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Shahragim Tajbakhsh - One of the best experts on this subject based on the ideXlab platform.
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loss of myod and MYF5 in skeletal muscle stem cells results in altered myogenic programming and failed regeneration
Stem cell reports, 2018Co-Authors: Masakazu Yamamoto, Shahragim Tajbakhsh, Nicholas P Legendre, Arpita A Biswas, Alexander Lawton, Shoko Yamamoto, Gabrielle Kardon, David J GoldhamerAbstract:MyoD and MYF5 are fundamental regulators of skeletal muscle lineage determination in the embryo, and their expression is induced in satellite cells following muscle injury. MyoD and MYF5 are also expressed by satellite cell precursors developmentally, although the relative contribution of historical and injury-induced expression to satellite cell function is unknown. We show that satellite cells lacking both MyoD and MYF5 (double knockout [dKO]) are maintained with aging in uninjured muscle. However, injured muscle fails to regenerate and dKO satellite cell progeny accumulate in damaged muscle but do not undergo muscle differentiation. dKO satellite cell progeny continue to express markers of myoblast identity, although their myogenic programming is labile, as demonstrated by dramatic morphological changes and increased propensity for non-myogenic differentiation. These data demonstrate an absolute requirement for either MyoD or MYF5 in muscle regeneration and indicate that their expression after injury stabilizes myogenic identity and confers the capacity for muscle differentiation.
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mrf4 determines skeletal muscle identity in MYF5 myod double mutant mice
Nature, 2004Co-Authors: Lina Kassarduchossoy, Margaret Buckingham, Didier Rocancourt, Barbara Gayraudmorel, Danielle Gomes, Vasily Shinin, Shahragim TajbakhshAbstract:In vertebrates, skeletal muscle is a model for the acquisition of cell fate from stem cells. Two determination factors of the basic helix-loop-helix myogenic regulatory factor (MRF) family, MYF5 and Myod, are thought to direct this transition because double-mutant mice totally lack skeletal muscle fibres and myoblasts. In the absence of these factors, progenitor cells remain multipotent and can change their fate. Gene targeting studies have revealed hierarchical relationships between these and the other MRF genes, Mrf4 and myogenin, where the latter are regarded as differentiation genes. Here we show, using an allelic series of three MYF5 mutants that differentially affect the expression of the genetically linked Mrf4 gene, that skeletal muscle is present in the new MYF5:Myod double-null mice only when Mrf4 expression is not compromised. This finding contradicts the widely held view that myogenic identity is conferred solely by MYF5 and Myod, and identifies Mrf4 as a determination gene. We revise the epistatic relationship of the MRFs, in which both MYF5 and Mrf4 act upstream of Myod to direct embryonic multipotent cells into the myogenic lineage.
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analysis of a key regulatory region upstream of the MYF5 gene reveals multiple phases of myogenesis orchestrated at each site by a combination of elements dispersed throughout the locus
Development, 2003Co-Authors: Juliette Hadchouel, Shahragim Tajbakhsh, Didier Rocancourt, Philippe Daubas, Lola Bajard, Dennis Summerbell, Jaime J Carvajal, Ted Chang, David G Cox, Peter W J RigbyAbstract:MYF5 is the first myogenic regulatory factor to be expressed in the mouse embryo and it determines the entry of cells into the skeletal muscle programme. A region situated between -58 kb and -48 kb from the gene directs MYF5 transcription at sites where muscles will form. We now show that this region consists of a number of distinct regulatory elements that specifically target sites of myogenesis in the somite, limbs and hypoglossal cord, and also sites of MYF5 transcription in the central nervous system. Deletion of these sequences in the context of the locus shows that elements within the region are essential, and also reveals the combinatorial complexity of the transcriptional regulation of MYF5 . Both within the -58 kb to -48 kb region and elsewhere in the locus, multiple sequences are present that direct transcription in subdomains of a single site during development, thus revealing distinct phases of myogenesis when subpopulations of progenitor cells enter the programme of skeletal muscle differentiation.
