Search PubMed⌕ Search

SEARCH · Search PubMed

Results for “Myogenin”

Search indexed PubMed citations on genomics, clinical trials, systematic reviews and public health. Explore titles, authors and supplied subject terms, then open the PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 19 recordsLinked to original sources
PubMed1992

Mapping of myogenin transcription during embryogenesis using transgenes linked to the myogenin control region.

During vertebrate embryogenesis, the muscle-specific helix-loop-helix protein myogenin is expressed in muscle cell precursors in the developing somite myotome and limb bud before muscle fiber formation and is further upregulated during myogenesis. We show that cis-acting DNA sequences within the 5' flanking region of the mouse myogenin gene are sufficient to direct appropriate temporal, spatial, and tissue-specific transcription of myogenin during mouse embryogenesis. Myogenin-lacZ transgenes trace the fate of embryonic cells that activate myogenin transcription and suggest that myogenic precursor cells that migrate from the somite myotome to the limb bud are committed to a myogenic fate in the absence of myogenin transcription. Activation of a myogenin-lacZ transgene can occur in limb bud explants in culture, indicating that signals required for activation of myogenin transcription are intrinsic to the limb bud and independent of other parts of the embryo. These results reveal multiple populations of myogenic precursor cells during development and suggest the existence of regulators other than myogenic helix-loop-helix proteins that maintain cells in the early limb bud in the myogenic lineage.

Animals↗
PubMed1990

Myogenin resides in the nucleus and acquires high affinity for a conserved enhancer element on heterodimerization.

Myogenin is a member of a family of muscle-specific factors that can activate the muscle differentiation program in nonmyogenin cells. Using antibodies directed against unique domains of myogenin, we show in the present study that myogenin resides in the nucleus of differentiated muscle cells. Myogenin translated in vitro does not exhibit detectable DNA binding activity; however, when dimerized with the ubiquitous enhancer-binding factor E12, it acquires high affinity for an element in the core of the muscle creatine kinase (MCK) enhancer that is conserved among many muscle-specific genes. Antibody disruption experiments show that myogenin, synthesized during differentiation of the BC3H1 and C2 muscle cell lines, is part of a complex that binds to the same site in the MCK enhancer as myogenin-E12 translated in vitro. Mutagenesis of the myogenin-E12-binding site in the MCK enhancer abolishes binding of the heterodimer and prevents trans-activation of the enhancer by myogenin. The properties of myogenin suggest that its functions as a sequence-specific DNA-binding factor that interacts directly with muscle-specific genes during myogenesis. The dependence of myogenin on E12 for high-affinity DNA binding activity also suggests that the susceptibility of various cell types to the actions of myogenin may be influenced by the cellular factors with which it may interact.

Base Sequence↗
PubMed1991

Myogenin and MyoD join a family of skeletal muscle genes regulated by electrical activity.

Myogenin and MyoD are proteins that bind to the regulatory regions of a battery of skeletal muscle genes and can activate their transcription during muscle differentiation. We have recently found that both proteins interact with the enhancer of the nicotinic acetylcholine receptor (nAChR) alpha subunit, a gene that is regulated by innervation. This observation prompted us to study if myogenin and MyoD transcript levels are also regulated by skeletal muscle innervation. Using Northern blot analysis, we found that MyoD and myogenin mRNA levels begin to decline at embryonic day 17 and attain adult levels in muscle of newborn and 3-week-old mice, respectively. In contrast, nAChR mRNAs are highest in newborn and 1-week-old mouse muscle and decline thereafter to reach adult levels in 3-week-old mice. To determine if the down-regulation of myogenin and MyoD mRNA levels during development is due to innervation, we quantitated message levels in adult calf muscles after denervation. We found that in denervated muscle myogenin and MyoD mRNAs reach levels that are approximately 40- and 15-fold higher than those found in innervated muscle. Myogenin mRNAs begin to accumulate rapidly between 8 and 16 hr after denervation, and MyoD transcripts levels begin to increase sharply between 16 hr and 1 day after denervation. The increases in myogenin and MyoD mRNA levels precede the rapid accumulation of nAChR alpha-subunit transcripts; receptor mRNAs begin to accumulate significantly after 1 day of denervation. The effects of denervation are specific because skeletal alpha-actin mRNA levels are not affected by denervation. In addition, we found that the repression of myogenin and MyoD expression by innervation is due, at least in part, to "electrical activity." Direct stimulation of soleus muscle with extracellular electrodes repressed the increase of myogenin and MyoD transcripts after denervation by 4- to 3-fold, respectively. In view of these results, it is interesting to speculate that myogenin and/or MyoD may regulate a repertoire of skeletal muscle genes that are down-regulated by electrical activity.

