长非编码RNA HULC通过调控PKM2促进肿瘤细胞增殖的分子机制研究

Increasing reports have revealed that long noncoding RNAs (lncRNAs) play important roles in a large spectrum of biological processes, including cell proliferation and differentiation. LncRNA HULC has been reported to be up-regulated in liver cancer and implicated in the regulation of tumor proliferation. However, the detailed molecular mechanism is still largely unknown. In our preliminary study, a systematic screening of the HULC interacting proteins was conducted by using a SILAC labeling based quantitative proteomics method. The results showed that HULC could bind with pyruvate kinase M2 (PKM2). PKM2 is a key glycolytic regulator. It has also been suggested to translocate to the cell nucleus and activate several transcriptional factors. Thus, we hypothesize that HULC may interact with PKM2 and regulate its enzyme activity to modulate glycolysis; furthermore, HULC may also affect gene transcription through PKM2. To test this hypothesis, we will investigate the interaction between HULC and PKM2 and further explore the mechanism whereby HULC regulates glycolysis and cell proliferation.

长非编码RNA(lncRNA)参与调控包括细胞增殖分化在内的许多重要基本生命过程。其中lncRNA HULC在肝癌中高表达并促进肿瘤细胞增殖，然而其具体分子机制还不清楚。我们在前期工作中通过SILAC定量蛋白质组学技术对HULC相互作用蛋白进行了系统性筛查，发现HULC与丙酮酸激酶PKM2存在相互作用。PKM2是调控肿瘤细胞有氧糖酵解的关键分子，同时通过调控基因转录影响肿瘤代谢、增殖和转移。我们推测HULC可能通过直接调控PKM2的聚合影响其酶活性从而作用于肝癌细胞有氧糖酵解代谢过程，同时HULC也可能通过PKM2影响其下游基因的表达进一步影响肿瘤细胞增殖。本项目拟深入探索HULC与PKM2的相互作用与调控机制，进一步阐明HULC调控有氧糖酵解的生理功能和分子调控机制，以及HULC调控PKM2下游基因表达的功能和机制，为解析lncRNA分子在调控肿瘤代谢和增殖中的新功能与相关机理奠定基础。

长非编码 RNA（lncRNA）与蛋白质相互作用是 lncRNA发挥生理作用的重要途径之一。本研究中，我们建立了一个新的提取lncRNA相互作用蛋白的实验方法TOBAP，结合SILAC稳定同位素标记、定量蛋白质组学分析和生物信息学分析，在此基础上构建了HULC的相互作用蛋白网络。研究发现，HULC直接结合LDHA和PKM2，并且HULC通过影响LDHA和PKM2的磷酸化影响LDH和PK的酶活性。HULC可能作为一个衔接分子，促进FGFR1与LDHA、PKM2的结合，进而使LDHA和PKM2的磷酸化水平升高。HULC可能通过平衡糖酵解和氧化磷酸化水平来调控肝癌细胞增殖。总之，本研究建立了一种研究细胞内 lncRNA 相互作用蛋白的简便方法，并阐述了 HULC 通过调控糖酵解关键酶活性、肿瘤细胞糖酵解水平，从而促进肝癌细胞增殖的分子机制。