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1.
Genes Dev ; 33(9-10): 524-535, 2019 05 01.
Article in English | MEDLINE | ID: mdl-30862660

ABSTRACT

The balance between proliferation and differentiation of muscle stem cells is tightly controlled, ensuring the maintenance of a cellular pool needed for muscle growth and repair. We demonstrate here that the transcriptional regulator Hes1 controls the balance between proliferation and differentiation of activated muscle stem cells in both developing and regenerating muscle. We observed that Hes1 is expressed in an oscillatory manner in activated stem cells where it drives the oscillatory expression of MyoD. MyoD expression oscillates in activated muscle stem cells from postnatal and adult muscle under various conditions: when the stem cells are dispersed in culture, when they remain associated with single muscle fibers, or when they reside in muscle biopsies. Unstable MyoD oscillations and long periods of sustained MyoD expression are observed in differentiating cells. Ablation of the Hes1 oscillator in stem cells interfered with stable MyoD oscillations and led to prolonged periods of sustained MyoD expression, resulting in increased differentiation propensity. This interfered with the maintenance of activated muscle stem cells, and impaired muscle growth and repair. We conclude that oscillatory MyoD expression allows the cells to remain in an undifferentiated and proliferative state and is required for amplification of the activated stem cell pool.


Subject(s)
Gene Expression Regulation, Developmental/genetics , MyoD Protein/metabolism , Stem Cells/cytology , Stem Cells/metabolism , Transcription Factor HES-1/metabolism , Animals , Cells, Cultured , Mice , MyoD Protein/genetics , Receptors, Notch/metabolism , Signal Transduction , Transcription Factor HES-1/genetics
2.
Dev Biol ; 395(2): 307-16, 2014 Nov 15.
Article in English | MEDLINE | ID: mdl-25220152

ABSTRACT

Craniofacial and trunk skeletal muscles are evolutionarily distinct and derive from cranial and somitic mesoderm, respectively. Different regulatory hierarchies act upstream of myogenic regulatory factors in cranial and somitic mesoderm, but the same core regulatory network - MyoD, Myf5 and Mrf4 - executes the myogenic differentiation program. Notch signaling controls self-renewal of myogenic progenitors as well as satellite cell homing during formation of trunk muscle, but its role in craniofacial muscles has been little investigated. We show here that the pool of myogenic progenitor cells in craniofacial muscle of Dll1(LacZ/Ki) mutant mice is depleted in early fetal development, which is accompanied by a major deficit in muscle growth. At the expense of progenitor cells, supernumerary differentiating myoblasts appear transiently and these express MyoD. The progenitor pool in craniofacial muscle of Dll1(LacZ/Ki) mutants is largely rescued by an additional mutation of MyoD. We conclude from this that Notch exerts its decisive role in craniofacial myogenesis by repression of MyoD. This function is similar to the one previously observed in trunk myogenesis, and is thus conserved in cranial and trunk muscle. However, in cranial mesoderm-derived progenitors, Notch signaling is not required for Pax7 expression and impinges little on the homing of satellite cells. Thus, Dll1 functions in satellite cell homing and Pax7 expression diverge in cranial- and somite-derived muscle.


Subject(s)
Facial Muscles/embryology , Intercellular Signaling Peptides and Proteins/metabolism , Morphogenesis/physiology , Muscle, Skeletal/embryology , Signal Transduction/physiology , Torso/embryology , Animals , Calcium-Binding Proteins , DNA Primers/genetics , Flow Cytometry , Immunohistochemistry , In Situ Hybridization , Intercellular Signaling Peptides and Proteins/genetics , Mice , Mice, Mutant Strains , PAX7 Transcription Factor/metabolism , Real-Time Polymerase Chain Reaction , Receptors, Notch/metabolism , Satellite Cells, Skeletal Muscle/metabolism
3.
Development ; 141(14): 2780-90, 2014 Jul.
Article in English | MEDLINE | ID: mdl-25005473