Aging↗
PubMed1991

Transforming growth factor beta represses the actions of myogenin through a mechanism independent of DNA binding.

Myogenin belongs to a family of regulatory factors that can activate myogenesis when transfected into nonmyogenic cells. A conserved DNA sequence, known as an E box, serves as the target for binding and trans-activation by myogenin. Using 10T1/2 fibroblasts that constitutively express a transfected myogenin cDNA, we show that myogenin accumulates in the nucleus but is unable to initiate myogenesis when cells are maintained with transforming growth factor beta (TGF-beta) or high serum. Although the final effect of TGF-beta and high serum--inhibition of myogenesis--was the same, their effects on the DNA-binding properties of myogenin in vitro differed. TGF-beta did not affect the ability of myogenin to bind DNA, whereas serum diminished the in vitro DNA-binding activity of myogenin. The helix-loop-helix (HLH) protein Id, postulated to inhibit DNA binding of other HLH proteins, was induced by high serum but not by TGF-beta. The presence of Id correlated with the failure of myogenin to bind the muscle creatine kinase enhancer in vitro. These findings suggest that serum can inhibit myogenesis by attenuating the DNA-binding activity of myogenin, possibly as a consequence of Id protein expression, whereas TGF-beta acts through a mechanism distal to DNA sequence recognition by myogenin and independent of Id.

Animals↗
PubMed1992

MyoD, myogenin independent differentiation of primordial myoblasts in mouse somites.

The accumulation of two myogenic regulatory proteins, MyoD and myogenin, was investigated by double-immunocytochemistry and correlated with myosin heavy chain expression in different classes of myoblasts in culture and during early myogenesis in vivo. During in vitro differentiation of fetal myoblasts, MyoD-positive cells were detected first, followed by the appearance of cells positive for both MyoD and myogenin and finally by the appearance of differentiated myocytes and myotubes expressing myosin heavy chain (MHC). A similar pattern of expression was observed in cultures of embryonic and satellite cells. In contrast, most myogenic cells isolated from newly formed somites, expressed MHC in the absence of detectable levels of myogenin or MyoD. In vivo, the appearance of both myogenin and MyoD proteins was only detected at 10.5 d postcoitum (d.p.c.), when terminally differentiated muscle cells could already be identified in the myotome. Parasagittal sections of the caudal myotomes of 10.5-d-old embryos showed that expression of contractile proteins preceded the expression of myogenin or MyoD and, when coexpressed, MHC and myogenin did not co-localize within all the cells of the myotome. In the limb bud, however, many myogenin (or MyoD) positive/MHC negative cells could be observed in the proximal region at day 11. During further embryonic development the expression of these proteins remained constant in all the muscle anlagens examined, decreasing to a low level during the late fetal period. Western and Northern analysis confirmed that the myogenin protein could only be detected after 10.5 d.p.c. while the corresponding message was clearly present at 9.5 d.p.c., strongly suggesting a posttranscriptional regulation of myogenin during this stage of embryonic development. These data show that the first myogenic cells which appear in the mouse myotome, and can be cultured from it, accumulate muscle structural proteins in their cytoplasm without expressing detectable levels of myogenin protein (although the message is clearly accumulated). Neither MyoD message or protein are detectable in these cells, which may represent a distinct myogenic population whose role in development remains to be established.

