Antibody-Free Fluorine-Assisted Metabolic Sequencing of RNA N⁴-Acetylcytidine

N⁴-Acetylcytidine (ac⁴C) has been found to affect a variety of cellular and biological processes. For a mechanistic understanding of the roles of ac⁴C in biology and disease, we present an antibody-free, fluorine-assisted metabolic sequencing method to detect RNA ac⁴C, called "FAM-seq". We successfully applied FAM-seq to profile ac⁴C landscapes in human 293T, HeLa, and MDA cell lines in parallel with the reported acRIP-seq method. By comparison with the classic ac⁴C antibody sequencing method, we found that FAM-seq is a convenient and reliable method for transcriptome-wide mapping of ac⁴C. Because this method holds promise for detecting nascent RNA ac⁴C modifications, we further investigated the role of ac⁴C in regulating chemotherapy drug resistance in chronic myeloid leukemia. The results indicated that drug development or combination therapy could be enhanced by appreciating the key role of ac⁴C modification in cancer therapy.