CARM1-mediated methylation of protein arginine methyltransferase 5 represses human γ-globin gene expression in erythroleukemia cells

Abstract

Protein arginine methyltransferase 5 (PRMT5) is a member of the arginine methyltransferase protein family that critically mediates the symmetric dimethylation of Arg-3 at histone H4 (H4R3me2s) and is involved in many key cellular processes, including hematopoiesis. However, the post-translational modifications (PTMs) of PRMT5 that may affect its biological functions remain less well-understood. In this study, using MS analyses, we found that PRMT5 itself is methylated in human erythroleukemia Lys-562 cells. Biochemical assays revealed that coactivator-associated arginine methyltransferase 1 (CARM1) interacts directly with and methylates PRMT5 at Arg-505 both in vivo and in vitro. Substitutions at Arg-505 significantly reduced PRMT5's methyltransferase activity, decreased H4R3me2s enrichment at the ␥-globin gene promoter, and increased the expression of the ␥-globin gene in Lys-562 cells. Moreover, CARM1 knockdown consistently reduced PRMT5 activity and activated ␥-globin gene expression. Importantly, we show that CARM1-mediated methylation of PRMT5 is essential for the intracellular homodimerization of PRMT5 to its active form. These results thus reveal a critical PTM of PRMT5 that represses human ␥-globin gene expression. We conclude that CARM1-mediated asymmetric methylation of PRMT5 is critical for its dimerization and methyltransferase activity leading to the repression of ␥-globin expression. Given PRMT5's crucial role in diverse cellular processes, these findings may inform strategies for manipulating its methyltransferase activity for managing hemoglobinopathy or cancer.Post-translational modifications (PTMs) 3 of histone proteins play important roles in defining chromatin structure and controlling gene activities such as globin gene expression (1-6). Among the various modifications, arginine methylation is particularly critical for several cellular processes, including signal transduction, DNA repair, transcription, protein subcellular localization, and RNA processing (7,8). In eukaryotes, arginine methylation is catalyzed by a family of enzymes called protein arginine methyltransferases (PRMTs). In humans, this family currently consists of nine members subdivided into three categories based on differences in primary sequences and substrate specificity: Type I PRMTs, which catalyze monomethyl arginine formation and the asymmetric dimethylation (me2a) of arginine residues (PRMT1, 2, 3, 4, 6, and 8); type II PRMTs, which catalyze monomethyl arginine formation and the symmetric dimethylation (me2s) of arginine residues (PRMT5 and 9); and type III PRMTs, which catalyze only the monomethyla

PRMT5 function and targeting in cancer

“…LKB1, a kinase with tumor suppressor function, phosphorylates multiple threonines (T132, 139 and 144) in the PRMT5 TIM-barrel domains required for MEP50, pICln and RIOK1 interaction, suppressing PRMT5 enzymatic activity [118]. PRMT5 itself is arginine-methylated by CARM1 (PRMT4) in the erythroleukemia cells Lys-562, which is essential for PRMT5 methyltransferase activity to repress human γ-globin expression via H4R3me2s [119]. Moreover, PRMT5 adaptors are subject to modifications that impact overall PRMT5 activity.…”

Fine-Tuning of GLI Activity through Arginine Methylation: Its Mechanisms and Function

“…In contrast, JAK2V617F [ 42 ], a constitutive active mutant of Janus kinase 2 (JAK2) that is found in patients with myeloproliferative neoplasms, phosphorylates PRMT5 and downregulates its activity [ 43 ]. Furthermore, another member of the PRMT family, PRMT4, methylates PRMT5 and downregulates its activity [ 44 ]. It has also been reported that casein kinase 1 (CK1)-mediated PRMT1 phosphorylation regulates its specificity of chromatin binding [ 45 ], and that p38 mitogen-activated kinase (MAPK)-mediated PRMT4 phosphorylation regulates its localization [ 46 ].…”

“…Because the roles of these additional methylations have not yet been defined, it would be interesting to understand the roles of other arginine residues that are methylated by PRMT5. Recently, it was reported that PRMT4 regulates PRMT5 activity [ 44 ]. Therefore, PRMT4 may be also involved in the regulation of GLI1 activity.…”

Fine-Tuning of GLI Activity Through Arginine Methylation: Its Mechanisms and Function

“…In contrast, JAK2V617F [41], a constitutive active mutant of Janus kinase 2 (JAK2) that is found in patients with myeloproliferative neoplasms, phosphorylates PRMT5 and downregulates its activity [42]. Furthermore, another member of the PRMT family, PRMT4, methylates PRMT5 and downregulates its activity [43]. It has also been reported that casein kinase 1 (CK1)-mediated PRMT1 methylation regulates its specificity of chromatin binding, and that p38 mitogen-activated kinase (MAPK)-mediated PRMT4 phosphorylation regulates its localization.…”

“…Because the roles of these additional methylations have not yet been defined, it would be interesting to understand the roles of other arginine residues that are methylated by PRMT5. Recently, it was reported that PRMT4 regulates PRMT5 activity [43]. Therefore, PRMT4 may be also involved in the regulation of GLI1 activity.…”