Animals↗
PubMed1992

Regulation of myogenin expression in normal and transformed myogenic cell lines.

The control of myogenin (Myf-4), one of the muscle-specific regulatory proteins, is particularly interesting since its expression appears obligatory in myoblasts at the onset of differentiation. We isolated the human Myf-4 (myogenin) gene and determined promoter elements which direct cell type-specific expression and are subject to transactivation by the muscle transcription factors Myf-5 and MyoD1 in fibroblasts. Extrinsic signals such as serum components and purified growth factors or potential intracellular signals such as cAMP down-regulate transcription of the myogenin gene. Constitutive expression of the catalytic subunit of PKA completely suppresses transactivation of the myogenin promoter by Myf-5 or MyoD1 suggesting that cAMP may act via phosphorylation by PKA. In contrast to normal myogenic cell lines in which differentiation and myogenin expression can be induced by the removal of serum components, retinoic acid (RA) is required for differentiation in the rat rhabdomyosarcoma cell line BA-Han-1C. This model system was utilized to investigate factors which influence the balance between the transformed state and differentiation. Administration of retinoic acid to BA-Han-1C cells leads to the accumulation of myogenin mRNA approximately 48 h after the addition of RA. This late induction requires ongoing protein- and DNA-synthesis suggesting that trans- and cis-acting factors may be involved in the control. The critical involvement of myogenin in the process of terminal muscle differentiation was also demonstrated in the rat L6 muscle cell line which has been blocked for differentiation by the transforming protein E1a of Ad5 adenovirus. In cells which stably express E1a, myogenin expression is completely suppressed while Myf-5 continues to be synthesized normally. However, E1a inhibits the transactivator function of Myf-5, as demonstrated on GAL4-Myf5 chimeric proteins. A possible interpretation of this result is that Myf-5 or factors activated by Myf-5 are required for the expression of myogenin and myogenin itself is necessary for the terminal differentiation of myoblasts.

Blood↗
PubMed1991

Inefficient homooligomerization contributes to the dependence of myogenin on E2A products for efficient DNA binding.

Myogenin is a muscle-specific transcription factor that can activate myogenesis; it belongs to a family of transcription factors that share homology within a basic region and an adjacent helix-loop-helix (HLH) motif. Although myogenin alone binds DNA inefficiently, in the presence of the widely expressed HLH proteins E12 and E47 (encoded by the E2A gene), it forms heterooligomers that bind with high affinity to a DNA sequence known as a kappa E-2 site. In contrast, E47 and to a lesser extent E12 are both able to bind the kappa E-2 site relatively efficiently as homooligomers. To define the relative contributions of the basic regions of myogenin and E12 to DNA binding and muscle-specific gene activation, we created chimeras of the two proteins by swapping their basic regions. We showed that myogenin's weak affinity for the kappa E-2 site is attributable to inefficient homooligomerization and that the myogenin basic domain alone can mediate high-affinity DNA binding when placed in E12. Within a heterooligomeric complex, two basic regions were required to form a high-affinity DNA-binding domain. Basic-domain mutants of myogenin or E2A gene products that cannot bind DNA retained the ability to oligomerize and could abolish DNA binding of the wild-type proteins in vitro. These myogenin and E2A mutants also acted as trans-dominant inhibitors of muscle-specific gene activation in vivo. These findings support the notion that muscle-specific gene activation requires oligomerization between myogenin and E2A gene products and that E2A gene products play an important role in myogenesis by enhancing the DNA-binding activity of myogenin, as well as other myogenic HLH proteins.

Amino Acid Sequence↗
PubMed1992

Retinoic acid induces myogenin synthesis and myogenic differentiation in the rat rhabdomyosarcoma cell line BA-Han-1C.

