Abstract
Alternative RNA processing of the human fibroblast growth factor receptor-1 transcript results in receptor forms that vary in their affinity for fibroblast growth factor. An alternative RNA processing event involving recognition of the alpha-exon is deregulated during neoplastic transformation of glial cells. We have previously established a splicing reporter/transfection cell culture model system to identify sequences involved in recognition of this exon. In this study, the system was used to identify two sequence elements that differentially function to regulate splicing of this exon. Exclusion of the alpha-exon in glioblastoma cells specifically required the downstream intron sequence comprising the 5'-splice site. Replacement or mutation of this sequence increasing complementarity to U1 RNA resulted in enhanced exon recognition in SNB-19 glioblastoma cells. Sequences within the exon were found to be required for alpha-exon inclusion. Deletion and gain-of-function experiments identified a 69-nucleotide exon sequence that was specifically required for alpha-exon inclusion. These studies indicate that multiple sequences are required for the regulated recognition of the alpha-exon.
Highlights
We have continued to examine the mechanisms involved in alternative RNA splicing of transcripts derived from the fibroblast growth factor receptor-1 (FGFR-1)1 gene
Cell-specific Splicing of the FGFR-1 ␣-Exon Is Maintained by Minimal Sequence—Previous studies found that the exclusion of the ␣-exon from the final FGFR-1 mRNA shows a strong correlation with glial cell malignancy [12, 13]
The construct pFGFR-17 contains a 4-kilobase fragment of the FGFR-1 gene inserted into the splicing reporter RSV/hMT2 [14] (Fig. 1A)
Summary
We have continued to examine the mechanisms involved in alternative RNA splicing of transcripts derived from the fibroblast growth factor receptor-1 (FGFR-1)1 gene. Comparison of the splicing patterns for these clones suggests that the FGFR-1 sequence in pFGFR-32 contains elements required for cell-specific recognition of the ␣-exon, but additional sequences might function in the exclusion pathway. FGFR-1 Gene Sequence Requirements for ␣-Exon Skipping in SNB-19 Glioblastoma Cells—The cell-specific RNA splicing pattern of transcripts derived from pFGFR-32 suggests that this process is regulated.
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