DNMT3A Knockout Cell Line (HEK293)

Introduction:DNA methylation is an important epigenetic modification associated with transcriptional repression and plays key roles in normal cell growth as well as oncogenesis. Among the three main DNA methyltransferases (DNMT1, DNMT3A, and DNMT3B), DNMT3A mediates DNA methylation. However, the general effect of DNMT3A on cell proliferation, metabolism, and downstream gene regulation is still to be unveiled. Methods:In this study, we successfully created -deficient HEK293 cells with frameshift mutations in the catalytic domain using CRISPR/Cas9 technology. The deficient cells showed a 21.5% reduction in global DNA methylation levels, leading to impaired cell proliferation as well as a blockage of MAPK and PI3K-Akt pathways in comparison with wild-type cells. Results and Discussion:RNA-seq analysis demonstrated that knockout resulted in the up-regulation of genes and pathways related to cell metabolism but down-regulation of those involved in ribosome function, potentially explaining the growth and signaling pathways inhibition. Furthermore, DNMT3A ablation reduced gene methylation, explaining the down-regulated profiles of genes. Conclusion:Our findings suggest a complex epigenetic regulatory role for , and the compensatory upregulation of in response to deficiency warrants further investigation to be validated in future studies.