Two clonal rat rhabdomyosarcoma cell lines BA-Han-1B and BA-Han-1C with different capacities for myogenic differentiation have been examined for the expression of muscle regulatory basic helix-loop-helix (bHLH) proteins of the MyoD family. Whereas cells of the BA-Han-1C subpopulation constitutively expressed MyoD1 and could be induced to differentiate with retinoic acid (RA), BA-Han-1B cells did not express any of the myogenic control factors and appeared to be largely differentiation-defective. Upon induction with RA, BA-Han-1C cells expressed also myogenin, in contrast to BA-Han-1B cells which never activated any of the genes encoding muscle bHLH factors. The onset of myogenin transcription in BA-Han-1C cells required de novo protein synthesis and DNA replication suggesting that RA probably did not act directly on the myogenin gene. Although MyoD1 was expressed in proliferating BA-Han-1C myoblasts, muscle-specific reporter genes were not activated indicating that MyoD was biologically inactive. However, transfections with plasmid expressing additional MyoD1 protein resulted in the transactivation of muscle genes even in the absence of RA. mRNA encoding the negative regulatory HLH protein Id was expressed in proliferating BA-Han-1C cells and disappeared later after RA induction which suggested that it may be involved in the regulation of MyoD1 activity. The myogenic differentiation of malignant rhabdomyosarcoma cells strictly correlated with the activation of the myogenin gene. In fact, stable transfections of BA-Han-1C cells with myogenin expressing plasmids resulted in spontaneous differentiation. Together, our results suggest that the transformed and undifferentiated phenotype of BA-Han-1C rhabdomyosarcoma cells is associated with the inactivation of the myogenic factor MyoD1 as well as lack of myogenin expression. RA alleviates the inhibition of myogenic differentiation, probably by activating MyoD protein and myogenin gene transcription. BA-Han-1B cells did not respond to RA and the differentiated phenotype could not be restored by overexpression of MyoD1 or myogenin.

Animals↗
PubMed1991

Myogenin induces the myocyte-specific enhancer binding factor MEF-2 independently of other muscle-specific gene products.

The myocyte-specific enhancer-binding factor MEF-2 is a nuclear factor that interacts with a conserved element in the muscle creatine kinase and myosin light-chain 1/3 enhancers (L. A. Gossett, D. J. Kelvin, E. A. Sternberg, and E. N. Olson, Mol. Cell. Biol. 9:5022-5033, 1989). We show in this study that MEF-2 is regulated by the myogenic regulatory factor myogenin and that mitogenic signals block this regulatory interaction. Induction of MEF-2 by myogenin occurs in transfected 10T1/2 cells that have been converted to myoblasts by myogenin, as well as in CV-1 kidney cells that do not activate the myogenic program in response to myogenin. Through mutagenesis of the MEF-2 site, we further defined the binding site requirements for MEF-2 and identified potential MEF-2 sites within numerous muscle-specific regulatory regions. The MEF-2 site was also found to bind a ubiquitous nuclear factor whose binding specificity was similar to but distinct from that of MEF-2. Our results reveal that MEF-2 is controlled, either directly or indirectly, by a myogenin-dependent regulatory pathway and suggest that growth factor signals suppress MEF-2 expression through repression of myogenin expression or activity. The ability of myogenin to induce MEF-2 activity in CV-1 cells, which do not activate downstream genes associated with terminal differentiation, also demonstrates that myogenin retains limited function within cell types that are nonpermissive for myogenesis and suggests that MEF-2 is regulated independently of other muscle-specific genes.

Amino Acid Sequence↗
PubMed1992

The basic region of myogenin cooperates with two transcription activation domains to induce muscle-specific transcription.

