KLF4 调控MGLL-Integrinβ1 轴抑制肝癌转移的分子机制研究

Metastasis is the key cause of Hepatocellular carcinoma (HCC) recurrence, thus uncovering its molecular mechanism is very important for HCC treatment. Previously, our data indicated the expression level of Monoglyceride lipase (MGLL) was downregulated in HCC. The decrease of MGLL was associated with HCC metastasis and survival time of HCC patients. However, the mechanism of molecular regulation involved by MGLL is still unclear. We identified Integrin β1 was a potential interacted protein of MGLL by IP-Mass Spectrometry and found overexpression of MGLL inhibited the stability of Integrin β1. Subsequently, we found KLF4 was a transcription factor of MGLL via promoter fishing system and Mass Spectrometry. Based on these, we will focus on the molecular mechanism by which MGLL regulates Integrinβ1 expression and confirm their relationship as well as the effect of MGLL on HCC metastasis. Meanwhile, we will try to reveal that MGLL is a new target gene of KLF4 and KLF4 inhibits cell metastasis via MGLL- Integrinβ1 axis. Finally, we will analyze the correlation of KLF4, MGLL and Integrin β1 by clinical data. Taken together, this project will provide a novel strategy for HCC treatment.

转移是肝癌术后复发的重要原因，阐明其分子机制对肝癌的治疗具有重要意义。前期研究表明单甘油酯脂肪酶（MGLL）表达水平在肝癌中显著降低，并与肝癌转移及病人的生存关系密切，但其分子机制尚未明确。我们通过质谱鉴定出MGLL潜在相互作用蛋白Integrinβ1，发现过表达MGLL降低Integrinβ1的稳定性；随后利用启动子垂钓系统结合质谱分析，寻找出调控MGLL表达的转录因子KLF4。基于此，本课题拟研究MGLL调控Integrinβ1稳定性的分子机制，明确二者的关系，阐明MGLL抑制Integrinβ1在肝癌转移中的作用；进一步揭示KLF4对MGLL的转录调控机制，明确MGLL是KLF4新的靶基因，并证实KLF4通过调控MGLL-Integrinβ1轴抑制肝癌转移；最后结合临床病例分析KLF4、MGLL 和Integrinβ1 三者的相关性及与病人预后的关系；为肝癌的诊断和治疗提供新的策略。

原发性肝癌是临床上常见的恶性肿瘤，其恶性度极高、预后极差、严重威胁国人生命健康。研究显示全世界范围内原发性肝癌的平均生存率仅有6-20个月。转移是肝癌术后复发和不良预后的重要原因，虽然有研究表明一些关键分子参与调控肝癌细胞的复发转移，但其分子机制仍不清楚。在本研究中我们发现单甘油酯脂肪酶(MGLL)在肝癌中表达水平降低， MGLL的表达与肝癌转移和病人生存期负相关。高表达MGLL 抑制肝癌细胞的转移。反之，敲低MGLL的表达促进细胞转移。为揭示MGLL在肝癌中低表达的分子机制，我们通过启动子垂钓系统证实转录因子KLF4可以直接结合在MGLL启动子区域，通过荧光素酶报告基因和染色体免疫共沉淀实验进一步确定KLF4对MGLL的调控作用。并且我们通过荧光定量PCR和western blot实验证实敲低KLF4减少MGLL mRNA和蛋白的表达水平。反之过表达促进MGLL的表达。最后通过免疫组化实验证实KLF4和MGLL在肝癌组织中表达水平呈正相关。总之，在本课题中我们揭示MGLL在肝癌转移中的功能，并证实KLF4通过转录上调MGLL的表达抑制肝癌的转移，为肝癌的临床治疗提供新的策略。