Mechanism research of PRMT5 regulating m6A modification to mediate sensitivity of triple-negative breast cancer cells to doxorubicin

FAN Cong; WANG Jiajia; WANG Ting; ZHANG Jian

doi:10.13276/j.issn.2097-1656.2024.07.011

Abstract：Objective To explore the molecular mechanism of protein arginine methyltransferases 5(PRMT5)reducing the sensitivity of triple-negative breast cancer cells to doxorubicin(DOX)by regulating RNA m6A modification.Methods The TCGA database was used to analyze the expression level of PRMT5 in breast cancer tissues and its correlation with prognosis of patients.Human breast cancer MDA-MB-231 cell lines with stable overexpression and knockdown of PRMT5 were constructed.The effect of PRMT5 on DOX sensitivity of breast cancer cells was analyzed by CCK-8 and colony formation assay.The effect of PRMT5 on the expression level of m6A-related molecules in MDA-MB-231 cells under DOX was detected by qRT-PCR and Western blotting.MeRIP-seq was used to analyze the target genes with significant differences in m6A modification under the regulation of DOX by PRMT5,and the expression level and RNA stability of the target genes were detected through qRT-PCR.Results The results of TCGA database analysis showed that PRMT5 was highly expressed in breast cancer and other various tumors(P＜0.05),and high expression of PRMT5 predicted poor prognosis.Overexpression of PRMT5 reduced the sensitivity of MDA-MB-231 cells to DOX(P＜0.01),while knockdown of PRMT5 or using PRMT5 inhibitor JNJ significantly increased it(P＜0.01,P＜0.05).Overexpression of PRMT5 notably inhibited the increase of m6A levels in cells induced by DOX(P＜0.01),particularly reduced the RNA methylation levels of molecules associated with DNA damage,such as ERCC4 and REV3L,and promoted RNA stability of these molecules.Conclusion PRMT5 reduces RNA methylation level of molecules related to DNA damage repair in MDA-MB-231 cells to increase its stability,thereby reducing the cell sensitivity to DOX.