PIWIL2对TBCB及下游肿瘤细胞微管聚合的调控作用及分子机制的研究

PIWIL2, one member of Piwi/Ago proteins, belongs to a group of cancer/testis antigens and plays multiple functions in cellular process. Previous studies including ours have shown that PIWIL2 plays essential roles in the proliferation and apoptosis of tumor cells. However, the molecular mechanism underlying its functions in tumor cells remains largely unclear. Our recent work have shown that PIWIL2 is involved in regulating the cytoskeleton system, which is known to be highly associated with transformation, proliferation, cell cycle, migration and invasion ability of tumor cells. But the regulating mechanism that leads to the abnormity of the cytoskeleton, especially the microtubule in tumor cells requires further investigation. Only recently, we revealed that the interaction of PIWIL2 and tubulin folding cofactor B (TBCB) is involved in microtubule dynamics. To further investigate these underlying molecular mechanism in this study, a series of experiments such as immunoprecipitation, western blot, confocal-immunofluorescence and etc will be performed, and some specific cell lines and animal models will be constructed. We aim to clarify the mechanism that how PIWIL2 and its variant PL2L60 regulate TBCB and microtubule dynamics, which is involved in tumor cell proliferation and metastasis. This study will reveal some novel signaling pathways in which PIWIL2 is involved in tumor cells, providing new insights into the function of PIWIL2 and TBCB in the proliferation and metastasis of tumor cells.

PIWIL2是一种新发现的多功能癌/睾丸蛋白，包括本课题组的已有研究显示PIWIL2在肿瘤细胞增殖、凋亡中具有重要的调控作用，但其在肿瘤细胞中的作用仍有待进一步阐明。近期我们发现PIWIL2能够对肿瘤细胞的细胞骨架系统进行调控。现有研究显示，细胞骨架与肿瘤细胞的恶性转化、细胞增殖、细胞周期和迁移侵袭密切相关，但对于肿瘤细胞中细胞骨架，尤其是微管系统异常变化的机理尚不清楚。我们前期初步研究发现，PIWIL2可以通过与TBCB作用对肿瘤细胞微管聚合/解聚产生影响。我们拟在此基础上进一步深入研究并阐明肿瘤细胞中PIWIL2及其剪接异构体PL2L60对TBCB的调控作用，并进一步影响TBCB下游微管聚合/解聚的分子机制，阐明PIWIL2通过调控TBCB及下游微管聚合调节肿瘤细胞增殖和迁移侵袭等方面新的功能机理。

人PIWIL2蛋白在生殖细胞增殖、分化、以及转座子调控上具有非常重要的作用，而最近的研究发现PIWIL2在肿瘤细胞中也广泛表达。而TBCB是一种广泛存在于细胞中的微管蛋白折叠辅助因子，能够促进α-微管蛋白的折叠，有利于α/β微管蛋白异二聚体的形成。在本课题研究中，我们首次报道了PIWIL2在肿瘤细胞中与TBCB存在相互作用。PIWIL2通过与TBCB及HSP90结合，促进TBCB与HSP90的结合，进而抑制E3泛素连接酶GAN介导的TBCB泛素化和PAK1介导的TBCB蛋白磷酸化水平。通过对TBCB泛素化及磷酸化的调控，PIWIL2能够抑制肿瘤细胞中的微管聚合进程，进而增加肿瘤细胞的细胞增殖、迁移和侵袭水平。此外，PIWIL2在肿瘤细胞中通过激活PI3K-AKT途径，抑制GSK3β介导的β-catenin磷酸化和泛素化降解。β-catenin的稳定与积聚入核能够启动CCND1的转录，最终对肿瘤细胞的细胞周期和细胞增殖产生调控。同时，PIWIL2能够抑制SIAH2介导的HDAC3泛素化降解；并促进CK2α介导的HDAC3磷酸化。通过调控HDAC3蛋白的稳定性和HDAC酶活性，PIWIL2能够降低P53基因启动子区域的组蛋白乙酰化水平，从而抑制P53-P21途径以促进肿瘤细胞增殖并抑制其凋亡水平。上述结果初步阐明了PIWIL2通过TBCB调控肿瘤细胞增殖和迁移侵袭等方面新的功能机理。这将有助于进一步认识PIWIL2在肿瘤细胞中新的调控功能，为临床医学和肿瘤治疗上的应用研究提供新的理论依据。