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kinetics of myoblast proliferation show that resident satellite cells are competent to fully regenerate skeletal muscle fibers
Experimental Cell Research, 2002Co-Authors: Peter S. Zammit, L Heslop, Shahragim Tajbakhsh, Margaret Buckingham, Valerie Hudon, David J Rosenblatt, Jonathan R Beauchamp, Terence A PartridgeAbstract:The satellite cell compartment provides skeletal muscle with a remarkable capacity for regeneration. Here, we have used isolated myofibers to investigate the activation and proliferative potential of satellite cells. We have previously shown that satellite cells are heterogeneous: the majority express MYF5 and M-cadherin protein, presumably reflecting commitment to myogenesis, while a minority is negative for both. Although MyoD is rarely detected in quiescent satellite cells, over 98% of satellite cells contain MyoD within 24 h of stimulation. Significantly, MyoD is only observed in cells that are already expressing MYF5. In contrast, a minority population does not activate by the criteria of MYF5 or MyoD expression. Following the synchronous activation of the myogenic regulatory factor+ve satellite cells, their daughter myoblasts proliferate with a doubling time of ∼17 h, irrespective of the fiber type (type I, IIa, or IIb) from which they originate. Although fast myofibers have fewer associated satellite cells than slow, and accordingly produce fewer myoblasts, each myofiber phenotype is associated with a complement of satellite cells that has sufficient proliferative potential to fully regenerate the parent myofiber within 4 days. This time course is similar to that observed in vivo following acute injury and indicates that cells other than satellite cells are not required for complete myofiber regeneration.
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expression of cd34 and MYF5 defines the majority of quiescent adult skeletal muscle satellite cells
Journal of Cell Biology, 2000Co-Authors: Jonathan R Beauchamp, L Heslop, Shahragim Tajbakhsh, Terence A Partridge, Robert G. Kelly, Margaret Buckingham, Anton Wernig, Peter S. ZammitAbstract:Skeletal muscle is one of a several adult post-mitotic tissues that retain the capacity to regenerate. This relies on a population of quiescent precursors, termed satellite cells. Here we describe two novel markers of quiescent satellite cells: CD34, an established marker of hematopoietic stem cells, and MYF5, the earliest marker of myogenic commitment. CD34+ve myoblasts can be detected in proliferating C2C12 cultures. In differentiating cultures, CD34+ve cells do not fuse into myotubes, nor express MyoD. Using isolated myofibers as a model of synchronous precursor cell activation, we show that quiescent satellite cells express CD34. An early feature of their activation is alternate splicing followed by complete transcriptional shutdown of CD34. This data implicates CD34 in the maintenance of satellite cell quiescence. In heterozygous MYF5nlacZ/+ mice, all CD34+ve satellite cells also express β-galactosidase, a marker of activation of MYF5, showing that quiescent satellite cells are committed to myogenesis. All such cells are positive for the accepted satellite cell marker, M-cadherin. We also show that satellite cells can be identified on isolated myofibers of the myosin light chain 3F-nlacZ-2E mouse as those that do not express the transgene. The numbers of satellite cells detected in this way are significantly greater than those identified by the other three markers. We conclude that the expression of CD34, MYF5, and M-cadherin defines quiescent, committed precursors and speculate that the CD34−ve, MYF5−ve minority may be involved in maintaining the lineage-committed majority.
Margaret Buckingham - One of the best experts on this subject based on the ideXlab platform.
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direct molecular regulation of the myogenic determination gene MYF5 by pax3 with modulation by six1 4 factors is exemplified by the 111 kb MYF5 enhancer
Developmental Biology, 2013Co-Authors: P Daubas, Margaret BuckinghamAbstract:Abstract The MYF5 gene plays an important role in myogenic determination during mouse embryo development. Multiple genomic regions of the Mrf4–MYF5 locus have been characterised as enhancer sequences responsible for the complex spatiotemporal expression of the MYF5 gene at the onset of myogenesis. These include an enhancer sequence, located at −111 kb upstream of the MYF5 transcription start site, which is responsible of MYF5 activation in ventral somitic domains (Ribas et al., 2011. Dev. Biol. 355, 372–380). We show that the −111 kb-MYF5 enhancer also directs transgene expression in some limb muscles, and is active at foetal as well as embryonic stages. We have carried out further characterisation of the regulation of this enhancer and show that the paired-box Pax3 transcription factor binds to it in vitro as in vivo, and that Pax binding sites are essential for its activity. This requirement is independent of the previously reported regulation by TEAD transcription factors. Six1/4 which, like Pax3, are important upstream regulators of myogenesis, also bind in vivo to sites in the −111 kb-MYF5 enhancer and modulate its activity. The −111 kb-MYF5 enhancer therefore shares common functional characteristics with another MYF5 regulatory sequence, the hypaxial and limb 145 bp-MYF5 enhancer, both being directly regulated in vivo by Pax3 and Six1/4 proteins. However, in the case of the −111 kb-MYF5 enhancer, Six has less effect and we conclude that Pax regulation plays a major role in controlling this aspect of the MYF5 gene expression at the onset of myogenesis in the embryo.
