1B-O2-1820(1P-07) PDF
Mass Spectrometric Approach for the Identification of the Splicing Regulatory Factors
Alternative splicing is a mechanism for creating proteomic diversity. This mechanism is regulated by splicing regulatory factors that bind to specific motifs located within pre-mRNA. The insulin receptor (IR) gene consists of 22 exons and exon 11 alternatively spliced to produce two isoforms. To identify the splicing regulatory factors involved in exon 11 splicing, we developed an in vitro RNA binding assay. The separated protein, which binds to the exon 11, was in-gel digested with trypsin and peptide fragments were eluted. This peptide mixture was applied to LC-MS/MS and identified the binding protein using Proteome Discoverer software. As a result, we found that hnRNPA1 (Heterogeneous nuclear ribonucleoprotein A1) protein binds to exon 11. Further investigation will be required to unravel the mechanism of alternative splicing of IR exon 11.