Myogenin is a skeletal muscle-specific transcription factor that can activate myogenesis when introduced into a variety of nonmuscle cell types. Activation of the myogenic program by myogenin is dependent on its binding to a DNA sequence known as an E box, which is associated with numerous muscle-specific genes. Myogenin shares homology with MyoD and other myogenic regulatory factors within a basic region and a helix-loop-helix (HLH) motif that mediate DNA binding and dimerization, respectively. Here we show that the basic region-HLH motif of myogenin alone lacks transcriptional activity and is dependent on domains in the amino and carboxyl termini to activate transcription. Analysis of these N- and C-terminal domains through creation of chimeras with the DNA-binding domain of the Saccharomyces cerevisiae transcription factor GAL4 revealed that they act as strong transcriptional activators. These transcription activation domains are dependent for activity on a specific amino acid sequence within the basic region, referred to as the myogenic recognition motif (MRM), when an E box is the target for DNA binding. However, the activation domains function independent of the MRM when DNA binding is mediated through a heterologous DNA-binding domain. The activation domain of the acidic coactivator VP16 can substitute for the myogenin activation domains and restore strong myogenic activity to the basic region-HLH motif. Within a myogenin-VP16 chimera, however, the VP16 activation domain also relies on the MRM for activation of the myogenic program. These findings reveal that DNA binding and transcriptional activation are separable functions, encoded by different domains of myogenin, but that the activity of the transcriptional activation domains is influenced by the DNA-binding domain. Activation of muscle-specific transcription requires collaboration between the DNA-binding and activation domains of myogenin and is dependent on events in addition to DNA binding.

Amino Acid Sequence↗
PubMed1992

Analysis of the myogenin promoter reveals an indirect pathway for positive autoregulation mediated by the muscle-specific enhancer factor MEF-2.

Transcriptional cascades that specify cell fate have been well described in invertebrates. In mammalian development, however, gene hierarchies involved in determination of cell lineage are not understood. With the recent cloning of the MyoD family of myogenic regulatory factors, a model system has become available with which to study the dynamics of muscle determination in mammalian development. Myogenin, along with other members of the MyoD gene family, possesses the apparent ability to redirect nonmuscle cells into the myogenic lineage. This ability appears to be due to the direct activation of an array of subordinate or downstream genes which are responsible for formation and function of the muscle contractile apparatus. Myogenin-directed transcription has been shown to occur through interaction with a DNA consensus sequence known as an E box (CANNTG) present in the control regions of numerous downstream genes. In addition to activating the transcription of subordinate genes, members of the MyoD family positively regulate their own expression and cross-activate one another's expression. These autoregulatory interactions have been suggested as a mechanism for induction and maintenance of the myogenic phenotype, but the molecular details of the autoregulatory circuits are undefined. Here we show that the myogenin promoter contains a binding site for the myocyte-specific enhancer-binding factor, MEF-2, which can function as an intermediary of myogenin autoactivation. Since MEF-2 can be induced by myogenin, these results suggest that myogenin and MEF-2 participate in a transcriptional cascade in which MEF-2, once induced by myogenin, acts to amplify and maintain the myogenic phenotype by acting as a positive regulator of myogenin expression.

Amino Acid Sequence↗
PubMed1991

Insulin-like growth factor-I stimulates terminal myogenic differentiation by induction of myogenin gene expression.

Stimulation of myogenic differentiation by the insulin-like growth factors (IGFs) has been established for many years, but our attempts to elucidate the mechanism of that stimulation have been successful only in eliminating some likely possibilities. The recent discovery of a family of muscle determination genes has opened a new approach to this question, allowing specific focus on those genes that might play central roles in controlling myogenesis. We now report that IGF-I stimulates terminal myogenic differentiation in L6A1 cells by inducing a large increase in expression of the myogenin gene. This conclusion is supported by the following observations. 1) Myogenin mRNA is elevated by IGF-I, with a concentration dependency that parallels the stimulation of differentiation, including a decrease in stimulation at higher concentrations. 2) The time course of elevation of myogenin mRNA is consistent with its acting as an intermediate in the signalling pathway between occupancy of the IGF-I receptor and induction of expression of muscle-specific genes. 3) Inhibitors of myogenesis also inhibit elevation of myogenin mRNA in response to IGF-I. 4) An antisense oligonucleotide to the N-terminus of myogenin prevents the stimulation of differentiation by IGF-I and IGF-II, but has no effect on other actions of IGF-I on myoblasts. MyoD has been reported not to be expressed in L6 cells, and the expression of myf-5 and herculin/myf-6/MRF4 is reportedly low or undetectable. Thus, the stimulation of differentiation by IGF-I can be attributed largely, if not entirely, to increased expression of the myogenin gene. However, the relatively long time period between addition of the IGFs and elevation of myogenin mRNA as well as the inhibition of this process by several inhibitors indicate that increased myogenin mRNA levels are not a simple direct result of occupation of the IGF-I receptor.