ABSTRACT

A central question in development is to define how the equilibrium between cell proliferation and differentiation is temporally and spatially regulated during tissue formation. Here, we address how interactions between cyclin-dependent kinase inhibitors essential for myogenic growth arrest (p21(cip1) and p57(kip2)), the Notch pathway and myogenic regulatory factors (MRFs) orchestrate the proliferation, specification and differentiation of muscle progenitor cells. We first show that cell cycle exit and myogenic differentiation can be uncoupled. In addition, we establish that skeletal muscle progenitor cells require Notch signaling to maintain their cycling status. Using several mouse models combined with ex vivo studies, we demonstrate that Notch signaling is required to repress p21(cip1) and p57(kip2) expression in muscle progenitor cells. Finally, we identify a muscle-specific regulatory element of p57(kip2) directly activated by MRFs in myoblasts but repressed by the Notch targets Hes1/Hey1 in progenitor cells. We propose a molecular mechanism whereby information provided by Hes/Hey downstream of Notch as well as MRF activities are integrated at the level of the p57(kip2) enhancer to regulate the decision between progenitor cell maintenance and muscle differentiation.


Subject(s)
Basic Helix-Loop-Helix Transcription Factors/metabolism , Cell Cycle Proteins/metabolism , Cyclin-Dependent Kinase Inhibitor p57/metabolism , Homeodomain Proteins/metabolism , Muscle, Skeletal/growth & development , MyoD Protein/metabolism , Myogenic Regulatory Factor 5/metabolism , Receptors, Notch/metabolism , Animals , Cell Cycle Checkpoints , Cell Differentiation , Cyclin-Dependent Kinase Inhibitor p21/metabolism , Enhancer Elements, Genetic/genetics , Extremities/embryology , Gene Expression Regulation, Developmental , Immunoglobulin J Recombination Signal Sequence-Binding Protein/metabolism , Mice , Mice, Transgenic , Muscle Development , Muscle, Skeletal/cytology , Muscle, Skeletal/embryology , Muscle, Skeletal/metabolism , MyoD Protein/genetics , Myoblasts/cytology , Myoblasts/metabolism , Organ Specificity , PAX7 Transcription Factor/metabolism , Signal Transduction , Stem Cells/cytology , Stem Cells/metabolism , Transcription Factor HES-1
4.
J Neurosci ; 33(17): 7299-307, 2013 Apr 24.
Article in English | MEDLINE | ID: mdl-23616538

ABSTRACT

Inhibitory interneurons of the spinal dorsal horn play critical roles in the processing of noxious and innocuous sensory information. They form a family of morphologically and functionally diverse neurons that likely fall into distinct subtypes. Traditional classifications rely mainly on differences in dendritic tree morphology and firing patterns. Although useful, these markers are not comprehensive and cannot be used to drive specific genetic manipulations targeted at defined subsets of neurons. Here, we have used genome-wide expression profiling of spinal dorsal horns of wild-type mice and of two strains of transcription factor-deficient mice (Ptf1a(-/-) and Ascl1/Mash1(-/-) mice) to identify new genetic markers for specific subsets of dorsal horn inhibitory interneurons. Ptf1a(-/-) mice lack all inhibitory interneurons in the dorsal horn, whereas only the late-born inhibitory interneurons are missing in Ascl1(-/-) mice. We found 30 genes that were significantly downregulated in the dorsal horn of Ptf1a(-/-) mice. Twenty-one of those also showed reduced expression in Ascl1(-/-) mice. In situ hybridization analyses of all 30 genes identified four genes with primarily non-overlapping expression patterns in the dorsal horn. Three genes, pDyn coding the neuropeptide dynorphin, Kcnip2 encoding a potassium channel associated protein, and the nuclear receptor encoding gene Rorb, were expressed in Ascl1-dependent subpopulations of the superficial dorsal horn. The fourth gene, Tfap2b, encoding a transcription factor, is expressed mainly in a Ascl1-independent subpopulation of the deep dorsal horn. Functional experiments in isolated spinal cords showed that the Ascl1-dependent inhibitory interneurons are key players of nociceptive reflex plasticity.


