Axin-RhoBTB3-VHL复合体调控HIF1信号通路的分子机制及其生物学功能

低氧诱导因子HIF1α信号通路对肿瘤细胞的代谢与生长有着非常重要的影响。它通过正调节下游基因如GLUT1、HK、LDH、VEGF的表达，促进肿瘤的生存和发展。我们通过包括酵母双杂交系统、免疫共沉淀、细胞共定位多种方法，鉴定了Axin-RhoBTB3-VHL这个新的复合体。我们发现，RhoBTB3在肾癌样本文库中出现显著下调，而HIF1α的下游基因出现相应上调；用siRNA敲低Hela细胞的RhoBTB3, 并注射裸鼠，可以促进裸鼠移植瘤的生长；过表达RhoBTB3对于HIF1α的蛋白水平具有下调作用，而MG132处理可以抑制这种下调作用。我们拟通过免疫共沉淀、免疫荧光、实时定量PCR等实验探索低氧刺激下Axin-RhoBTB3-VHL复合体调节HIF1α蛋白水平的分子机制及其对肿瘤细胞生长和代谢的影响。

The hypoxia-inducible factor (HIF1α) signal pathway plays very important roles in tumor metabolism, which promotes tumor survival and development by regulating its downstream target genes such as GLUT1, HK, LDH, and VEGF expresstion. These target genes are cloasely related to metabolism. We have employed several methods such as yeast two-hybrid screen, co-immunoprecipitation, subcellular co-localization to identify additional complexes centered on Axin, revealing a novel complex comprised of Axin, RhoBTB3, and VHL. We found that RhoBTB3 is significantly downregulated and HIF1α signal is upregulated in kidney cancer samples in the Oncomine. Knockdown of RhoBTB3 in Hela cells promoted xenograft growth in nude mice. Moreover, overexpressed RhoBTB3 in cells could decrease protein levels of HIF1α, which could be blocked by prior MG132 treatment. We hereby propose to explore the molecular mechanism and the effect on tumor metabolism of regulation HIF1α protein levels by the Axin-RhoBTB3-VHL complex under hypoxic conditions using immunoprecipitation, immunostaining, real-time quantitative PCR and etc. Emphasis will placed on how the complex is assembled and the mechanism of the dynamic regulation of the complex. Our study will make major contributions to the understanding of the regulation of the important factor HIF1α, and the mechanism of the tumor suppression by VHL.