The Role And Mechanism Of LSD1 Methylation Mediated By PRMT4 In Breast Cancer Metastasis

Breast cancer is one of the most common tumors in women and has high morbidity and mortality worldwide.The distant metastasis was found in 90% patients dying of breast cancer,which has become one of the main causes of death.Therefore,it's important for exploring the molecular mechanism of metastasis in breast cancer treatment.Protein post-translational modifications such as phosphorylation,methylation,ubiquitination,etc.are covalent modifications catalyzed by epigenetic modifying enzymes that can occur on both histones and non-histones and involved in multiple cellular processes.Protein arginine methylation was involved in multiple cellular processes,such as signal transduction,transcriptional control,and protein stability etc.Recent studies have shown that deregulated arginine methylation is associated with tumor development.PRMT4(CARM1)wa found to dimethylate histones H3R17 and H3R26,and activate gene expression at the transcriptional level.Non-histone substrates of PRMT4 have been discovered in recent years,such as BAF155,PKM2,etc.PRMT4 is involved in a variety of cellular processes such as gene expression,transcription and mRNA processing,and protein stability regulation.It is highly expressed in breast cancer,particularly in triple negative tumors,and elevated levels of PRMT4 were associated with poor prognosis.PRMT4 plays important roles in breast cancer progression,but the mechanism remains elusive.Recent studies showed that PRMT4 promotes cancer progression and metastasis through methylation of cancer-relevant substrates,which provide us a way to explore the molecular mechanism of PRMT4 in breast cancer progression.Lysine-specific demethylase 1(LSD1),as a first identified histone demethylase,can selectively removes the methylation of histones H3K4me1/2 and H3K9me1/2 and then mediates gene repression and activating.We have also found that LSD1 regulate E-cadherin and vimentin expression via decreased H3K4me2 and H3K9me2 respectively,thereby promoting breast cancer metastasis.Overexpression of LSD1 has been found in a wide range of cancers and the high levels of LSD1 caused tumor aggressiveness and poor prognosis via promoting cancer cells migration and invasion,however,how it upregulated in tumor progression is largely unknown.As is known,protein levels are associated with its stability.Previous studies have shown that methylation of LSD1 at K322 inhibits LSD1 ubiquitination and enhances its stability;phosphorylation of LSD1 at S683 promotes LSD1 deubiquitination and enhances its stability.In this report,we discovered that LSD1 was methylated by PRMT4,which is necessary to maintain the stability of LSD1.We found that PRMT4 and LSD1 are positively correlated in human breast cancer tissues,and then we found that PRMT4 enhances LSD1 stability by inhibiting its ubiquitination.Furthermore,by mass spectrometry and in vitro methylation,we confirmed that PRMT4 mediates the methylation of LSD1 at R838,and this modification plays a role in maintaining LSD1 stability.Further studies revealed that PRMT4 methylated LSD1 enhancing the binding of LSD1 with and deubiquitination by USP7,which leads to increased stability of LSD1.The addition of PRMT4 and USP7 inhibitors significantly increased LSD1 ubiquitination,resulting in reduced stability of LSD1 in breast cancer cells.ChIP assay confirmed that inhibition of PRMT4 and USP7 significantly reduced the enrichment of LSD1 in the promoter of E-cadherin and vimentin,and as a result to repress E-cadherin and activate vimentin gene expression by attenuating H3K4me2 and H3K9me2 respectively,leading to promotion of the migration and invasion ability of cancer cells and then facilitate cancer metastasis.Finally,LSD1 methylation plays a pivotal role in promoting breast cancer metastasis in vivo.In addition,LSD1 methylation levels are correlated with LSD1 levels in human breast cancer tissues.Conclusively,data arising from this study depicts a series of molecular events in which PRMT4 methylates and stabilizes LSD1 by promoting the recruitment of USP7,leading to the repression of E-cadherin and activation of vimentin genes,respectively.Consequently,these reactions result in the promotion of migration and invasion of breast cancer cells,and facilitate cancer metastasis.We define a novel functional post-translational modification of LSD1 and demonstrate that PRMT4-and USP7-mediated LSD1 stabilization is essential for breast cancer tumorigenesis and provides us a way for potential treatment strategy against breast cancer.