Leicester Institute of Structural & Chemical Biology and Department of Molecular & Cell Biology, University of Leicester, Leicester LE1 7RH, UK.
College of Pharmacy and College of Pharmacy and BIO5 Institute, University of Arizona, Tucson, AZ 85721, USA.
Nucleic Acids Res. 2018 Jan 25;46(2):886-896. doi: 10.1093/nar/gkx1122.
Sequences with the potential to form RNA G-quadruplexes (G4s) are common in mammalian introns, especially in the proximity of the 5' splice site (5'SS). However, the difficulty of demonstrating that G4s form in pre-mRNA in functional conditions has meant that little is known about their effects or mechanisms of action. We have shown previously that two G4s form in Bcl-X pre-mRNA, one close to each of the two alternative 5'SS. If these G4s affect splicing but are in competition with other RNA structures or RNA binding proteins, then ligands that stabilize them would increase the proportion of Bcl-X pre-mRNA molecules in which either or both G4s had formed, shifting Bcl-X splicing. We show here that a restricted set of G4 ligands do affect splicing, that their activity and specificity are strongly dependent on their structures and that they act independently at the two splice sites. One of the ligands, the ellipticine GQC-05, antagonizes the major 5'SS that expresses the anti-apoptotic isoform of Bcl-X and activates the alternative 5'SS that expresses a pro-apoptotic isoform. We propose mechanisms that would account for these see-saw effects and suggest that these effects contribute to the ability of GQC-05 to induce apoptosis.
具有形成 RNA G-四链体（G4s）潜力的序列在哺乳动物内含子中很常见，特别是在 5'剪接位点（5'SS）附近。然而，证明 G4s 在功能性条件下形成于前体 mRNA 中的困难意味着人们对它们的作用或作用机制知之甚少。我们之前已经表明，两个 G4s 形成在 Bcl-X 前体 mRNA 中，一个靠近两个替代 5'SS 中的每一个。如果这些 G4s 影响剪接，但与其他 RNA 结构或 RNA 结合蛋白竞争，那么稳定它们的配体将增加形成任一个或两个 G4s 的 Bcl-X 前体 mRNA 分子的比例，从而改变 Bcl-X 剪接。我们在这里表明，一组受限的 G4 配体确实会影响剪接，它们的活性和特异性强烈依赖于它们的结构，并且它们在两个剪接位点独立作用。其中一种配体，椭圆碱 GQC-05，拮抗表达抗凋亡 Bcl-X 异构体的主要 5'SS，并激活表达促凋亡异构体的替代 5'SS。我们提出了可以解释这些跷跷板效应的机制，并表明这些效应有助于 GQC-05 诱导细胞凋亡的能力。
