Journal of Developmental Biology Review Bu-M-P-ing Iron: How BMP Signaling Regulates Muscle Growth and Regeneration 1,2, 1,2, 1,2,3,4,5, Matthew J Borok y, Despoina Mademtzoglou y and Frederic Relaix * 1 Inserm, IMRB U955-E10, 94010 Créteil, France; [email protected] (M.J.B.); [email protected] (D.M.) 2 Faculté de santé, Université Paris Est, 94000 Creteil, France 3 Ecole Nationale Veterinaire d’Alfort, 94700 Maison Alfort, France 4 Etablissement Français du Sang, 94017 Créteil, France 5 APHP, Hopitaux Universitaires Henri Mondor, DHU Pepsy & Centre de Référence des Maladies Neuromusculaires GNMH, 94000 Créteil, France * Correspondence: [email protected]; Tel.: +33-149-813-940 These authors contributed equally to this work. y Received: 10 January 2020; Accepted: 7 February 2020; Published: 11 February 2020 Abstract: The bone morphogenetic protein (BMP) pathway is best known for its role in promoting bone formation, however it has been shown to play important roles in both development and regeneration of many different tissues. Recent work has shown that the BMP proteins have a number of functions in skeletal muscle, from embryonic to postnatal development. Furthermore, complementary studies have recently demonstrated that specific components of the pathway are required for efficient muscle regeneration. Keywords: development; regeneration; TGFβ; stem cells; satellite cells 1. Introduction As its name implies, the bone morphogenetic protein (BMP) signaling pathway was first identified as a key regulator of bone formation. Urist and colleagues isolated proteins from rabbit bone and then used them to induce bone formation in vitro or in vivo in the rat [1]. Later work showed that these proteins were highly conserved during evolution and most species had a number of paralogues [2–4]. Mapping of the mouse short ear gene, essential for skeletal patterning and bone repair, to the Bmp5 locus, was the first demonstration of the importance of this pathway in mice [5]. In contrast to the relatively mild phenotype of Bmp5 deletions in short-eared mice, deletion of either Bmp4 or Bmp2 and their receptors Bmpr1a and Bmpr2 caused early embryonic lethality, with severe defects in mesoderm formation, extraembryonic tissues and cardiac development [6–9] (Table A1). In addition to demonstrating that certain BMP genes are essential for development, this work implicated this signaling pathway to play important roles in multiple organ systems. 2. Signaling Mechanism After decades of biochemical work, many components of the BMP signaling pathway and their mode of action have been described. Signaling activation requires two different receptors, first discovered by experiments with a human cell line [10]. Today the pathway consists of at least 20 different ligands, four type 1 receptors, and three type 2 receptors, varying combinations of which give distinct responses [11]. After binding of the BMP ligand, the receptors phosphorylate Smad cofactors, allowing an interaction with Smad4 and translocation of the Smad complex into the nucleus (Figure1)[ 12–14]. The BMP-specific cofactors are Smad1, Smad5, and Smad9. Surprisingly, while single J. Dev. Biol. 2020, 8, 4; doi:10.3390/jdb8010004 www.mdpi.com/journal/jdb J. Dev. Biol. 2020, 8, 4 2 of 14 J.knockouts Dev. Biol. 2020 for, 8 Smad1, 4 and Smad5 are embryonic lethal, Smad9 is dispensable for development [152 –of19 14]. The interaction between Smad1/5/8 and Smad4 is BMP-specific, while TGFβ signaling induces an developmentinteraction between [15–19]. Smad2 The interaction/3 and Smad4 between [20–22 ].Sm Furthermore,ad1/5/8 and Smad1Smad4 andis BMP-specific, Smad4 have homologouswhile TGFβ signalingtranscriptional induces activation an interaction domains between and their Smad2/3 nuclear import and Smad4 leads to [20–22]. transcriptional Furthermore, competence Smad1 [14 and,23]. Smad4While therehave ishomologous some overlap transcriptional in the two pathways, activation the domains specific components,and their nuclear including import extracellular leads to transcriptionaland intracellular competence inhibitors, [14,23]. receptors, While and there Smads isdistinguish some overlap BMP infrom the two TGF pathways,β signaling, the leading specific to components,distinct outcomes including and specific extracellular developmental and intracellu consequenceslar inhibitors, [24,25 ].receptors, The regulation and Smads of skeletal distinguish muscle BMPgrowth from and TGF regenerationβ signaling, by leading the TGF toβ distinctpathway outcomes falls beyond and specific the scope developmental of this review; consequences readers may [24,25].refer to previousThe regulation works [of26 ,27skeletal]. muscle growth and regeneration by the TGFβ pathway falls beyond the scope of this review; readers