Ajiro Masahiko, Zheng Zhi-Ming
Tumor Virus RNA Biology Section, Gene Regulation and Chromosome Biology Laboratory, National Cancer Institute, National Institutes of Health, Frederick, MD 21702 USA.
Cell Biosci. 2015 Dec 21;5:70. doi: 10.1186/s13578-015-0061-7. eCollection 2015.
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.
黑色素瘤相关BRAF激酶对其小分子抑制剂维莫非尼产生耐药性的一种机制是其内含子8发生点突变，导致外显子4 - 8跳跃。在本报告中，我们进行了体外BRAF RNA剪接试验和套索RT-PCR，以定位野生型和BRAF突变体中内含子8的分支点。我们在野生型（wt）和维莫非尼耐药BRAF RNA的内含子8中鉴定出多个分支点（BP）。在wt BRAF中，分支点位于 - 29A、 - 28A和 - 26A，而在一个维莫非尼耐药的BRAF剪接突变体中，分支点定位到靠近内含子8 3'剪接位点的 - 22A、 - 18A和 - 15A。这一关于BRAF剪接中分支点序列因内含子8中的点突变而发生从远端到近端移位的发现，为维莫非尼耐药时BRAF可变剪接的调控提供了见解。
