Effect of Protein Arginine Methyltransferase 1 Gene Knockout on the Proliferation of Human Embryonic Kidney 293T Cells

• Published in: Biology bulletin of the Russian Academy of Sciences Vol. 49; no. Suppl 2; pp. S1 - S11
• Main Authors: Mei-Lin Zhou, Ma, Jin-Ni, Xue, Lu
• Format: Journal Article
• Language: English
• Published: Moscow Pleiades Publishing 01.12.2022; Springer Nature B.V
• Subjects: Apoptosis; Biochemistry; Biomedical and Life Sciences; c-Myc protein; Caspase-8; Cell Biology; Cell cycle; Cell lines; Cell proliferation; CRISPR; DNA sequencing; Ecology; Flow cytometry; Gene flow; Growth rate; Immunoblotting; Life Sciences; Myc protein; Protein arginine methyltransferase; Proteins; Western blotting; Zoology; caspase-8; CRISPR/Cas9; proliferation; PRMT1; cell cycle; c-Myc; apoptosis
• ISSN: 1062-3590; 1608-3059
• DOI: 10.1134/S1062359022140163

To construct a protein arginine methyltransferase 1 (PRMT1) knockout human embryonic kidney cell line 293T (HEK293T/293T) and elucidate the effect on the proliferation of 293T cells, we generated a PRMT1 protein knockout 293T cell line with the clustered regularly interspaced short palindromic repeats/CRISPR-associated nuclease9 (CRISPR/Cas9) gene editing and performed flow cytometry. Single edited 293T cells were screened by DNA sequencing and protein immunoblotting. Furthermore, the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) and colony formation assays were performed to detect the effect of PRMT1 knockout on the proliferation of 293T cells, and the effects on 293T cell cycle distribution and apoptosis after PRMT1 knockout were analyzed by flow cytometry. The expression levels of c-Myc, a proliferation-related protein, and caspase-8, which classically triggers the extrinsic apoptotic pathway, in the wild-type (WT) 293T cell line and PRMT1 knockout cell lines were detected via western blotting. The cellular experimental results showed that knockout of PRMT1 inhibited the growth rate and clonal formation ability of 293T cells. In addition, 293T cells were arrested at the G2/M phase, and the proportion of apoptotic cells was increased as a result of PRMT1 depletion. Moreover, the expression levels of c-Myc and caspase-8 in the PRMT1 knockout cells were decreased compared with those in the WT 293T cells. Conclusion: Knockout of PRMT1 repressed the proliferation of 293T cells, more specifically, altering the distribution of cell cycle phases in 293T cells and promoting cell apoptosis of 293T cells.