Animals↗
PubMed1992

Induction and peak gene expression of insulin-like growth factor II follow that of myogenin during differentiation of BC3H-1 muscle cells.

Acting through hormonal and/or autocrine/paracrine mechanisms, the insulin-like growth factors (IGFs) stimulate the differentiation of muscle cells. Previous studies have suggested that one mechanism by which IGFs stimulate muscle cell differentiation is by increasing the expression of myogenin, a DNA binding protein that regulates the expression of muscle-specific genes. While exogenous IGF peptides increase myogenin mRNA, the role of endogenously produced IGF peptides in myogenin expression has not been established. In addition, the potential role of IGFs in regulating the expression of Id, a protein in myoblasts that can inhibit the action of myogenin-like peptides, is also unknown. In the present study, we have examined the kinetics of accumulation of myogenin and IGF-II mRNAs during differentiation of BC3H-1 mouse muscle cells and have explored the potential role of IGFs in regulating Id expression. During differentiation induced by serum withdrawal, induction of myogenin expression preceded that of IGF-II, the principal IGF peptide expressed by these cells. In addition, Id expression decreased within two hours in serum-free medium and was not affected by IGF treatment. Thus, these studies suggest that endogenously-produced IGF-II may stimulate muscle cell differentiation after both the decrease in Id and the induction of myogenin gene expression have occurred.

Animals↗
PubMed1991

c-myc inhibition of MyoD and myogenin-initiated myogenic differentiation.

In vertebrate development, a prominent feature of several cell lineages is the coupling of cell cycle regulation with terminal differentiation. We have investigated the basis of this relationship in the skeletal muscle lineage by studying the effects of the proliferation-associated regulator, c-myc, on the differentiation of MyoD-initiated myoblasts. Transient cotransfection assays in NIH 3T3 cells using MyoD and c-myc expression vectors demonstrated c-myc suppression of MyoD-initiated differentiation. A stable cell system was also developed in which MyoD expression was constitutive, while myc levels could be elevated conditionally. Induction of this conditional c-myc suppressed myogenesis effectively, even in the presence of MyoD. c-myc suppression also prevented up-regulation of a relative of MyoD, myogenin, which is normally expressed at the onset of differentiation in all muscle cell lines examined and may be essential for differentiation. Additional experiments tested whether failure to differentiate in the presence of myc could be overcome by providing myogenin ectopically. Cotransfection of c-myc with myogenin, MyoD, or a mixture of myogenin and MyoD showed that neither myogenin alone nor myogenin plus MyoD together could bypass the c-myc block. The effects of c-myc were further dissected by showing that c-myc can inhibit differentiation independently of Id, a negative regulator of muscle differentiation. These results lead us to propose that c-myc and Id constitute independent negative regulators of muscle differentiation, while myogenin and any of the other three related myogenic factors (MyoD, Myf-5, and MRF4/herculin/Myf-6) act as positive regulators.

Animals↗
PubMed1990

Role of myogenin in myoblast differentiation and its regulation by fibroblast growth factor.

In BC3H-1 cells, a model for studying the events that occur when myoblasts transform into myocytes, we observed that differentiation was associated with a 10-20-fold increase in both gene transcription and mRNA levels for myogenin, a member of the myc gene family. In contrast, the expression of the related protein MyoD1 was not influenced. This enhanced expression of myogenin was followed by morphological differentiation and increased expression of the nicotinic acetylcholine receptor, a muscle-specific gene product. Exposure of BC3H-1 cells to a 20-base myogenin antisense oligomer blocked morphological differentiation and resulted in nearly complete inhibition of acetylcholine receptor protein expression. To further study the relationship between muscle cell differentiation and myogenin gene expression, fibroblast growth factor (FGF), a known inhibitor of myogenic differentiation, was employed. FGF treatment inhibited myogenin gene transcription and BC3H-1 cell differentiation. These results demonstrate therefore that myogenin is an important regulator of skeletal muscle cell differentiation, and the expression of myogenin is under the control of FGF.

