Presentation Authors: Haiwei Yang*, Shuhui Si, Hao Yu, Jun Tao, Jie Han, Jingzi Wang, Xiao Yang, Qiang Lu, Zengjun Wang, Nanjing, China, People's Republic of
Introduction: N6-methyladenosine (m6A) modification of mRNA plays a role in regulating tumor initiation and progression through various mechanisms. ALKBH5, described as m6A demethylases, was reported to mediate the hypoxia-induced and hypoxia-inducible factor (HIF)-dependent breast CSC phenotype, suggesting an important tumorigenic role. Among all kidney cancers, clear cell renal cell carcinoma (ccRCC) is the most common with a molecularly distinct phenotype and contain up to 91% nonfunctional VHL mutations, which may leads to the constitutive expression of HIF-1Î± and HIF-2Î±.
Methods: The expression of ALKBH5 was measured by western blotting, qRT-PCR and immunohistochemistry. The bioprocesses of RCC cells affected by ALKBH5 were demonstrated by a series of experiments in vitro and in vivo. RNA immunoprecipitation and luciferase reporter assay were conducted to identify the potential mechanisms of ALKBH5 in RCC.
Results: In the present study, we found that ALKBH5 is up-regulated in RCC tissues and cell lines. RCC patients with high ALKBH5 expression had shortened survival. Enforced- expression of ALKBH5 promoted cells proliferation, migration, invasion and metastasis both in vitro and in vivo. Furthermore, we demonstrated that ALKBH5 up-regulates the expression of AURKB and stabilized AURKB transcript in m6A manner to enhance AURKB expression via binding to 3â€²-UTR of AURKB transcript, which promote the aggressive biological behaviors of RCC. Additionally, specific inhibitors of AURKB activity and small interfering RNA (siRNA) of AURKB expression inhibited ALKBH5-mediated promotion of proliferation. Finally, we found hypoxia induced HIF-dependent ALKBH5 expression when RCC cells exposed in 1% O2, and decreased m6A methylation of AURKB mRNA.
Conclusions: In summary, our study tested the hypothesis that exposure of RCC cells to hypoxia is sufficient to stimulate HIF-dependent ALKBH5 expression, leading to decreased m6A methylation of AURKB mRNA, increased AURKB mRNA stability and RCC cells proliferation, thereby establishing hypoxia as a major physiological stimulus in the tumor microenvironment that directly connects regulation of m6A RNA methylation to the proliferation of RCC.