein modification, as well as to interrogate FIR?exon2 for other functions relating to altered FIR pre-mRNA splicing. Knockdown of SAP155 or FIR was used to investigate their reciprocal influence on each other and on c-myc transcription, pre-mRNA splicing, and protein expression. Pull down from HeLa cell nuclear extracts revealed the association of FIR, FIR?exon2, and SF3b subunits. FIR and FIR?exon2 were coimmunoprecipitated with SAP155. FIR and FIR?exon2 adenovirus vector (Ad-FIR and Ad-FIR?exon2, respectively) were prepared to test for their influence on c-myc expression. FIR, SAP155, SAP130, and c-myc were coordinately upregulated in human colorectal cancer. These results reveal that SAP155 and FIR/FIR?exon2 form a complex and are mutually upregulating. Ad-FIR?exon2 antagonized Ad-FIR transcriptional repression of c-myc in HeLa cells. Because FIR?exon2 still carries RRM1 and RRM2 and binding activity to FUSE, it is able to displace repression competent FIR from FUSE in electrophoretic mobility shift assays, thus thwarting FIR-mediated transcriptional repression by FUSE. Thus aberrant FIR?exon2 production in turn sustained c-Myc expression. In conclusion, altered FIR and c-myc pre-mRNA splicing, in addition to c-Myc expression by augmented FIR/FIR?exon2-SAP155 complex, potentially contribute to colorectal cancer development.

SUBMITTER: Matsushita K 

PROVIDER: S-EPMC7469880 | biostudies-literature | 2012 Jun

REPOSITORIES: biostudies-literature