Y. box superfamily (named after the first four members identified to contain this motif: MCM1, Agamous, Deficiens, and serum response factor) due to the presence of a highly conserved 59-amino-acid (59-aa) MADS box at their amino termini (Fig. ?(Fig.1A).1A). In addition, all MEF2 proteins contain a conserved 26-aa region carboxyl to the MADS box, termed the MEF2 domain, which is unique to MEF2 members (Fig. ?(Fig.1A).1A). Apart from the conserved MADS box and MEF2 domain, MEF2 proteins differ significantly in their central regions and carboxyl termini, which harbor transactivation domains (Fig. ?(Fig.1A)1A) (1). MEF2 preferentially binds to the MEF2 site, an AT-rich consensus sequence [(C/T)TA(A/T)4TA(A/G)], which is found in the promoters and enhancers of many genes (1). Open in a separate window FIG. 1. MEF2 is acetylated in vitro and in vivo. (A) Schematic representation of human MEF2C protein used in this study. The numbers give the positions of amino acids marking the boundaries of domains. (B) Purified His-thioredoxin-MEF2C (lane 1) and His-thioredoxin (lane 2) were subjected to in vitro AZD7507 acetylation assays with the immunoprecipitated Flag-p300. The dried gel was first subjected to fluorography (top panel), followed by rehydration and Coomassie blue staining (bottom panel). Ig-H, immunoglobulin heavy chain. (C) 293T cells were transfected with the plasmid encoding Flag-MEF2C, followed by in vivo sodium [3H]acetate labeling. The immunoprecipitated Flag-MEF2 was resolved by SDS-PAGE, and the gel was subjected to fluorography (top panel). The expression level of Flag-MEF2C was detected by immunoblotting (IB) (bottom panel). (D) Proliferating (grown in GM) and differentiating (grown in DM) C2C12 cells were labeled in vivo with sodium [3H]acetate. The endogenous MEF2s were separately immunoprecipitated (IP) from GM and DM cell lysates (twice as much GM lysate was used, because the MEF2 level is lower in GM) and resolved by SDS-PAGE. The gel was subjected to fluorography (top panel). The expression levels of MEF2s (center panel) and -tubulin (bottom panel) present in 10% of cell lysates used in the above immunoprecipitation were revealed by immunoblotting. MEF2 LUC7L2 antibody plays important roles in various biological processes. During myogenesis, many skeletal-muscle-specific genes (e.g., muscle creatine kinase, myogenin, and desmin) contain indispensable MEF2 sites in their promoters or enhancers (1). In the mouse myogenin promoter, a key proximal MEF2 site controls the expression level and tissue expression pattern of myogenin (5, 7, 38). MEF2 is also capable of directly interacting with AZD7507 members of the myogenic regulatory factors (MRFs) (i.e., MyoD, Myf5, myogenin, and MRF4) to synergistically activate many muscle-specific genes (22). In addition, MEF2 also plays a decisive role in cardiogenesis. It controls the expression of gene in mice is embryonically lethal due to a failure in heart development (17), whereas loss of in mice leads to deficiency in cardiac mitochondria and sudden death due to cardiac abnormality (25). Loss of a single gene in is also embryonically lethal, due to aborted differentiation of AZD7507 muscle cells in somites and heart (16). Mutations in also lead to a form of severe coronary artery disease in humans (35). In lymphocyte development, expression of the gene, an immediate-early gene involved in apoptosis of autoreactive T cells in response to T-cell-receptor signaling, is critically regulated by MEF2 (39). In neuronal cells, MEF2 is implicated in neuronal-activity-dependent cell survival (19). Recently, MEF2 has also been found to play a role in epithelial cell survival and maintenance of blood vessel integrity (12). In cells, MEF2 activity is regulated by diverse signaling pathways. The p38 mitogen-activated protein kinase- and BMK1/ERK5-mediated signaling pathways directly phosphorylate MEF2A and MEF2C in their transactivation domains and enhance AZD7507 their AZD7507 transcriptional activity (11, 13, 36, 41). In addition, the calcium/calmodulin-dependent protein kinase (CaMK) and phosphatase (calcineurin) also activate the transcriptional activity of MEF2. Whereas it remains unclear how calcineurin activates MEF2, the mechanism by which CaMK activates MEF2 has recently been elucidated. MEF2 interacts directly with the class II histone deacetylases (HDACs) via the MADS/MEF2 domain (20, 21)..