Subject(s)
Basic Helix-Loop-Helix Transcription Factors/deficiency , Interneurons/metabolism , Neuronal Plasticity/physiology , Nociception/physiology , Posterior Horn Cells/metabolism , Reflex/physiology , Transcription Factors/deficiency , Animals , Basic Helix-Loop-Helix Transcription Factors/biosynthesis , Basic Helix-Loop-Helix Transcription Factors/genetics , Biomarkers/metabolism , Female , Genome-Wide Association Study/methods , Mice , Mice, Knockout , Transcription Factors/biosynthesis , Transcription Factors/genetics
5.
Dev Cell ; 23(3): 469-81, 2012 Sep 11.
Article in English | MEDLINE | ID: mdl-22940113

ABSTRACT

Skeletal muscle growth and regeneration rely on myogenic progenitor and satellite cells, the stem cells of postnatal muscle. Elimination of Notch signals during mouse development results in premature differentiation of myogenic progenitors and formation of very small muscle groups. Here we show that this drastic effect is rescued by mutation of the muscle differentiation factor MyoD. However, rescued myogenic progenitors do not assume a satellite cell position and contribute poorly to myofiber growth. The disrupted homing is due to a deficit in basal lamina assembly around emerging satellite cells and to their impaired adhesion to myofibers. On a molecular level, emerging satellite cells deregulate the expression of basal lamina components and adhesion molecules like integrin α7, collagen XVIIIα1, Megf10, and Mcam. We conclude that Notch signals control homing of satellite cells, stimulating them to contribute to their own microenvironment and to adhere to myofibers.


Subject(s)
Muscle, Skeletal/cytology , Receptors, Notch/metabolism , Satellite Cells, Skeletal Muscle/cytology , Satellite Cells, Skeletal Muscle/metabolism , Signal Transduction , Animals , Cell Adhesion , Mice , Mice, Knockout , Muscle, Skeletal/metabolism , MyoD Protein/genetics , MyoD Protein/metabolism , Receptors, Notch/genetics
6.
Dev Cell ; 21(2): 273-87, 2011 Aug 16.
Article in English | MEDLINE | ID: mdl-21782525

ABSTRACT

Coordinating the balance between progenitor self-renewal and myogenic differentiation is required for a regulated expansion of the developing muscles. Previous observation that neural crest cells (NCCs) migrate throughout the somite regions, where trunk skeletal muscles first emerge, suggests a potential role for these cells in influencing early muscle formation. However, specific signaling interactions between NCCs and skeletal muscle cells remain unknown. Here we show that mice with specific NCC and peripheral nervous system defects display impaired survival of skeletal muscle and show skeletal muscle progenitor cell (MPC) depletion due to precocious commitment to differentiation. We show that reduced NCC-derived Neuregulin1 (Nrg1) in the somite region perturbs ErbB3 signaling in uncommitted MPCs. Using a combination of explant culture experiments and genetic ablation in the mouse, we demonstrate that Nrg1 signals provided by the NCC lineage play a critical role in sustainable myogenesis, by restraining MPCs from precocious differentiation.


Subject(s)
Cell Differentiation/physiology , Cell Lineage/physiology , Muscle Development/physiology , Muscle, Skeletal/cytology , Neural Crest/cytology , Neuregulin-1/metabolism , Receptor, ErbB-3/metabolism , Signal Transduction/physiology , Animals , Caspase 3/metabolism , Cell Movement/genetics , Cell Movement/physiology , Embryo, Mammalian , Gene Expression Regulation, Developmental/genetics , Mice , Mice, Transgenic , Muscle Proteins/genetics , Muscle Proteins/metabolism , Muscle, Skeletal/embryology , MyoD Protein/metabolism , Neuregulin-1/genetics , Neurofilament Proteins/metabolism , Organ Culture Techniques , PAX7 Transcription Factor/metabolism , Receptor, ErbB-3/genetics , SOXE Transcription Factors/deficiency , SOXE Transcription Factors/genetics , Transcription Factors/genetics , Transcription Factors/metabolism , Wnt1 Protein/genetics
7.
Development ; 136(2): 295-305, 2009 Jan.
Article in English | MEDLINE | ID: mdl-19088088