may refer to previous works [26,27]. Figure 1. The BMP signaling pathway. Binding of the ligand to the type II receptor recruits the type I Figure 1. The BMP signaling pathway. Binding of the ligand to the type II receptor recruits the type I receptor and activates Smad 1/5/8 by phosphorylation. The later engages Smad4, which is shared with receptorthe TGFb and pathway, activates and Smad the complex 1/5/8 by translocates phosphorylat to theion. nucleus The later to regulate engages transcription Smad4, which of target is shared genes with(e.g. Id).the TheTGFb pathway pathway, is blockedand the by complex several inhibitorstranslocates (e.g. to Noggin)the nucleus as well to regulate as the inhibitory transcription Smad6. of target genes (e.g. Id). The pathway is blocked by several inhibitors (e.g. Noggin) as well as the inhibitoryThe most Smad6. common BMP targets are Id and Msx genes. The canonical downstream target of BMP signaling is Id1, first identified in osteoblast-like cells [28]. Later work in embryonic stem cells and a numberThe ofmost diff commonerent cell BMP lines targets confirmed are Id this and target Msx gene and alsos. The identified canonical Id2, downstream Id3, Msx1, target and Msx2of BMP as signalingdirect targets is Id1, of first the pathwayidentified [29 in]. osteoblast-like The Drosophila cellshomologue [28]. Later of work the Smad in embryonic proteins, stem Mad, cells binds and to a numberGCCG motif of different in target cell genes lines [ 30confirmed]. When thisthis sitetarget is presentand also in identified a multimerized Id2, Id3, form, Msx1, it is and suffi Msx2cient as to directbind Smad targets proteins of the andpathway to confer [29]. BMP-specificThe Drosophila activation homologue in several of the mammalianSmad proteins, cell Mad, lines [binds31]. to a GCCGSeveral motif inhibitorsin target genes of the [30]. pathway When have this beensite is identified. present in Work a multimerized in Xenopus identifiedform, it is Smad6sufficient as anto bindinhibitor Smad of proteins the pathway, and to which confer functions BMP-specific by blocking activation Smad4 in several association mammalian with Smad1 cell lines (Figure [31].1)[ 32]. A numberSeveral of inhibitors secreted of BMP the pathway inhibitors have have been also identified. been described. Work in XenopusXenopus identifiedexplant Smad6 experiments as an inhibitordemonstrated of the that pathway, Bmp4 andwhich noggin functions have by opposing blocking eff Smad4ects on embryonicassociation patterningwith Smad1 [33 (Figure]. Biochemical 1) [32]. Aexperiments number of demonstrated secreted BMP that inhibitors noggin ishave able toalso bind been to BMPdescribed. ligands, Xenopus rendering explant them experiments incapable of demonstratedbinding to their that receptors Bmp4 [34and]. Related noggin antagonists have opposing chordin effects and gremlin, on embr functionyonic inpatterning the same [33]. way Biochemical(Figure1, Table experiments A1)[35–37 demo]. nstrated that noggin is able to bind to BMP ligands, rendering them incapable of binding to their receptors [34]. Related antagonists chordin and gremlin, function in the same3. Role way in (Figure Skeletal 1, Muscle Table A1) [35–37]. Experiments with rodent myogenic cell lines were the first to demonstrate an inhibitory role for 3. Role in Skeletal Muscle BMP signaling on muscle differentiation (Figure2)[ 38,39]. The addition of BMP ligands to the myogenic L6 andExperiments C2C12 cell with lines, rodent as well myogenic as primary cell rat lines cell were cultures, the first inhibited to demo thenstrate formation an inhibitory of myotubes role and for BMPthe expression signaling of on the muscle myogenic differentiation differentiation (Figure genes 2) myogenin, [38,39]. The MyoD, addition and Myf5, of BMP while ligands it promoted to the a myogenic L6 and C2C12 cell lines, as well as primary rat cell cultures, inhibited the formation of myotubes and the expression of the myogenic differentiation genes myogenin, MyoD, and Myf5, while it promoted a concomitant trans-differentiation into osteogenic fate [40–50]. Of note, Bmp2-activated muscle tissue was as efficient as autologous bone grafting in the treatment of large J. Dev. Biol. 2020, 8, 4 3 of 14 concomitantJ. Dev. Biol. trans-di2020, 8, 4 fferentiation into osteogenic fate [40–50]. Of note, Bmp2-activated muscle3 of 14 tissue was as efficient as autologous bone grafting in the treatment of large bone defects [51]. Furthermore, the implantationbone defects of BMP2-soaked [51]. Furthermore, beads the in implantation rat calf muscle of BMP2-soaked resulted in osteogenic beads in rat di calffferentiation muscle resulted [52]. in On the contrary,osteogenic culture differentiation of myogenic cells[52]. On in the