title: Histone methyltransferase Setd2 is critical for the proliferation and differentiation of myoblasts.
abstract: Skeletal muscle cell proliferation and differentiation are tightly regulated. Epigenetic regulation is a major component of the regulatory mechanism governing these processes. Histone modification is part of the epigenetic code used for transcriptional regulation of chromatin through the establishment of an active or repressive state for genes involved in myogenesis in a temporal manner. Here, we uncovered the function of SET domain containing 2 (Setd2), an essential histone 3 lysine 36 trimethyltransferase, in regulating the proliferation and differentiation of myoblasts. Setd2 was silenced in the skeletal muscle myoblast cell line, C2C12, using the CRISPR/CAS9 system. The mutant cells exhibited defect in myotube formation. The myotube formation marker, myosin heavy chain (MHC), was downregulated earlier in Setd2 silenced cells compared to wild-type myoblasts during differentiation. The deficiency in Setd2 also resulted in repression of Myogenin (MyoG) expression, a key myogenic regulator during differentiation. In addition to the myoblast differentiation defect, decreased proliferation rate with significantly reduced levels of histone 3 phosphorylation, indicative of cell proliferation defect, were observed in the Setd2 silenced cells; suggesting an impaired proliferation phenotype. Furthermore, compromised G1/S- and G2/M-phase transition and decreased expression levels of major regulators of cell cycle G1/S checkpoints, cyclin D1, CDK4, CDK6, and cyclin E2 were detected in Setd2 silenced cells. Consistent with the cell cycle arrested phenotype, cyclin-dependent kinase inhibitor p21 was upregulated in Setd2 silenced cells. Together, this study demonstrates an essential role of Setd2 in myoblast proliferation and differentiation, and uncovers Setd2-mediated molecular mechanism through regulating MyoG and p21.
authors.affiliation: Department of Cardiology, Renmin Hospital of Wuhan University, Wuhan, Hubei 430060, China; Texas A&M University Health Science Center, Institute of Biosciences and Technology, Houston, TX 77030, USA.
authors.forename: Xin
authors.initials: X
authors.lastname: Yi
authors.affiliation: Texas A&M University Health Science Center, Institute of Biosciences and Technology, Houston, TX 77030, USA.
authors.forename: Ye
authors.initials: Y
authors.lastname: Tao
authors.affiliation: Department of Cardiology, Renmin Hospital of Wuhan University, Wuhan, Hubei 430060, China.
authors.forename: Xi
authors.initials: X
authors.lastname: Lin
authors.affiliation: Texas A&M University Health Science Center, Institute of Biosciences and Technology, Houston, TX 77030, USA.
authors.forename: Yuan
authors.initials: Y
authors.lastname: Dai
authors.affiliation: Texas A&M University Health Science Center, Institute of Biosciences and Technology, Houston, TX 77030, USA.
authors.forename: Tingli
authors.initials: T
authors.lastname: Yang
authors.affiliation: Department of Obstetrics and Gynecology, Nanfang Hospital, Southern Medical University, Guangzhou 510515, China.
authors.forename: Xiaojing
authors.initials: X
authors.lastname: Yue
authors.affiliation: Department of Cardiology, Renmin Hospital of Wuhan University, Wuhan, Hubei 430060, China.
authors.forename: Xuejun
authors.initials: X
authors.lastname: Jiang
authors.affiliation: Department of Cardiology, Renmin Hospital of Wuhan University, Wuhan, Hubei 430060, China.
authors.forename: Xiaoyan
authors.initials: X
authors.lastname: Li
authors.affiliation: Division of Cardiothoracic and Vascular Surgery, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430030, China.
authors.forename: Ding-Sheng
authors.initials: DS
authors.lastname: Jiang
authors.affiliation: Texas A&M University Health Science Center, Institute of Biosciences and Technology, Houston, TX 77030, USA.
authors.forename: Kelsey C
authors.initials: KC
authors.lastname: Andrade
authors.affiliation: Texas A&M University Health Science Center, Institute of Biosciences and Technology, Houston, TX 77030, USA. Electronic address: jchang@ibt.tamhsc.edu.
authors.forename: Jiang
authors.initials: J
authors.lastname: Chang
mesh_terms: D000818:Animals; D001483:Base Sequence; D064113:CRISPR-Cas Systems; D059447:Cell Cycle Checkpoints; D002454:Cell Differentiation; D002460:Cell Line; D049109:Cell Proliferation; D002843:Chromatin / Q000737:chemistry / Q000378:metabolism; D019938:Cyclin D1 / Q000235:genetics / Q000378:metabolism; D051358:Cyclin-Dependent Kinase 4 / Q000235:genetics / Q000378:metabolism; D051361:Cyclin-Dependent Kinase 6 / Q000235:genetics / Q000378:metabolism; D050759:Cyclin-Dependent Kinase Inhibitor p21 / Q000235:genetics* / Q000378:metabolism; D016213:Cyclins / Q000235:genetics / Q000378:metabolism; D000072669:Gene Editing; D020868:Gene Silencing; D011495:Histone-Lysine N-Methyltransferase / Q000172:deficiency / Q000235:genetics*; D006657:Histones / Q000235:genetics* / Q000378:metabolism; D051379:Mice; D018485:Muscle Fibers, Skeletal / Q000166:cytology / Q000378:metabolism*; D032446:Myoblasts / Q000166:cytology / Q000378:metabolism*; D018008:Myogenin / Q000235:genetics* / Q000378:metabolism; D018995:Myosin Heavy Chains / Q000235:genetics / Q000378:metabolism; D010766:Phosphorylation
chemical_list: C530105:Ccnd1 protein, mouse; C503689:Ccne2 protein, mouse; D002843:Chromatin; D050759:Cyclin-Dependent Kinase Inhibitor p21; D016213:Cyclins; D006657:Histones; C503801:Myog protein, mouse; D018008:Myogenin; D019938:Cyclin D1; D011495:Histone-Lysine N-Methyltransferase; C587921:SETD2 protein, mouse; C495902:Cdk4 protein, mouse; C495912:Cdk6 protein, mouse; D051358:Cyclin-Dependent Kinase 4; D051361:Cyclin-Dependent Kinase 6; D018995:Myosin Heavy Chains
keywords: C2C12 myoblasts; Epigenetics; H3K36me3; Proliferation and differentiation; Setd2