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a pax3 dmrt2 MYF5 regulatory cascade functions at the onset of myogenesis
PLOS Genetics, 2010Co-Authors: Takahiko Sato, Didier Rocancourt, Luis Marques, Solveig Thorsteinsdottir, Margaret BuckinghamAbstract:All skeletal muscle progenitor cells in the body derive from the dermomyotome, the dorsal epithelial domain of developing somites. These multipotent stem cells express Pax3, and this expression is maintained in the myogenic lineage where Pax3 plays an important role. Identification of Pax3 targets is therefore important for understanding the mechanisms that underlie the onset of myogenesis. In a microarray screen of Pax3-GFP sorted cells, with analysis on Pax3 gain and loss of function genetic backgrounds, we identify Dmrt2, expressed in the dermomyotome, as a Pax3 target. In vitro gel shift analysis and chromatin immunoprecipitation with in vivo extracts show that Pax3 binds to a conserved 286 bp sequence, situated at -18 kb from Dmrt2. This sequence directs reporter transgene expression to the somite, and this is severely affected when the Pax3 site is mutated in the context of the locus. In Dmrt2 mutant embryos, somite maturation is perturbed and the skeletal muscle of the myotome is abnormal. We now report that the onset of myogenesis is also affected. This depends on activation, in the epaxial dermomyotome, of the myogenic determination gene, MYF5, through its early epaxial enhancer. This sequence contains sites that bind Dmrt2, which belongs to the DM class of DNA-binding proteins. Mutation of these sites compromises activity of the enhancer in transgenic embryos where the reporter transgene is under the control of the MYF5 epaxial enhancer. Transactivation of this site by Dmrt2 is demonstrated in vitro, and conditional overexpression of Dmrt2 in Pax3 expressing cells in the somite confirms the role of this factor in the activation of MYF5. These results reveal a novel genetic network, comprising a Pax3/Dmrt2/MYF5 regulatory cascade that operates in stem cells of the epaxial dermomyotome to initiate skeletal muscle formation.
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the wnt β catenin pathway regulates gli mediated MYF5 expression during somitogenesis
Development, 2006Co-Authors: Ugo Borello, Margaret Buckingham, Lola Bajard, Barbara Berarducci, Paula Murphy, Viviana Buffa, Stefano Piccolo, Giulio CossuAbstract:Canonical Wnt/β-catenin signaling regulates the activation of the myogenic determination gene MYF5 at the onset of myogenesis, but the underlying molecular mechanism is unknown. Here, we report that the Wnt signal is transduced in muscle progenitor cells by at least two Frizzled (Fz) receptors (Fz1 and/or Fz6), through the canonical β-catenin pathway, in the epaxial domain of newly formed somites. We show that MYF5 activation is dramatically reduced by blocking the Wnt/β-catenin pathway in somite progenitor cells, whereas expression of activated β-catenin is sufficient to activate MYF5 in somites but not in the presomitic mesoderm. In addition, we identified Tcf/Lef sequences immediately 5′ to the MYF5 early epaxial enhancer. These sites determine the correct spatiotemporal expression of MYF5 in the epaxial domain of the somite, mediating the synergistic action of the Wnt/β-catenin and the Shh/Gli pathways. Taken together, these results demonstrate that MYF5 is a direct target of Wnt/β-catenin, and that its full activation requires a cooperative interaction between the canonical Wnt and the Shh/Gli pathways in muscle progenitor cells.
