To test whether NAT10-promoted malignant phenotypes of cancer cells were dependent on its ac4C activity, NAT10-knockout cells were used to express wild-type NAT10 or mutant NAT10 lacking a functional acetyltransferase domain (G641E) or RNA helicase domain (K290A) (Fig. 2A, B) due to point mutations (K290A or G641E) previously shown to disrupt the RNA acetyltransferase function of NAT10 [19–21]. Here, NAT10 is linked to cancer.