Quantitative Label‐Free Surface‐Enhanced Raman Spectroscopic Transcriptome‐Wide Profiling of RNA Adenosine Methylation

RNA N6‐methyladenosine (m6A) modification plays critical roles in diverse biological processes and human diseases, but its functional studies are greatly impeded by the inability to quantify the m6A methylation in whole‐transcriptomes. Here, An integrated label‐free surface‐enhanced Raman spectroscopy (SERS) profiling strategy, named m6A‐SERS‐profiler, is developed for quantitative transcriptome‐wide m6A detection of cellular and serum RNA. The m6A‐SERS‐profiler is rationally designed through coupling the plasmonic liquid microparticle‐based label‐free SERS biosensing with the custom deep learning‐assisted spectral analysis of m6A methylation signatures. With this m6A‐SERS‐profiler, direct identification of SERS spectral signatures of canonical nucleoside monophosphates and N6‐methyladenosine 5′‐monophosphate is achieved. The applicability of the m6A‐SERS‐profiler is verified by quantifying the m6A methylation status of microRNA and natural RNA in total RNA isolated from cancer cells and clinical serums. The results reveal a significant difference in the m6A RNA methylation among breast cancer cells of different subtypes and between clinical serums from healthy individuals and breast cancer patients. Moreover, the method can quantitatively track the m6A dynamics in cancer cells caused by pharmacological inhibition of the m6A methyltransferase. Thus, this m6A‐SERS‐profiler holds great promise for transcriptome‐wide quantitative detection of m6A RNA methylation to investigate biological functions and dynamics of m6A methylation in human diseases.