Similar to other MADS box proteins MEF factors associate

Similar to other MADS box proteins, MEF2 factors associate with a variety of transcriptional cofactors to control specific sets of downstream target genes. Some of the cofactors such as MyoD, GATA4, NFAT, TH receptor, p300, 14.3.3 and ERK5 stimulate MEF2 activity, whereas others, such as HDACs4-5-7-9, MITR and cabin, repress MEF2 function (reviewed in McKinsey et al., 2002). Although MEF2 proteins are expressed at high levels in muscle cells, they are unable to activate muscle gene expression alone, but they potentiate the transcriptional activity of myogenic bHLH proteins (MyoD family). In many muscle cell gene promoters, a MEF2 binding site and a MyoD binding site (E box) are found side by side, and the interaction of MEF2 with MyoD seems to be important for muscle cell differentiation Cserjesi and Olson, 1991, Molkentin et al., 1995. This cooperativity is mediated by direct interactions between the MADS box of MEF2 and the bHLH region of myogenic proteins. The synergy results through physical association of MEF2 proteins with myogenic bHLH proteins that must first heterodimerize with E proteins (such as E12) in order to associate with MEF2. Moreover, these factors can also cooperate to activate transcription through a single DNA binding site eliminating the requirement for both factors simultaneously binding DNA. Indeed, MEF2 can interact with MyoD/E12 heterodimers bound to E box, or MyoD/E12 heterodimer can interact with MEF2 bound to DNA (reviewed in Molkentin and Olson, 1996, Black and Olson, 1998). In cardiac and visceral muscle cells, where the MyoD family of transcription factors is not expressed, MEF2 proteins are recruited to target promoters by the cell-specific GATA transcription factors. The MEF2–GATA4 synergy is mediated by physical interaction between the MEF2 DNA binding domain and the carboxy zinc finger of GATA and requires the transactivation domains of both factors. GATA 22(R)-hydroxy Cholesterol sale are necessary and sufficient for cooperativity with MEF2 (Morin et al., 2000). Thus, MyoD and GATA4 proteins interact with the two types of MADS box proteins MEF2 and SRF, and it was recently reported that TEF1, another SRF coactivator, also activates synergistically many muscle-specific genes in association with MEF2. The MEF2 MADS box domain is not sufficient for MEF2 interaction with TEF1; additional sequences in the activation domains of both proteins are required for in vivo association (Maeda et al., 2002). Repression of MEF2 activity depends on its association with HDACs-4-5-7-9 McKinsey et al., 2001a, Zhou et al., 2001. This interaction does not affect MEF2 DNA binding activity and results in recruitment of HDACs to MEF2-dependent gene regulatory regions. Release of HDAC from MEF2 may occur by phosphorylation of two conserved serine residues in the N-terminal region of the HDAC through CaMKI signalling pathway (McKinsey et al., 2000). This phosphorylation creates docking sites for the intracellular chaperone protein 14.3.3. Upon binding to 14.3.3, HDACs are released from MEF2 and transported to the cytoplasm, as a result of masking HDAC nuclear localization signal and unmasking a nuclear export signal Grozinger and Schreiber, 2000, McKinsey et al., 2001b. The released MEF2 is then able to associate with the p300 acetyltransferase coactivator and stimulates MEF2-dependent genes. Similar to HDAC, p300 associates with the MADS/MEF2 domains of MEF2 factors, suggesting that interaction between MEF2 and HATs or HDACs are mutually exclusive (McKinsey et al., 2001a). Interestingly, 14.3.3 was also recently shown to stimulate MEF2D activity by directly associating with the MADS/MEF2 domains. It was proposed that 14.3.3 activates MEF2D by competitively inhibiting HDAC4 from binding to MEF2, and thereby affects muscle cell differentiation (Choi et al., 2001). It was proven that the cytoplasmic retention of HDAC is necessary for correct regulation of C2C12 myoblast differentiation at the step of myoblast fusion into multinucleated myotubes. Indeed, in differentiated myoblasts, HDAC4 is sequestered in the cytoplasm by 14.3.3, allowing activation of MEF2 by p300, whereas in myotubes, HDAC relocates to the nucleus where it associates with MEF2 to repress transcription (Miska et al., 2001).