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a novel genetic hierarchy functions during hypaxial myogenesis pax3 directly activates MYF5 in muscle progenitor cells in the limb
Genes & Development, 2006Co-Authors: Lola Bajard, Frederic Relaix, Didier Rocancourt, Mounia Lagha, Philippe Daubas, Margaret BuckinghamAbstract:We address the molecular control of myogenesis in progenitor cells derived from the hypaxial somite. Null mutations in Pax3, a key regulator of skeletal muscle formation, lead to cell death in this domain. We have developed a novel allele of Pax3 encoding a Pax3–engrailed fusion protein that acts as a transcriptional repressor. Heterozygote mouse embryos have an attenuated mutant phenotype, with partial conservation of the hypaxial somite and its myogenic derivatives, including some hindlimb muscles. At these sites, expression of MYF5 is compromised, showing that Pax3 acts genetically upstream of this myogenic determination gene. We have characterized a 145-base-pair (bp) regulatory element, at −57.5 kb from MYF5, that directs transgene expression to the mature somite, notably to myogenic cells of the hypaxial domain that form ventral trunk and limb muscles. A Pax3 consensus site in this sequence binds Pax3 in vitro and in vivo. Multimers of the 145-bp sequence direct transgene expression to sites of Pax3 function, and an assay of its activity in the chick embryo shows Pax3 dependence. Mutation of the Pax3 site abolishes all expression controlled by the 145-bp sequence in transgenic mouse embryos. We conclude that Pax3 directly regulates MYF5 in the hypaxial somite and its derivatives.
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mrf4 determines skeletal muscle identity in MYF5 myod double mutant mice
Nature, 2004Co-Authors: Lina Kassarduchossoy, Margaret Buckingham, Didier Rocancourt, Barbara Gayraudmorel, Danielle Gomes, Vasily Shinin, Shahragim TajbakhshAbstract:In vertebrates, skeletal muscle is a model for the acquisition of cell fate from stem cells. Two determination factors of the basic helix-loop-helix myogenic regulatory factor (MRF) family, MYF5 and Myod, are thought to direct this transition because double-mutant mice totally lack skeletal muscle fibres and myoblasts. In the absence of these factors, progenitor cells remain multipotent and can change their fate. Gene targeting studies have revealed hierarchical relationships between these and the other MRF genes, Mrf4 and myogenin, where the latter are regarded as differentiation genes. Here we show, using an allelic series of three MYF5 mutants that differentially affect the expression of the genetically linked Mrf4 gene, that skeletal muscle is present in the new MYF5:Myod double-null mice only when Mrf4 expression is not compromised. This finding contradicts the widely held view that myogenic identity is conferred solely by MYF5 and Myod, and identifies Mrf4 as a determination gene. We revise the epistatic relationship of the MRFs, in which both MYF5 and Mrf4 act upstream of Myod to direct embryonic multipotent cells into the myogenic lineage.
Michael A. Rudnicki - One of the best experts on this subject based on the ideXlab platform.
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carm1 regulates pax7 transcriptional activity through mll1 2 recruitment during asymmetric satellite stem cell divisions
Cell Stem Cell, 2012Co-Authors: Michael A. Rudnicki, Iain W Mckinnell, Yoh Ichi Kawabe, Yu Xin Wang, Mark T BedfordAbstract:Summary In skeletal muscle, asymmetrically dividing satellite stem cells give rise to committed satellite cells that transcribe the myogenic determination factor MYF5, a Pax7-target gene. We identified the arginine methyltransferase Carm1 as a Pax7 interacting protein and found that Carm1 specifically methylates multiple arginines in the N terminus of Pax7. Methylated Pax7 directly binds the C-terminal cleavage forms of the trithorax proteins MLL1/2 resulting in the recruitment of the ASH2L:MLL1/2:WDR5:RBBP5 histone H3K4 methyltransferase complex to regulatory enhancers and the proximal promoter of MYF5 . Finally, Carm1 is required for the induction of de novo MYF5 transcription following asymmetric satellite stem cell divisions. We defined the C-terminal MLL region as a reader domain for the recognition of arginine methylated proteins such as Pax7. Thus, arginine methylation of Pax7 by Carm1 functions as a molecular switch controlling the epigenetic induction of MYF5 during satellite stem cell asymmetric division and entry into the myogenic program.
