A new myocyte-specific enhancer-binding factor that recognizes a conserved element associated with multiple muscle-specific genes.

Abstract

Exposure of skeletal myoblasts to growth factor-deficient medium results in transcriptional activation of muscle-specific genes, including the muscle creatine kinase gene (mck). Tissue specificity, developmental regulation, and high-level expression of mck are conferred primarily by a muscle-specific enhancer located between base pairs (bp) -1350 and -1048 relative to the transcription initiation site (E. A. Sternberg, G. Spizz, W. M. Perry, D. Vizard, T. Weil, and E. N. Olson, Mol. Cell. Biol. 8:2896-2909. To begin to define the regulatory mechanisms that mediate the selective activation of the mck enhancer in differentiating muscle cells, we have further delimited the boundaries of this enhancer and analyzed its interactions with nuclear factors from a variety of myogenic and nonmyogenic cell types. Deletion mutagenesis showed that the region between 1,204 and 1,095 bp upstream of mck functions as a weak muscle-specffic enhancer that is dependent on an adjacent enhancer element for strong activity. This adjacent activating element does not exhibit enhancer activity in single copy but acts as a strong enhancer when multimerized. Gel retardation assays combined with DNase I footprinting and diethyl pyrocarbonate interference showed that a nuclear factor from differentiated C2 myotubes and BC3H1 myocytes recognized a conserved A+T-rich sequence within the peripheral activating region. This myocyte-specific enhancer-binding factor, designated MEF-2, was undetectable in nuclear extracts from C2 or BC3H1 myoblasts or several nonmyogenic cell lines. MEF-2 was first detectable within 2 h after exposure of myoblasts to mitogen-deficient medium and increased in abundance for 24 to 48 h thereafter. The appearance of MEF-2 required ongoing protein synthesis and was prevented by fibroblast growth factor and type transforming growth factor, which block the induction of muscle-specific genes. A myoblast-specific factor that is down regulated within 4 h after removal of growth factors was also found to bind to the MEF-2 recognition site. A 10-bp sequence, which was shown by DNase I footprinting and diethyl pyrocarbonate interference to interact directly with MEF-2, was identified within the rat and human mck enhancers, the rat myosin light-chain (m1c)-113 enhancer, and the chicken cardiac mkc-2A promoter. Oligomers corresponding to the region of the mlc-113 enhancer, which encompasses this conserved sequence, bound MEF-2 and competed for its binding to the mck enhancer. These results thus provide evidence for a novel myocyte-specific enhancer-binding factor, MEF-2, that is expressed early in the differentiation program and is suppressed by specific polypeptide growth factors. The ability of MEF-2 to recognize conserved activating elements associated with multiple muscle-specific genes suggests that this factor may participate in the coordinate regulation of genes during myogenesis.Development requires precisely coordinated control of transcription. Differentiation of skeletal myoblasts to myotubes represents a well-defined sy...

p53 protein is activated during muscle differentiation and participates with MyoD in the transcription of muscle creatine kinase gene

“…Nevertheless, the enhancer and p53 elements combined induced the transcription of MCK in a cooperative manner (Figure 5a). At least two elements of de®ned transcription factors are found in the enhancer, MEF2 and MyoD (Lassar et al, 1989;Gossett et al, 1989). Each element was separately placed with the distal p53 element to control a minimal MCK reporter gene (Figure 5b).…”

“…At least two elements of de®ned transcription factors constitute the MCK enhancer; MEF2 and MyoD (Gossett et al, 1989;Lassar et al, 1989). Reporter genes that contained each of these sites in conjunction with the p53 distal element were constructed and used in transfection studies of 10T1/2 cells.…”

Changes in the expression of mRNAs for myogenic factors and other muscle‐specific proteins in experimental autoimmune myasthenia gravis

“…However, later in development in innervated muscle, MCK and MLC are regulated either by other members of the MyoD family or by other factors (i.e. MEF 2) which have been implicated in the regulation of these genes in cultured muscle cells [32]. A slight increase in the level of muscle dystrophin was observed in EAMG and a larger one following denervation.…”

Mouse dystrophin enhancer preferentially targets lacZ expression in skeletal and cardiac muscle

“…All of these enhancers rely on myogenic factors that act in combination with other ubiquitous factors to target skeletal or cardiac muscle (Firulli and Olson, 1997). Because myogenic factors are not present in the heart (Apone and Hauschka, 1995), enhancer function in the cardiac muscle depends on other factors such as mef-2 (Gossett et al, 1989), GATA-4 (Arceci et al, 1993), and SRF (Treisman, 1995). These three transcription factors may activate the mouse dystrophin enhancer in the heart.…”