Animals↗
PubMed1989

Myogenin, a factor regulating myogenesis, has a domain homologous to MyoD.

In this report, we describe the isolation, sequence, and initial characterization of the cDNA for the muscle-specific regulatory factor skeletal myogenin. Transfection of myogenin into the mesenchymal cell line C3H10T1/2 produces cells expressing muscle-specific markers. Myogenin is absent in undifferentiated cells, peaks, and then declines following a stimulus to differentiate, and is overexpressed in myoblasts selected with 5-bromodeoxyuridine for the overproduction of factors that regulate the decision to differentiate. High levels of myogenin transcripts are present in the myotomal region of somites at 8.5 days of gestation in the mouse. Although myogenin and MyoD are different genes, they share the myc homology domain. Myogenin and MyoD thus form part of a gene family regulating myogenesis, and together with myd may constitute a set of factors that interact to regulate the determination and differentiation of muscle cells.

Amino Acid Sequence↗
PubMed1992

Cyclic amplification and selection of targets for multicomponent complexes: myogenin interacts with factors recognizing binding sites for basic helix-loop-helix, nuclear factor 1, myocyte-specific enhancer-binding factor 2, and COMP1 factor.

Myogenin is one of four muscle-specific basic helix-loop-helix regulatory factors involved in controlling myogenesis. We here describe various protein complexes that increase the affinity of myogenin for DNA. We mixed an oligonucleotide containing a degenerate center large enough to accommodate multiple binding sites with crude myotube nuclear extracts and used cyclic amplification and selection of targets with an antimyogenin monoclonal antibody to isolate protein-DNA complexes. Since each cycle of selection results in the enrichment for the sequences with the highest affinity, we isolated multicomponent sites in which myogenin binding was increased by its interaction with other DNA binding proteins. Myogenin interacts with members of the nuclear factor 1 family, the muscle-specific factor myocyte-specific enhancer-binding factor 2, and another factor, COMP1 (cooperates with myogenic proteins 1), that binds to the sequence TGATTGAC. Myogenin also exhibits cooperative binding with other proteins that recognize CANNTG motifs, and various constraints on spacing and orientation were observed. The application of this approach to other transcription factors should not only help identify the different functions of myogenin versus other members of the muscle basic helix-loop-helix regulatory family but also help define the general combinatorial mechanisms involved in eukaryotic gene regulation.

Base Sequence↗
PubMed1992

Fos and Jun repress transcriptional activation by myogenin and MyoD: the amino terminus of Jun can mediate repression.

Myogenin and MyoD belong to a family of muscle-specific helix-loop-helix (HLH) proteins that have the potential to activate muscle-specific genes in nonmyogenic cells. Peptide growth factors can block the ability of myogenin and MyoD to activate their target genes. Here, we show that the growth factor-inducible proto-oncogenes c-fos, c-jun, and junB mimic the effects of exogenous growth factors and suppress trans-activation of the muscle creatine kinase (MCK) enhancer by myogenin and MyoD. In contrast, JunD, which shares DNA-binding specificity with JunB and c-Jun but is expressed constitutively in muscle cells, is an inefficient inhibitor of the trans-activating capacity of myogenin and MyoD. Transcriptional repression by Fos and Jun is specific to myogenic HLH proteins and is not observed with the widely expressed HLH protein E47, which recognizes the same DNA sequence. Repression of the MCK enhancer by Fos and Jun is targeted at the myogenin and MyoD DNA recognition sequence and can be mediated by the amino terminus of c-Jun. Comparison of several myogenin mutants for their responsiveness to Fos and Jun shows that repression is directed at the basic-HLH region. These results indicate that members of the Jun family can be distinguished on the basis of their effects on muscle-specific transcription and suggest there is cross talk between transcription factors that control myogenesis and those involved in cell proliferation.

Animals↗