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polycomb ezh2 controls self renewal and safeguards the transcriptional identity of skeletal muscle stem cells
Genes & Development, 2011Co-Authors: Aster H Juan, Michael A. Rudnicki, Assia Derfoul, Xuesong Feng, James G Ryall, Stefania Dellorso, Alessandra Pasut, Hossein Zare, James M Simone, Vittorio SartorelliAbstract:Satellite cells (SCs) sustain muscle growth and empower adult skeletal muscle with vigorous regenerative abilities. Here, we report that EZH2, the enzymatic subunit of the Polycomb-repressive complex 2 (PRC2), is expressed in both Pax7+/MYF5− stem cells and Pax7+/MYF5+ committed myogenic precursors and is required for homeostasis of the adult SC pool. Mice with conditional ablation of Ezh2 in SCs have fewer muscle postnatal Pax7+ cells and reduced muscle mass and fail to appropriately regenerate. These defects are associated with impaired SC proliferation and derepression of genes expressed in nonmuscle cell lineages. Thus, EZH2 controls self-renewal and proliferation, and maintains an appropriate transcriptional program in SCs.
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the molecular regulation of muscle stem cell function
Cold Spring Harbor Symposia on Quantitative Biology, 2008Co-Authors: Michael A. Rudnicki, Le F Grand, Iain W Mckinnell, Shihuan KuangAbstract:Muscle satellite cells are responsible for the postnatal growth and robust regeneration capacity of adult skeletal muscle. A subset of satellite cells purified from adult skeletal muscle is capable of repopulating the satellite cell pool, suggesting that it has direct therapeutic potential for treating degenerative muscle disease. Satellite cells uniformly express the transcription factor Pax7, and Pax7 is required for satellite cell viability and to give rise to myogenic precursors that express the basic helix-loop-helic (bHLH) transcription factors MYF5 and MyoD. Pax7 activates expression of target genes such as MYF5 and MyoD through recruitment of the Wdr5/Ash2L/MLL2 histone methyltransferase complex. Extensive genetic analysis has revealed that MYF5 and MyoD are required for myogenic determination, whereas myogenin and MRF4 have roles in terminal differentiation. Using a MYF5-Cre knockin allele and an R26R-YFP Cre reporter, we observed that in vivo about 10% of satellite cells only express Pax7 and have never expressed MYF5. Moreover, we found that Pax7(+)/MYF5(-) satellite cells give rise to Pax7(+)/MYF5(+) satellite cells through basal-apical asymmetric cell divisions. Therefore, satellite cells in skeletal muscle are a heterogeneous population composed of satellite stem cells (Pax7(+)/MYF5(-)) and satellite myogenic cells (Pax7(+)/MYF5(+)). Evidence is accumulating that indicates that satellite stem cells represent a true stem cell reservoir, and targeting mechanisms that regulate their function represents an important therapeutic strategy for the treatment of neuromuscular disease.
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myod induces myogenic differentiation through cooperation of its nh2 and cooh terminal regions
Journal of Cell Biology, 2005Co-Authors: Jeff Ishibashi, Michael A. Rudnicki, Atsushi Asakura, Robert L. S. PerryAbstract:MyoD and MYF5 are basic helix-loop-helix transcription factors that play key but redundant roles in specifying myogenic progenitors during embryogenesis. However, there are functional differences between the two transcription factors that impact myoblast proliferation and differentiation. Target gene activation could be one such difference. We have used microarray and polymerase chain reaction approaches to measure the induction of muscle gene expression by MyoD and MYF5 in an in vitro model. In proliferating cells, MyoD and MYF5 function very similarly to activate the expression of likely growth phase target genes such as L-myc, m-cadherin, Mcpt8, Runx1, Spp1, Six1, IGFBP5, and Chrnβ1. MyoD, however, is strikingly more effective than MYF5 at inducing differentiation-phase target genes. This distinction between MyoD and MYF5 results from a novel and unanticipated cooperation between the MyoD NH2- and COOH-terminal regions. Together, these results support the notion that MYF5 functions toward myoblast proliferation, whereas MyoD prepares myoblasts for efficient differentiation.
