Study on the function of TRMT61A in liver cancer cell and its mechanism

doi:10.3969/j.issn.1674-8115.2022.06.008

Abstract

Abstract: Objective

·To investigate the effect of tRNA methyltransferase 61 homolog A (TRMT61A) on liver cancer cell function and its mechanism.

Methods

·The expression of TRMT61A in tumor and paired peri-tumor tissues of hepatocellular carcinoma patients was analyzed from TCGA database and the survival curves were plotted by using bioinformatics software. Stably-TRMT61A-knockdown Huh7 cells and HepG2 cells were established by using CRISPR-Cas9 system. TRMT61A protein levels in negative control group and knockdown group were detected by Western blotting. M¹A methylation levels in the two groups were detected by Dot blot assay. The cell proliferation of the two groups was investigated through CCK-8 assay and colony formation assay. Flow cytometry was used to analyze cell cycle after propidium iodide (PI) staining. Cell cycle-related protein level was detected by Western blotting. Cell apoptosis was detected by using Annexin V/PI kit. Cell apoptosis-related protein level was detected by Western blotting.

Results

·TRMT61A was highly expressed in hepatocellular carcinoma samples from TCGA database through bioinformatic analysis. Survival curves showed that TRMT61A was negatively correlated with patient prognosis. TRMT61A protein level and m¹A modification level were lower in TRMT61A-knockdown liver cancer cells. CCK-8 assay showed that the proliferation ability was aberrantly inhibited in Huh7 and HepG2 cells after TRMT61A knockdown. Colony formation assay showed that the number of colonies was reduced in Huh7 and HepG2 cells after TRMT61A knockdown. Mechanism investigation showed that TRMT61A-knockdown Huh7 and HepG2 cells both displayed cell cycle arrest on G0/G1 phase with elevated P21 protein level and decreased cyclin D1 protein level. Cell apoptosis rates were higher in Huh7 and HepG2 cells after TRMT61A knockdown with elevated cleaved-caspase3 protein level.

Conclusion

·TRMT61A knockdown inhibits the proliferation of liver cancer cells and induces cell apoptosis.

Key words: N1-methyladenosine (m¹A) methylation, TRMT61A, hepatocellular carcinoma, cell proliferation, cell apoptosis

CLC Number:

R735.7

HU Zhexuan, ZHANG Xin, WO Lulu, LI Jingchi, WANG Jiao, ZHOU Cixiang, ZHAO Qian. Study on the function of TRMT61A in liver cancer cell and its mechanism[J]. Journal of Shanghai Jiao Tong University (Medical Science), 2022, 42(6): 742-750.

Figures/Tables 6

Fig 1 Expression of TRMT61A in hepatocellular carcinoma samples and its correlation with prognosis

Fig 2 Establishment and confirmation of stably-TRMT61A-knockdown Huh7 and HepG2 cells

Fig 3 Effects of TRMT61A knockdown on proliferation of Huh7 and HepG2 cells

Fig 4 Effects of TRMT61A knockdown on cell cycle of Huh7 and HepG2 cells

Fig 5 Effects of TRMT61A knockdown on cell cycle-related protein levels in Huh7 and HepG2 cells

Fig 6 Effects of TRMT61A knockdown on cell apoptosis in Huh7 cells and HepG2 cells

TrendMD

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