Molkentin J D, Li L, Olson E N
Department of Molecular Biology, University of Texas, Southwestern Medical Center at Dallas, 75235-9148, USA.
J Biol Chem. 1996 Jul 19;271(29):17199-204. doi: 10.1074/jbc.271.29.17199.
Members of the myocyte enhancer factor-2 (MEF2) family of transcription factors activate muscle gene expression by binding an A/T-rich DNA sequence in the control regions of muscle-specific genes. There are four MEF2 factors in vertebrates, MEF2A-D, which share homology in an amino-terminal MADS domain and an adjacent region known as the MEF2 domain, that together mediate DNA binding and dimerization. We show that serine 59 located between the MADS and MEF2 domains of MEF2C is phosphorylated in vivo and can be phosphorylated in vitro by casein kinase-II (CKII). Phosphorylation of this site enhanced the DNA binding and transcriptional activity of MEF2C by increasing its DNA binding activity 5-fold. In vivo 32P labeling experiments showed that serine 59 is the only phosphorylation site in the MADS and MEF2 domains. Mutagenesis of this serine to an aspartic acid resulted in an increase in DNA binding and transcriptional activity of MEF2C comparable to that observed when this site was phosphorylated, suggesting that phosphorylation augments DNA binding activity by introducing negative charge. This phosphorylation site, which corresponds to a CKII recognition site, is conserved in all known MEF2 factors in organisms ranging from flies to humans, consistent with its importance for the functions of MEF2C.
肌细胞增强因子2（MEF2）转录因子家族的成员通过结合肌肉特异性基因控制区域中的富含A/T的DNA序列来激活肌肉基因表达。脊椎动物中有四种MEF2因子，即MEF2A - D，它们在氨基末端的MADS结构域和一个称为MEF2结构域的相邻区域具有同源性，这两个区域共同介导DNA结合和二聚化。我们发现，位于MEF2C的MADS结构域和MEF2结构域之间的丝氨酸59在体内被磷酸化，并且在体外可被酪蛋白激酶II（CKII）磷酸化。该位点的磷酸化通过将MEF2C的DNA结合活性提高5倍，增强了其DNA结合和转录活性。体内32P标记实验表明，丝氨酸59是MADS结构域和MEF2结构域中唯一的磷酸化位点。将该丝氨酸突变为天冬氨酸导致MEF2C的DNA结合和转录活性增加，与该位点被磷酸化时观察到的情况相当，这表明磷酸化通过引入负电荷增强了DNA结合活性。这个磷酸化位点对应于一个CKII识别位点，在从果蝇到人类的所有已知生物的MEF2因子中都是保守的，这与其对MEF2C功能的重要性一致。
