Department of Biochemistry, The Catholic University of Korea, 222, Banpo-daero, Seocho-gu, Seoul 06591, Korea.
Institute of Aging and Metabolic Diseases, College of Medicine, The Catholic University of Korea, 222, Banpo-daero, Seocho-gu, Seoul 06591, Korea.
Cells. 2020 Oct 19;9(10):2325. doi: 10.3390/cells9102325.
BCL-2 interacting cell death suppressor (BIS), also known as BAG3, is a multifunctional protein. Aberrant expression and mutation of BIS have been implicated in cancers and myopathy. However, there have only been a few studies on the splicing of BIS pre-mRNA. In the present study, through RT-PCR and sequencing in various cell lines and mouse tissues, we identified for the first time the presence of BIS mRNA isomers in which exon 3 or exons 2-3 are skipped. We also demonstrated that the depletion of SRSF3 promoted the skipping of exon 3 of BIS pre-mRNA in endogenous BIS and the GFP-BIS minigene. SRSF3 specifically interacts with the putative binding sites in exon 3, in which deletion promoted the skipping of exon 3 in the GFP-BIS minigene, which was comparable to the effect of SRSF knockdown. Even though acceleration of exon 3 skipping was not observed in response to various stimuli, SRSF3 depletion, accompanied by the production of a truncated BIS protein, inhibited the nuclear translocation of HSF1, which was restored by the wild-type BIS, not by exon 3-depleted BIS. Therefore, our results suggested that the maintenance of SRSF3 levels and subsequent preservation of the intact BIS protein is an important factor in modulating HSF1 localization upon cellular stress.
BCL-2 相互作用细胞死亡抑制因子(BIS),也称为 BAG3,是一种多功能蛋白。BIS 的异常表达和突变与癌症和肌病有关。然而,目前关于 BIS 前体 mRNA 剪接的研究较少。在本研究中,通过在各种细胞系和小鼠组织中的 RT-PCR 和测序,我们首次鉴定出 BIS mRNA 异构体中存在外显子 3 或外显子 2-3 缺失。我们还证明,SRSF3 的耗竭促进了内源性 BIS 和 GFP-BIS 基因座中 BIS 前体 mRNA 外显子 3 的跳过。SRSF3 特异性地与外显子 3 中的假定结合位点相互作用,其中缺失促进了 GFP-BIS 基因座中外显子 3 的跳过,这与 SRSF 敲低的效果相当。尽管外显子 3 跳过的加速没有观察到对各种刺激的反应,但 SRSF3 的耗竭伴随着截断的 BIS 蛋白的产生,抑制了 HSF1 的核易位,野生型 BIS 可以恢复核易位,而不是外显子 3 缺失的 BIS。因此,我们的结果表明,SRSF3 水平的维持以及随后完整 BIS 蛋白的保留是调节细胞应激时 HSF1 定位的重要因素。