Targeted Profiling of Epitranscriptomic Reader, Writer, and Eraser Proteins Accompanied with Radioresistance in Breast Cancer Cells

Epitranscriptomic reader, writer,
and eraser (RWE) proteins recognize,
install, and remove modified nucleosides in RNA, which are known to
play crucial roles in RNA processing, splicing, and stability. Here,
we established a liquid chromatography-parallel-reaction monitoring
(LC-PRM) method for high-throughput profiling of a total of 152 epitranscriptomic
RWE proteins. We also applied the LC-PRM method, in conjunction with
stable isotope labeling by amino acids in cell culture (SILAC), to
quantify these proteins in two pairs of matched parental/radioresistant
breast cancer cells (i.e., MDA-MB-231 and MCF-7 cells and their corresponding
radioresistant C5 and C6 clones), with the goal of assessing the roles
of these proteins in radioresistance. We found that eight epitranscriptomic
RWE proteins were commonly altered by over 1.5-fold in the two pairs
of breast cancer cells. Among them, TRMT1 (an m2,2G writer)
may play a role in promoting breast cancer radioresistance due to
its clinical relevance and its correlation with DNA repair gene sets.
To our knowledge, this is the first report of a targeted proteomic
method for comprehensive quantifications of epitranscriptomic RWE
proteins. We envision that the LC-PRM method is applicable for studying
the roles of these proteins in the metastatic transformation of cancer
and therapeutic resistance of other types of cancer in the future.