Transcriptional activation of the SERCA2 gene by calcium in cardiac myocytes (original) (raw)
Related papers
Molecular and cellular …, 1996
There are four members of the myocyte enhancer factor 2 (MEF2) family of transcription factors in vertebrates, MEF2A, -B, -C, and -D, which have homology within a MADS box at their amino termini and an adjacent motif known as the MEF2 domain. These factors activate muscle gene expression by binding as homoand heterodimers to an A/T-rich DNA sequence in the control regions of muscle-specific genes. To understand the mechanisms of muscle gene activation by MEF2 factors, we generated a series of deletion and site-directed mutants of MEF2C. These mutants demonstrated that the MADS and MEF2 domains mediate DNA binding and dimerization, whereas the carboxyl terminus is required for transcriptional activation. Amino acids that are essential for MEF2 site-dependent transcription but which do not affect DNA binding were also identified in the MEF2 domain. This type of positive-control mutant demonstrates that the transcription activation domain of MEF2C, although separate from the MEF2 domain, is dependent on this domain for transcriptional activation through the MEF2 site. MEF2 mutants that are defective for DNA binding act as dominant negative mutants and can inhibit activation of MEF2-dependent genes by wild-type MEF2C.
MEF2C DNA-binding activity is inhibited through its interaction with the regulatory protein Ki-1/57
FEBS Letters, 2005
Myocyte enhancer factor (MEF2) are MADS box transcription factors that play important roles in the regulation of myogenesis and morphogenesis of muscle cells. MEF2 proteins are activated by mechanical overload in the heart. In this study, we found the interaction of MEF2C with the regulatory protein Ki-1/57 using yeast two-hybrid system. This interaction was confirmed by GST-pull down assay in vitro and by co-immunoprecipitation in vivo. This interaction is also dependent on pressure overload in the heart. Co-imunoprecipitation assay with anti-MEF2 and anti-Ki-1/57 antibodies demonstrated a basal association between these proteins in the left ventricles of control rats. Pressure overload caused a reduction in this association. Ki-1/57 co-localizes with MEF2 in the nucleus of myocytes of control rats. However, after submitting the animals to pressure overload Ki-1/57 leaves the nucleus thereby decreasing this co-localization. Ki-1/57 also exerts an inhibitory effect upon MEF2C DNA binding activity. These results suggest that Ki-1/57 is a new interacting partner of MEF2 protein and may be involved in the regulation of MEF2 at the onset of hypertrophy.
Journal of Biological Chemistry, 1998
The murine adult IIB myosin heavy chain (IIB MyHC) gene is expressed only in certain skeletal muscle fibers. Within the proximal promoter are two A ؉ T-rich motifs, mAT1 and mAT2, which greatly enhance muscle-specific transcription; myogenic cells contain proteins that bind to these sequences. MEF-2 binds to both mAT1 and mAT2; a mutation abolishing its binding to mAT1 greatly diminishes the activity of the promoter. Both mAT motifs also form complexes with a protein requiring a target sequence typical of POU domain proteins, which migrate in electrophoretic mobility shift assays to the same position as a complex containing purified Oct-1 and which are supershifted by an antibody specific to Oct-1; this protein is therefore probably Oct-1. Footprinting experiments demonstrate that mAT1 is preferentially occupied by MEF-2 and mAT2 by Oct-1 and that these two proteins appear to bind cooperatively to their respective sites. Although the two mAT motifs have sequences that are very similar, they nonetheless exhibit distinct behaviors and perform differently in the activation of the promoter. The contribution of the IIB MyHC gene to specification of the myogenic phenotype is thus at least in part regulated by MEF-2 and Oct-1.
MEF2B is a potent transactivator expressed in early myogenic lineages
Molecular and cellular biology, 1996
There are four members of the myocyte enhancer binding factor 2 (MEF2) family of transcription factors, MEF2A, -B, -C, and -D, that have homology within an amino-terminal MADS box and an adjacent MEF2 domain that together mediate dimerization and DNA binding. MEF2A, -C, and -D have previously been shown to bind an A/T-rich DNA sequence in the control regions of numerous muscle-specific genes, whereas MEF2B was reported to be unable to bind this sequence unless the carboxyl terminus was deleted. To further define the functions of MEF2B, we analyzed its DNA binding and transcriptional activities. In contrast to previous studies, our results show that MEF2B binds the same DNA sequence as other members of the MEF2 family and acts as a strong transactivator through that sequence. Transcriptional activation by MEF2B is dependent on the carboxyl terminus, which contains two conserved sequence motifs found in all vertebrate MEF2 factors. During mouse embryogenesis, MEF2B transcripts are exp...
