m6A-seq assays and analysis of breast cancer MDA-MB-231 cells with or without PRMT5 overexpression and Doxrubicin treatment

We report the application of immunoprecipitation of m6A methylated RNA followed by sequencing (MeRIP-Seq) in breast cancer MDA-MB-231 cells to identify PRMT5-mediated transcriptome-wide methylation patterns for doxorubicin response. Total RNAs from breast cancer cells were extracted with TRIzol. Then, mRNA sequencing and m6A sequencing were simultaneously performed. We find that doxorubicin induced the increasing m6A peaks compared to the control group, whereas inhibited by PRMT5. The transcripts exhibiting m6A methylation alteration by doxorubicin and PRMT5 were enriched for gene ontology (GO) terms related to DNA damage response and repair. The GO terms and KEGG annotations analysis identified BRCA1 signaling as being significantly altered through m6A methylation which induced by doxorubicin but inhibited by PRMT5. This study provides that PRMT5 promotes DNA damage repair through regulating m6A methylation of BRCA1 mRNA. Overall design: Examination of PRMT5-mediated transcriptome-wide methylation patterns for doxorubicin response in breast cancer MDA-MB-231 cells