title: Vemurafenib-resistant BRAF selects alternative branch points different from its wild-type BRAF in intron 8 for RNA splicing.
abstract: One mechanism of resistance of the melanoma-associated BRAF kinase to its small molecule inhibitor vemurafenib is by point mutations in its intron 8 resulting in exons 4-8 skipping. In this report, we carried out in vitro BRAF RNA splicing assays and lariat RT-PCR to map the intron 8 branch points in wild-type and BRAF mutants. We identify multiple branch points (BP) in intron 8 of both wild-type (wt) and vemurafenib-resistant BRAF RNA. In wt BRAF, BPs are located at -29A, -28A and -26A, whereas in a vemurafenib-resistant BRAF splicing mutant, BPs map to -22A, -18A and -15A, proximal to the intron 8 3' splice site. This finding of a distal-to-proximal shift of the branch point sequence in BRAF splicing in response to point-mutations in intron 8 provides insight into the regulation of BRAF alternative splicing upon vemurafenib resistance.
authors.affiliation: Tumor Virus RNA Biology Section, Gene Regulation and Chromosome Biology Laboratory, National Cancer Institute, National Institutes of Health, Frederick, MD 21702 USA.
authors.forename: Masahiko
authors.initials: M
authors.lastname: Ajiro
authors.affiliation: Tumor Virus RNA Biology Section, Gene Regulation and Chromosome Biology Laboratory, National Cancer Institute, National Institutes of Health, Frederick, MD 21702 USA.
authors.forename: Zhi-Ming
authors.initials: ZM
authors.lastname: Zheng
keywords: BRAF; Branch point mapping; Lariat RT-PCR; RNA splicing; Vemurafenib-resistance