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activation of fgf4 gene expression in the myotomes is regulated by myogenic bhlh factors and by sonic hedgehog
Developmental Biology, 2000Co-Authors: Diego Fraidenraich, Michael A. Rudnicki, Akiyo Iwahori, Claudio BasilicoAbstract:Abstract The Fgf4 gene encodes an important signaling molecule which is expressed in specific developmental stages, including the inner cell mass of the blastocyst, the myotomes, and the limb bud apical ectodermal ridge (AER). Using a transgenic approach, we previously identified overlapping but distinct enhancer elements in the Fgf4 3′ untranslated region necessary and sufficient for myotome and AER expression. Here we have investigated the hypothesis that Fgf4 is a target of myogenic bHLH factors. We show by mutational analysis that a conserved E box located in the Fgf4 myotome enhancer is required for Fgf4-lacZ expression in the myotomes. A DNA probe containing the E box binds MYF5, MYOD, and bHLH-like activities from nuclear extracts of differentiating C2-7 myoblast cells, and both MYF5 and MYOD can activate gene expression of reporter plasmids containing the E-box element. Analyses of MYF5 and MyoD knockout mice harboring Fgf4-lacZ transgenes show that MYF5 is required for Fgf4 expression in the myotomes, while MyoD is not, but MyoD can sustain Fgf4 expression in the ventral myotomes in the absence of MYF5. Sonic hedgehog (Shh) signaling has been shown to have an essential inductive function in the expression of MYF5 and MyoD in the epaxial myotomes, but not in the hypaxial myotomes. We show here that expression of an Fgf4-lacZ transgene in Shh−/− embryos is suppressed not only in the epaxial but also in the hypaxial myotomes, while it is maintained in the AER. This suggests that Shh mediates Fgf4 activation in the myotomes through mechanisms independent of its role in the activation of myogenic factors. Thus, a cascade of events, involving Shh and bHLH factors, is responsible for activating Fgf4 expression in the myotomes in a spatial- and temporal-specific manner.
Peter W J Rigby - One of the best experts on this subject based on the ideXlab platform.
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Musculin and TCF21 coordinate the maintenance of myogenic regulatory factor expression levels during mouse craniofacial development
Development (Cambridge England), 2012Co-Authors: Natalia Moncaut, Peter W J Rigby, Joe W. Cross, Christine Siligan, Annette Keith, Kevin Taylor, Jaime J CarvajalAbstract:The specification of the skeletal muscle lineage during craniofacial development is dependent on the activity of MYF5 and MYOD, two members of the myogenic regulatory factor family. In the absence of MYF5 or MYOD there is not an overt muscle phenotype, whereas in the double MYF5;MyoD knockout branchiomeric myogenic precursors fail to be specified and skeletal muscle is not formed. The transcriptional regulation of MYF5 is controlled by a multitude of regulatory elements acting at different times and anatomical locations, with at least five operating in the branchial arches. By contrast, only two enhancers have been implicated in the regulation of MyoD. In this work, we characterize an enhancer element that drives MYF5 expression in the branchial arches from 9.5 days post-coitum and show that its activity in the context of the entire locus is dependent on two highly conserved E-boxes. These binding sites are required in a subset of MYF5-expressing cells including both progenitors and those which have entered the myogenic pathway. The correct levels of expression of MYF5 and MyoD result from activation by musculin and TCF21 through direct binding to specific enhancers. Consistent with this, we show that in the absence of musculin the timing of activation of MYF5 and MyoD is not affected but the expression levels are significantly reduced. Importantly, normal levels of MYF5 expression are restored at later stages, which might explain the absence of particular muscles in the Msc;Tcf21 double-knockout mice.
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the myogenic factor MYF5 supports efficient skeletal muscle regeneration by enabling transient myoblast amplification
Stem Cells, 2007Co-Authors: Svetlana Ustanina, Peter W J Rigby, Jaime J Carvajal, Thomas BraunAbstract:The myogenic factor MYF5 defines the onset of myogenesis in mammals during development. Mice lacking both MYF5 and MyoD fail to form myoblasts and are characterized by a complete absence of skeletal muscle at birth. To investigate the function of MYF5 in adult skeletal muscle, we generated MYF5 and mdx compound mutants, which are characterized by constant regeneration. Double mutant mice show an increase of dystrophic changes in the musculature, although these mice were viable and the degree of myopathy was modest. MYF5 mutant muscles show a small decrease in the number of muscle satellite cells, which was Within the range of physiological variations. We also observed a significant delay in the regeneration of MYF5 deficient skeletal muscles after injury. Interestingly, MYF5 deficient skeletal muscles were able to even out this flaw during the course of regeneration, generating intact muscles 4 weeks after injury. Although we did not detect a striking reduction of MyoD positive activated myoblasts or of MYF5-LacZ positive cells in regenerating muscles, a clear decrease in the proliferation rate of satellite cell-derived myoblasts was apparent in satellite cell-derived cultures. The reduction of the proliferation rate of MYF5 mutant myoblasts was also reflected by a delayed transition from proliferation to differentiation, resulting in a reduced number of myotube nuclei after 6 and 7 days of culture. We reason that MYF5 supports efficient skeletal muscle regeneration by enabling transient myoblast amplification.
