VDAC在人纤溶酶原K5诱导宫颈癌细胞和内皮细胞凋亡中的关键作用及机制

We have previously shown that K5 induced endothelial cell apoptosis via activating the mitochondrial apoptotic pathway. Recently, we have also found K5 directly induced cervical cancer cells Hela apoptosis, however the mechanism was not certain. VDAC is the hinge molecule in mitochondria-mediated apoptosis. Our preliminary data showed K5 up-regulated VDAC, suggestting that K5 may induced endothelial cells and cancer cells apoptosis both through VDAC activation. This study aims to clarify: ① How did K5 up-regulate VDAC? We had found that K5 actuated GSK3β activity, which was the upstream kinase of VDAC. We will identify whether K5 promote the phosphorylation of VDAC and aviod the VDAC ubiquitination to ascend its level. ② How did K5 induce apoptosis when VDAC expression was increased? It is known that all the phosphorylation form of VDAC could not bind to HK, but could bind to Bax. We will study whether K5 induced apoptosis via promoting VDAC's phosphorylation, reducing the incorporation of VDAC-HK and increasing the combination of VDAC-Bax. ③ It is clear that VDAC is a receptor for K5, our results showed that K5 facilitated the translocation of VDAC to plasma membrane. We will discuss whether there have a positive feedback regulation loop, by which enlarge the effect of K5. This study will demonstrate the mechanism and key molecules of K5-inhibited the growth of cervical cancer.

我们以往研究表明：K5激活线粒体通路诱导内皮细胞凋亡；最近又发现K5可直接诱导宫颈癌HeLa细胞凋亡；但机制不明。VDAC是调控线粒体凋亡的关键分子；预实验显示K5上调内皮细胞和HeLa细胞VDAC，提示K5可能通过调控VDAC诱导两种细胞凋亡。本项目拟探讨其机制：① K5如何上调VDAC？预实验显示K5可激活VDAC上游激酶GSK3β的活性，本项目拟明确K5是否促进VDAC磷酸化，避免其泛素化降解上调VDAC；②VDAC上调后如何诱导细胞凋亡？已知VDAC磷酸化后不能与HK结合，但可与Bax结合。本项目拟明确K5可否促进VDAC磷酸化，减少VDAC-HK、增加VDAC-Bax结合，诱导细胞凋亡；③已知VDAC是K5的受体，预实验显示K5可促进VDAC转位到细胞膜上，本项目拟探讨是否存在配体（K5）和受体（VDAC）正反馈调节环路放大K5的作用。旨在阐明K5抑制宫颈癌生长的机制和关键分子。

本项目对新生血管抑制分子K5诱导内皮细胞凋亡的作用及机制进行了较为深入的研究。① K5诱导内皮细胞凋亡的信号通路：发现K5通过调节Bak/Bcl-xL在线粒体和胞浆中的分布，降低线粒体膜电位，促进Cyt c释放，激活线粒体途径诱导内皮细胞凋亡；②明确VDAC1在K5诱导内皮细胞凋亡中的作用：阐明VDAC1同时作为受体和线粒体膜通道蛋白在K5激活线粒体通路的关键作用；③ K5对VDAC1含量及线粒体和细胞膜转位的调节：发现K5促进VDAC1磷酸化抑制其泛素化降解从而上调VDAC1；发现K5可与其受体VDAC特异性结合并调控VDAC在内皮细胞的表达与分布，通过正反馈调节环路放大K5的凋亡诱导作用。本项目阐明了K5诱导血管内皮细胞凋亡的信号通路；明确了VDAC1在K5诱导血管内皮细胞凋亡中的关键作用和机制。为K5防治血管增生性疾病提供理论依据，同时关键分子的发现也为提供新的干预靶点提供线索。项目执行过程中已发表标注该基金资助论文15篇，其中SCI论文12篇，获得授权专利1项；项目组成员在四年中1人晋升为副主任技师（蓝秀健）、广东省高等学校“千百十工程”省级培养对象1人次（杨霞）、培养博士生3名，其中中山大学优秀研究生2名（姚亚超、李磊）、硕士生1名，获得国家奖学金（黄莉钧）。