ABSTRACT

The Olig3 gene encodes a bHLH factor that is expressed in the ventricular zone of the dorsal alar plate of the hindbrain. We found that the Olig3(+) progenitor domain encompassed subdomains that co-expressed Math1, Ngn1, Mash1 and Ptf1a. Olig3(+) cells give rise to neuronal types in the dorsal alar plate that we denote as class A neurons. We used genetic lineage tracing to demonstrate that class A neurons contribute to the nucleus of the solitary tract and to precerebellar nuclei. The fate of class A neurons was not correctly determined in Olig3 mutant mice. As a consequence, the nucleus of the solitary tract did not form, and precerebellar nuclei, such as the inferior olivary nucleus, were absent or small. At the expense of class A neurons, ectopic Lbx1(+) neurons appeared in the alar plate in Olig3 mutant mice. By contrast, electroporation of an Olig3 expression vector in the chick hindbrain suppressed the emergence of Lbx1(+) neurons. Climbing fiber neurons of the inferior olivary nucleus express Foxd3 and require Olig3 as well as Ptf1a for the determination of their fate. We observed that electroporation of Olig3 and Ptf1a expression vectors, but not either alone, induced Foxd3. We therefore propose that Olig3 can cooperate with Ptf1a to determine the fate of climbing fiber neurons of the inferior olivary nucleus.


Subject(s)
Basic Helix-Loop-Helix Transcription Factors/metabolism , Brain Stem/embryology , Rhombencephalon/embryology , Animals , Basic Helix-Loop-Helix Transcription Factors/genetics , Brain Stem/cytology , Brain Stem/metabolism , Chick Embryo , Electroporation , Female , Mice , Mice, Knockout , Mice, Mutant Strains , Neuroepithelial Cells/classification , Neuroepithelial Cells/cytology , Neurogenesis/genetics , Neurogenesis/physiology , Pregnancy , Rhombencephalon/cytology , Rhombencephalon/metabolism , Spinal Cord/cytology , Spinal Cord/embryology , Spinal Cord/metabolism , Transcription Factors/genetics , Transcription Factors/metabolism
8.
Dev Biol ; 322(2): 381-93, 2008 Oct 15.
Article in English | MEDLINE | ID: mdl-18721803

ABSTRACT

Dorsal horn neurons express many different neuropeptides that modulate sensory perception like the sensation of pain. Inhibitory neurons of the dorsal horn derive from postmitotic neurons that express Pax2, Lbx1 and Lhx1/5, and diversify during maturation. In particular, fractions of maturing inhibitory neurons express various neuropeptides. We demonstrate here that a coordinate molecular mechanism determines inhibitory and peptidergic fate in the developing dorsal horn. A bHLH factor complex that contains Ptf1a acts as upstream regulator and initiates the expression of several downstream transcription factors in the future inhibitory neurons, of which Pax2 is known to determine the neurotransmitter phenotype. We demonstrate here that dynorphin, galanin, NPY, nociceptin and enkephalin expression depends on Ptf1a, indicating that these neuropeptides are expressed in inhibitory neurons. Furthermore, we show that Neurod1/2/6 and Lhx1/5, which act downstream of Ptf1a, control distinct aspects of peptidergic differentiation. In particular, the Neurod1/2/6 factors are essential for dynorphin and galanin expression, whereas the Lhx1/5 factors are essential for NPY expression. We conclude that a transcriptional network operates in maturing dorsal horn neurons that coordinately determines transmitter and peptidergic fate.


Subject(s)
Neuropeptides/biosynthesis , Posterior Horn Cells/embryology , Spinal Cord/embryology , Transcription Factors/metabolism , Transcription, Genetic , Animals , Basic Helix-Loop-Helix Transcription Factors/metabolism , Ganglia, Spinal/cytology , Ganglia, Spinal/embryology , Ganglia, Spinal/metabolism , Gene Expression Profiling , Homeodomain Proteins/metabolism , LIM-Homeodomain Proteins , Mice , Mice, Mutant Strains , Nerve Tissue Proteins/metabolism , Neuropeptides/metabolism , Posterior Horn Cells/metabolism , Spinal Cord/cytology , Spinal Cord/metabolism
9.
Exp Cell Res ; 313(14): 3127-40, 2007 Aug 15.
Article in English | MEDLINE | ID: mdl-17617404

ABSTRACT

Mutations in genes encoding epidermal keratins cause skin disorders, while those in internal epithelial keratins, such as K8 and K18, are risk factors for liver diseases. The effect of dominant mutations in K8 or K18 during embryonic development and tissue homeostasis has not been examined so far. Here we demonstrate that the dominant mutation hK18 R89C, that is highly similar to hK14 R125C, causing EBS in humans, leads to cell type-specific lethality in mice, depending on the ratio of mutant to endogenous keratins. Mice expressing hK18 R89C in the absence of endogenous K19 and K18 died at mid-gestation from defects in trophoblast giant cells, accompanied by haematomas. A single, endogenous K18 allele rescued embryonic lethality but caused aggregation of keratins in all adult internal epithelia, surprisingly without spontaneous cell fragility. Closer analysis revealed that both filaments and aggregates coexisted in the same cell, depending on the ratio of mutant to endogenous keratins. Our results demonstrate that balanced overexpression of a wild-type keratin rescued the lethal consequences of a dominant-negative mutation. This has important implications for therapy approaches of keratinopathies, suggesting that suppressing the mutant allele is not necessary in vivo.