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analysis of a key regulatory region upstream of the MYF5 gene reveals multiple phases of myogenesis orchestrated at each site by a combination of elements dispersed throughout the locus
Development, 2003Co-Authors: Juliette Hadchouel, Shahragim Tajbakhsh, Didier Rocancourt, Philippe Daubas, Lola Bajard, Dennis Summerbell, Jaime J Carvajal, Ted Chang, David G Cox, Peter W J RigbyAbstract:MYF5 is the first myogenic regulatory factor to be expressed in the mouse embryo and it determines the entry of cells into the skeletal muscle programme. A region situated between -58 kb and -48 kb from the gene directs MYF5 transcription at sites where muscles will form. We now show that this region consists of a number of distinct regulatory elements that specifically target sites of myogenesis in the somite, limbs and hypoglossal cord, and also sites of MYF5 transcription in the central nervous system. Deletion of these sequences in the context of the locus shows that elements within the region are essential, and also reveals the combinatorial complexity of the transcriptional regulation of MYF5 . Both within the -58 kb to -48 kb region and elsewhere in the locus, multiple sequences are present that direct transcription in subdomains of a single site during development, thus revealing distinct phases of myogenesis when subpopulations of progenitor cells enter the programme of skeletal muscle differentiation.
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Expression of the myogenic regulatory factor Mrf4 precedes or is contemporaneous with that of MYF5 in the somitic bud.
Mechanisms of development, 2002Co-Authors: Dennis Summerbell, Chandrika Halai, Peter W J RigbyAbstract:The development of skeletal muscle in vertebrate embryos is controlled by a transcriptional cascade involving the four myogenic regulatory factors. In the somites of the mouse embryo the order of expression is thought to be MYF5, Myogenin, Mrf4 and MyoD. We have re-examined the expression pattern of Mrf4 and show that in the hypaxial domain of thoracic somites (the somitic bud) Mrf4 expression precedes or is contemporaneous with that of MYF5, suggesting that this transcription factor plays a hitherto unsuspected role in myogenesis.
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a bac transgenic analysis of the mrf4 MYF5 locus reveals interdigitated elements that control activation and maintenance of gene expression during muscle development
Development, 2001Co-Authors: Jaime J Carvajal, Dennis Summerbell, Peter W J RigbyAbstract:The muscle-specific transcription factors MYF5 and Mrf4 are two of the four myogenic regulatory factors involved in the transcriptional cascade responsible for skeletal myogenesis in the vertebrate embryo. MYF5 is the first of these four genes to be expressed in the mouse. We have previously described discrete enhancers that drive MYF5 expression in epaxial and hypaxial somites, branchial arches and central nervous system, and argued that additional elements are required for proper expression (Summerbell, D., Ashby, P. R., Coutelle, O., Cox, D., Yee, S. P. and Rigby, P. W. J. (2000) Development 127, 3745-3757). We have now investigated the transcriptional regulation of both MYF5 and Mrf4 using bacterial artificial chromosome transgenesis. We show that a clone containing MYF5 and 140 kb of upstream sequences is sufficient to recapitulate the known expression patterns of both genes. Our results confirm and reinforce the conclusion of our earlier studies, that MYF5 expression is regulated differently in each of a considerable number of populations of muscle progenitors, and they begin to illuminate the evolutionary origins of this complex regulation. We further show that separate elements are involved in the activation and maintenance of expression in the various precursor populations, reflecting the diversity of the signals that control myogenesis. Mrf4 expression requires at least four elements, one of which may be shared with MYF5, providing a possible explanation for the linkage of these genes throughout vertebrate phylogeny. Further complexity is revealed by the demonstration that elements which control Mrf4 and MYF5 are embedded in an unrelated neighbouring gene.
Peter S. Zammit - One of the best experts on this subject based on the ideXlab platform.