Journal of Biological Chemistry, 1997
The expression of the cardiac myosin light chain 2 (MLC2) gene is repressed in skeletal muscle as a result of the negative regulation of its transcription. Two regulatory elements, the cardiac specific sequence (CSS) located upstream (؊360 base pairs) and a downstream negative modulatory sequence (NMS), which function in concert with each other, are required for repression of the MLC2 promoter activity in skeletal muscle. Individually, CSS and NMS have no effect. Transient transfection analysis with recombinant plasmids indicated that CSS-and NMS-mediated repression of transcription is position-and orientation-dependent and is transferable to heterologous promoters. A minimal conserved motif, GAAG/CTTC, present in both CSS and NMS, is responsible for repression as the mutation in the core CTTC sequence alone was sufficient to abrogate its repressor activity. The DNA binding assay by gel mobility shift analysis revealed that one of the two complexes, CSSBP2, is significantly enriched in embryonic skeletal muscle relative to cardiac muscle. In extracts from adult skeletal muscle, where the cardiac MLC2 expression is suppressed, both complexes, CSSBP1 and CSSBP2, were present, whereas the cardiac muscle extracts contained CSSBP1 alone, suggesting that the protein(s) in the CSSBP2 complex accounts for the negative regulation of cardiac MLC2 in skeletal muscle. A partial cDNA clone (Nished) specific for the candidate repressor factor was isolated by expression screening of the skeletal muscle cDNA library by multimerized CSS-DNA as probe. The recombinant Nished protein binds to the CSS-DNA, but not to ⌬CSS-DNA where the core CTTC sequence was mutated. The amino acid sequence of Nished showed a significant structural similarity to the sequence of transcription factor "runt," a known repressor of gap and pair-rule gene expression in Drosophila.
Journal of Molecular Biology, 2010
Myocyte enhancer factor 2 (MEF2) regulates specific gene expression in diverse developmental programs and adaptive responses. MEF2 recognizes DNA and interacts with transcription cofactors through a highly conserved N-terminal domain referred to as the MADS-box/MEF2 domain. Here we present the crystal structure of the MADS-box/MEF2 domain of MEF2A bound to DNA. In contrast to previous structural studies showing that the MEF2 domain of MEF2A is partially unstructured, the present study reveals that the MEF2 domain participates with the MADS-box in both dimerization and DNA binding as a single domain. The sequence divergence at and immediately following the C-terminal end of the MEF2 domain may allow different MEF2 dimers to recognize different DNA sequences in the flanking regions. The current structure also suggests that the ligand-binding pocket previously observed in the Cabin1-MEF2B-DNA complex and the HDAC9 (histone deacetylase 9)-MEF2B-DNA complex is not induced by cofactor binding but rather preformed by intrinsic folding. However, the structure of the ligand-binding pocket does undergo subtle but significant conformational changes upon cofactor binding. On the basis of these observations, we generated a homology model of MEF2 bound to a myocardin family protein, MASTR, that acts as a potent coactivator of MEF2-dependent gene expression. The model shows excellent shape and chemical complementarity at the binding interface and is consistent with existing mutagenesis data. The apo structure presented here can also serve as a target for virtual screening and soaking studies of small molecules that can modulate the function of MEF2 as research tools and therapeutic leads.
Post-translational control of the MEF2A transcriptional regulatory protein
Nucleic Acids Research, 1999
Myocyte enhancer factor 2 (MEF2) transcriptional regulatory proteins are key regulators of musclespecific gene expression and also play a general role in the cellular response to growth factors, cytokines and environmental stressors. To identify signaling pathway components that might mediate these events, the potential role of MAP kinase and PKC signaling in the modulation of MEF2A phosphorylation and transcriptional activity were therefore studied. In transient transfection reporter assays, activated p38 MAP kinase potently increased MEF2A trans-activating potential, PKCδ δ δ δ and ε ε ε ε isotypes enhanced MEF2A transactivation to a lesser extent, while the ERK1/2 and JNK/SAPK pathways were without effect. A GAL4-based assay system showed that p38 MAP kinase and PKCδ δ δ δ target the MEF2A transactivation domain. We also observed an increase in p38 MAP kinase activity in congruence with the increase in MEF2A expression in differentiating primary muscle cells. COS cells overexpressing MEF2A alone or with one of the kinases were metabolically labeled with [ 32 P]orthophosphate and MEF2A was immunoprecipitated using specific anti-MEF2A antibodies. MEF2A from cells co-transfected with activated p38 MAP kinase showed a decreased electrophoretic mobility due to phosphorylation. Subsequent phosphopeptide mapping and phosphoamino acid analysis indicated the appearance of several phoshopeptides due to p38 MAP kinase activation of MEF2A which were due to phosphorylation on serine and threonine residues. These studies position MEF2A as a nuclear target for the p38 MAP kinase signaling pathway.