EHMT2 (G9a) activation in mantle cell lymphoma and its associated DNA methylation and gene expression

Abstract

Objective: The function of euchromatic histone-lysine N-methyltransferase 2 (EHMT2) has been studied in several cancers; however, little is known about its role in mantle cell lymphoma (MCL). Thus, this study aimed to characterize the significance and function of EHMT2 in MCL. Methods: EHMT2 expression in MCL and reactive hyperplasia (RH) were investigated by immunohistochemistry. Genome-wide analysis of DNA methylation was performed on EHMT2 + MCL samples. The function of EHMT2 was determined by CCK8, flow cytometry, and western blot assays. Gene expression profile analysis was performed before and after EHMT2 knockdown to search for EHMT2-regulated genes. Co-immunoprecipitation (Co-IP) experiments were conducted to identify the proteins interacting with EHMT2. Results: EHMT2 was expressed in 68.57% (24/35) of MCLs but not in any RHs. Genome-wide analysis of DNA methylation on EHMT2 + MCLs revealed that multiple members of the HOX, FOX, PAX, SOX, and CDX families were hypermethylated or hypomethylated in EHMT2 + MCLs. BIX01294, a EHMT2 inhibitor, inhibited MCL cell growth and stalled cells in the G1 phase. Additionally, BIX01294 downregulated the expressions of cell cycle proteins, cyclin D1, CDK4, and P21, but upregulated the expressions of apoptosis-related proteins, Bax and caspase-3. Co-IP experiments revealed that EHMT2 interacted with UHRF1, HDAC1, and HDAC2 but not with HDCA3. After EHMT2 knockdown, multiple genes were regulated, including CD5 and CCND1, mostly enriched in the Tec kinase signaling pathway. In addition, several genes (e.g., MARCH1, CCDC50, HIP1, and WNT3) were aberrantly methylated in EHMT2 + MCLs. Conclusions: For the first time, we determined the significance of EHMT2 in MCL and identified potential EHMT2-regulated genes.