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Function of the myogenic regulatory factors MYF5, MyoD, Myogenin and MRF4 in skeletal muscle, satellite cells and regenerative myogenesis.
Seminars in cell & developmental biology, 2017Co-Authors: Peter S. ZammitAbstract:Discovery of the myogenic regulatory factor family of transcription factors MYF5, MYOD, Myogenin and MRF4 was a seminal step in understanding specification of the skeletal muscle lineage and control of myogenic differentiation during development. These factors are also involved in specification of the muscle satellite cell lineage, which becomes the resident stem cell compartment inadult skeletal muscle. While MYF5, MYOD, Myogenin and MRF4 have subtle roles in mature muscle, they again play a crucial role in directing satellite cell function to regenerate skeletal muscle: linking the genetic control of developmental and regenerative myogenesis. Here, I review the role of the myogenic regulatory factors in developing and mature skeletal muscle, satellite cell specification and muscle regeneration.
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kinetics of myoblast proliferation show that resident satellite cells are competent to fully regenerate skeletal muscle fibers
Experimental Cell Research, 2002Co-Authors: Peter S. Zammit, L Heslop, Shahragim Tajbakhsh, Margaret Buckingham, Valerie Hudon, David J Rosenblatt, Jonathan R Beauchamp, Terence A PartridgeAbstract:The satellite cell compartment provides skeletal muscle with a remarkable capacity for regeneration. Here, we have used isolated myofibers to investigate the activation and proliferative potential of satellite cells. We have previously shown that satellite cells are heterogeneous: the majority express MYF5 and M-cadherin protein, presumably reflecting commitment to myogenesis, while a minority is negative for both. Although MyoD is rarely detected in quiescent satellite cells, over 98% of satellite cells contain MyoD within 24 h of stimulation. Significantly, MyoD is only observed in cells that are already expressing MYF5. In contrast, a minority population does not activate by the criteria of MYF5 or MyoD expression. Following the synchronous activation of the myogenic regulatory factor+ve satellite cells, their daughter myoblasts proliferate with a doubling time of ∼17 h, irrespective of the fiber type (type I, IIa, or IIb) from which they originate. Although fast myofibers have fewer associated satellite cells than slow, and accordingly produce fewer myoblasts, each myofiber phenotype is associated with a complement of satellite cells that has sufficient proliferative potential to fully regenerate the parent myofiber within 4 days. This time course is similar to that observed in vivo following acute injury and indicates that cells other than satellite cells are not required for complete myofiber regeneration.
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expression of cd34 and MYF5 defines the majority of quiescent adult skeletal muscle satellite cells
Journal of Cell Biology, 2000Co-Authors: Jonathan R Beauchamp, L Heslop, Shahragim Tajbakhsh, Terence A Partridge, Robert G. Kelly, Margaret Buckingham, Anton Wernig, Peter S. ZammitAbstract:Skeletal muscle is one of a several adult post-mitotic tissues that retain the capacity to regenerate. This relies on a population of quiescent precursors, termed satellite cells. Here we describe two novel markers of quiescent satellite cells: CD34, an established marker of hematopoietic stem cells, and MYF5, the earliest marker of myogenic commitment. CD34+ve myoblasts can be detected in proliferating C2C12 cultures. In differentiating cultures, CD34+ve cells do not fuse into myotubes, nor express MyoD. Using isolated myofibers as a model of synchronous precursor cell activation, we show that quiescent satellite cells express CD34. An early feature of their activation is alternate splicing followed by complete transcriptional shutdown of CD34. This data implicates CD34 in the maintenance of satellite cell quiescence. In heterozygous MYF5nlacZ/+ mice, all CD34+ve satellite cells also express β-galactosidase, a marker of activation of MYF5, showing that quiescent satellite cells are committed to myogenesis. All such cells are positive for the accepted satellite cell marker, M-cadherin. We also show that satellite cells can be identified on isolated myofibers of the myosin light chain 3F-nlacZ-2E mouse as those that do not express the transgene. The numbers of satellite cells detected in this way are significantly greater than those identified by the other three markers. We conclude that the expression of CD34, MYF5, and M-cadherin defines quiescent, committed precursors and speculate that the CD34−ve, MYF5−ve minority may be involved in maintaining the lineage-committed majority.