Subject(s)
Amino Acid Motifs , Keratin-18/genetics , Keratin-18/metabolism , Mutation , Protein Structure, Secondary , Animals , Cytoskeleton/metabolism , Embryo, Mammalian/anatomy & histology , Embryo, Mammalian/physiology , Humans , Intestinal Mucosa/metabolism , Intestines/embryology , Intestines/pathology , Intestines/ultrastructure , Keratin-18/chemistry , Keratin-18/ultrastructure , Mice , Mice, Inbred C57BL , Mice, Knockout , Trophoblasts/cytology
10.
Development ; 133(11): 2105-13, 2006 Jun.
Article in English | MEDLINE | ID: mdl-16690754

ABSTRACT

dILA and dILB neurons comprise the major neuronal subtypes generated in the dorsal spinal cord, and arise in a salt-and-pepper pattern from a broad progenitor domain that expresses the bHLH factor Mash1. In this domain, Mash1-positive and Mash1-negative cells intermingle. Using a Mash1(GFP) allele in mice, we show here that Mash1+ progenitors give rise to dILA and dILB neurons. Using retroviral tracing in the chick, we demonstrate that a single progenitor can give rise to a dILA and a dILB neuron, and that dILA neurons are the product of asymmetric progenitor cell divisions. In Mash1-null mutant mice, the development of dILA, but not of dILB neurons is impaired. We provide evidence that a dual function of Mash1 in neuronal differentiation and specification accounts for the observed changes in the mutant mice. Our data allow us to assign to Mash1 a function in asymmetric cell divisions, and indicate that the factor coordinates cell cycle exit and specification in the one daughter that gives rise to a dILA neuron.


Subject(s)
Basic Helix-Loop-Helix Transcription Factors/metabolism , Neurons/cytology , Neurons/metabolism , Spinal Cord/cytology , Stem Cells/cytology , Stem Cells/metabolism , Animals , Basic Helix-Loop-Helix Transcription Factors/genetics , Cell Differentiation , Cell Division , Cell Lineage , Gene Expression Regulation, Developmental , Mice , Mice, Transgenic , Mutation/genetics , Spinal Cord/embryology , Spinal Cord/metabolism , Transcription Factors/metabolism
11.
Eur J Cell Biol ; 85(1): 1-10, 2006 Jan.
Article in English | MEDLINE | ID: mdl-16373170

ABSTRACT

To get new insights into the function of the intermediate filament (IF) protein vimentin in cell physiology, we generated two mutant cDNAs, one with a point mutation in the consensus motif in coil1A (R113C) and one with the complete deletion of coil 2B of the rod domain. In keratins and glia filament protein (GFAP), analogous mutations cause keratinopathies and Alexander disease, respectively. Both mutants prevented filament assembly in vitro and inhibited assembly of wild-type vimentin when present in equal amounts. In stably transfected preadipocytes, these mutants caused the complete disruption of the endogenous vimentin network, demonstrating their dominant-negative behaviour. Cytoplasmic vimentin aggregates colocalised with the chaperones alphaB-crystallin and HSP40. Moreover, vimR113C mutant cells were more resistant against staurosporine-induced apoptosis compared to controls. We hypothesise that mutations in the vimentin gene, like in most classes of IF genes, may contribute to distinct human diseases.


Subject(s)
Apoptosis , Cytoskeleton/metabolism , Fibroblasts/cytology , Fibroblasts/metabolism , Mutation/genetics , Vimentin/genetics , Vimentin/metabolism , 3T3 Cells , Animals , Cells, Cultured , Cytoplasm , Inclusion Bodies/metabolism , Mice , Molecular Chaperones/metabolism , Multiprotein Complexes , Protein Transport , Subcellular Fractions , Vimentin/ultrastructure , Viscosity
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