Published online 12 September 2014 Nucleic Acids Research, 2014, Vol. 42, No. 18 11349–11362 doi: 10.1093/nar/gku813 Global MEF2 target gene analysis in cardiac and skeletal muscle reveals novel regulation of DUSP6 by p38MAPK-MEF2 signaling Stephanie Wales1,2,3, Sara Hashemi1,2,3, Alexandre Blais4 and John C. McDermott1,2,3,5,*
1Department of Biology, York University, 4700 Keele Street Toronto, Ontario, M3J 1P3 Canada, 2Muscle Health Research Centre (MHRC), York University, 4700 Keele Street, Toronto, Ontario, M3J 1P3 Canada, 3Centre for Research on Biomolecular Interactions (CRBI), 4700 Keele Street, Toronto, Ontario, M3J 1P3 Canada, 4Ottawa Institute of Systems Biology, University of Ottawa, Health Sciences Campus, 451 Smyth Road, Ottawa, Ontario, K1H 8M5 Canada and 5Centre for Research in Mass Spectrometry (CRMS), York University, 4700 Keele Street, Toronto, Ontario, M3J 1P3 Canada Downloaded from
Received June 12, 2014; Revised August 23, 2014; Accepted August 28, 2014
ABSTRACT INTRODUCTION http://nar.oxfordjournals.org/ MEF2 plays a profound role in the regulation of tran- Myocyte enhancer factor-2 (MEF2) is a member of the scription in cardiac and skeletal muscle lineages. MADS-box super family of transcriptional regulatory pro- To define the overlapping and unique MEF2A ge- teins originally identified in skeletal muscle but are now nomic targets, we utilized ChIP-exo analysis of car- an established component in the regulation of a diverse diomyocytes and skeletal myoblasts. Of the 2783 and number of tissues, including smooth, cardiac and skele- tal muscle, neurons and T cells (1–4). In vertebrates 1648 MEF2A binding peaks in skeletal myoblasts and there are four MEF2 isoforms (A–D) which bind to the cardiomyocytes, respectively, 294 common binding consensus sequence (C/T TA(A/T)4TA G/A) within the sites were identified. Genomic targets were com- promoter/regulatory regions of genes to regulate gene tran- at York University Libraries on June 2, 2015 pared to differentially expressed genes in RNA-seq scription (5,6). The transcriptional activation properties of analysis of MEF2A depleted myogenic cells, reveal- MEF2 is regulated by a variety of post-translational mech- ing two prominent genetic networks. Genes largely anisms including regulation by MAPKs, such as p38 and associated with muscle development were down- ERK5 (7–9), and PKA (10), and also through interaction regulated by loss of MEF2A while up-regulated genes with class II HDACs which inhibit MEF2-dependent gene reveal a previously unrecognized function of MEF2A activation (11,12). in suppressing growth/proliferative genes. Several The transcriptional networks underlying both cardiac up-regulated (Tprg, Mctp2, Kitl, Prrx1, Dusp6)and and skeletal muscle gene expression require MEF2 dur- ing embryonic and fetal development and for post-natal down-regulated (Atp1a2, Hspb7, Tmem182, Sorbs2, control of gene expression for tissue homeostasis in adult- Lmod3) MEF2A target genes were chosen for fur- hood (13–16). During embryonic development Mef2 is ex- ther investigation. Interestingly, siRNA targeting of pressed in the somite and the presumptive vertebrate heart the MEF2A/D heterodimer revealed a somewhat di- in successive waves, beginning with Mef2c (2). This is fol- vergent role in the regulation of Dusp6, a MAPK phos- lowed shortly thereafter by Mef2a and Mef2d. MEF2A and phatase, in cardiac and skeletal myogenic lineages. MEF2C are required at different stages of the life cycle. Furthermore, MEF2D functions as a p38MAPK- Global deletion of Mef2c is embryonic lethal due to im- dependent repressor of Dusp6 in myoblasts. These paired heart morphogenesis (13) while Mef2a is necessary data illustrate that MEF2 orchestrates both common for post-natal function since gene targeting results in mito- and non-overlapping programs of signal-dependent chondrial and contractile defects in the heart (17). Mef2d gene expression in skeletal and cardiac muscle lin- homozygous null mice have no phenotypic abnormalities unless exposed to cardiac stress (18). Due to the impaired eages. development and embryonic lethality associated with Mef2 null mice, tissue-specific conditional mutant mice have been useful in fully dissecting the role of MEF2 in a plethora of tissues. Interestingly, individual skeletal muscle deletion
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