RESUMEN
Pacific oyster (Crassostrea gigas), the most productive economical bivalve mollusc, is identified as an attractive model for developmental studies due to its classical mosaic developmental pattern. Myosin heavy chain is a structural and functional component of myosin, the key muscle protein of thick filament. Here, full length cDNA of striated myosin heavy chains in C. gigas (CgSmhc) was obtained, and the expression profiles were examined in different development stage. CgSmhc had a high expression level in trochophore and D-shaped stage during embryo-larval stage. In adult, CgSmhc was a muscle-specific gene and primarily expressed in muscle tissues. Then, activity of 5' flanking region of CgSmhc were examined through an reconstructed EGFP vector. The results indicated that 3098 bp 5'-flanking region of CgSmhc owned various conserved binding sites of myogenesis-related regulatory elements, and the 2000 bp 5'-flanking sequence was sufficient to induce the CgSmhc expression. Subsequently, the CRISPR/Cas9-mediated target disruption of CgSmhc was generated by co-injection of Cas9mRNA and CgSmhc-sgRNAs into one-cell stage embryos of C. gigas. Loss of CgSmhc had a visible effect on the sarcomeric organization of thin filaments in larval musculature, indicating that CgSmhc was required during larval myogenesis to regulate the correct assembly of sarcomere.
Asunto(s)
Crassostrea , Desarrollo de Músculos , Músculo Estriado/crecimiento & desarrollo , Cadenas Pesadas de Miosina/fisiología , Animales , Crassostrea/crecimiento & desarrollo , Expresión Génica , Larva/crecimiento & desarrolloRESUMEN
The well-orchestrated turnover of proteins in cross-striated muscles is one of the fundamental processes required for muscle cell function and survival. Dysfunction of the intricate protein degradation machinery is often associated with development of cardiac and skeletal muscle myopathies. Most muscle proteins are degraded by the ubiquitin-proteasome system (UPS). The UPS involves a number of enzymes, including E3-ligases, which tightly control which protein substrates are marked for degradation by the proteasome. Recent data reveal that E3-ligases of the cullin family play more diverse and crucial roles in cross striated muscles than previously anticipated. This review highlights some of the findings on the multifaceted functions of cullin-RING E3-ligases, their substrate adapters, muscle protein substrates, and regulatory proteins, such as the Cop9 signalosome, for the development of cross striated muscles, and their roles in the etiology of myopathies.
Asunto(s)
Proteínas Cullin/metabolismo , Músculo Estriado/fisiología , Enfermedades Musculares/metabolismo , Complejo del Señalosoma COP9/metabolismo , Regulación del Desarrollo de la Expresión Génica , Humanos , Proteínas Musculares/metabolismo , Músculo Estriado/crecimiento & desarrollo , ProteolisisRESUMEN
Chronic pressure overload due to aortic valve stenosis leads to pathological cardiac hypertrophy and heart failure. Hypertrophy is accompanied by an increase in myocyte surface area, which requires a proportional increase in the number of cell-cell and cell-matrix contacts to withstand enhanced workload. In a proteomic analysis we identified nerve injury-induced protein 1 (Ninjurin1), a 16kDa transmembrane cell-surface protein involved in cell adhesion and nerve repair, to be increased in hypertrophic hearts from patients with aortic stenosis. We hypothesised that Ninjurin1 is involved in myocyte hypertrophy. We analyzed cardiac biopsies from aortic-stenosis patients and control patients undergoing elective heart surgery. We studied cardiac hypertrophy in mice after transverse aortic constriction and angiotensin II infusions, and performed mechanistic analyses in cultured myocytes. We assessed the physiological role of ninjurin1 in zebrafish during heart and skeletal muscle development. Ninjurin1 was increased in hearts of aortic stenosis patients, compared to controls, as well as in hearts from mice with cardiac hypertrophy. Besides the 16kDa Ninjurin1 (Ninjurin1-16) we detected a 24kDa variant of Ninjurin1 (Ninjurin1-24), which was predominantly expressed during myocyte hypertrophy. We disclosed that the higher molecular weight of Ninjurin1-24 was caused by N-glycosylation. Ninjurin1-16 was contained in the cytoplasm of myocytes where it colocalized with stress-fibers. In contrast, Ninjurin1-24 was localized at myocyte membranes. Gain and loss-of-function experiments showed that Ninjurin1-24 plays a role in myocyte hypertrophy and myogenic differentiation in vitro. Reduced levels of ninjurin1 impaired cardiac and skeletal muscle development in zebrafish. We conclude that Ninjurin1 contributes to myocyte growth and differentiation, and that these effects are mainly mediated by N-glycosylated Ninjurin1-24.
Asunto(s)
Estenosis de la Válvula Aórtica/genética , Cardiomegalia/genética , Moléculas de Adhesión Celular Neuronal/genética , Músculo Estriado/crecimiento & desarrollo , Factores de Crecimiento Nervioso/genética , Animales , Estenosis de la Válvula Aórtica/patología , Cardiomegalia/patología , Diferenciación Celular/genética , Modelos Animales de Enfermedad , Femenino , Humanos , Mutación con Pérdida de Función/genética , Masculino , Ratones , Desarrollo de Músculos/genética , Músculo Estriado/metabolismo , Músculo Estriado/patología , Miocitos Cardíacos/metabolismo , Miocitos Cardíacos/patología , Transducción de Señal/genética , Pez CebraRESUMEN
Metabolic reprogramming is an active regulator of stem cell fate choices, and successful stem cell differentiation in different compartments requires the induction of oxidative phosphorylation. However, the mechanisms that promote mitochondrial respiration during stem cell differentiation are poorly understood. Here we demonstrate that Stat3 promotes muscle stem cell myogenic lineage progression by stimulating mitochondrial respiration in mice. We identify Fam3a, a cytokine-like protein, as a major Stat3 downstream effector in muscle stem cells. We demonstrate that Fam3a is required for muscle stem cell commitment and skeletal muscle development. We show that myogenic cells secrete Fam3a, and exposure of Stat3-ablated muscle stem cells to recombinant Fam3a in vitro and in vivo rescues their defects in mitochondrial respiration and myogenic commitment. Together, these findings indicate that Fam3a is a Stat3-regulated secreted factor that promotes muscle stem cell oxidative metabolism and differentiation, and suggests that Fam3a is a potential tool to modulate cell fate choices.
Asunto(s)
Diferenciación Celular , Citocinas/fisiología , Desarrollo de Músculos/fisiología , Mioblastos/fisiología , Factor de Transcripción STAT3/fisiología , Células Madre/fisiología , Animales , Animales Recién Nacidos , Linaje de la Célula/fisiología , Células Cultivadas , Embrión de Mamíferos , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Mitocondrias/metabolismo , Músculo Estriado/citología , Músculo Estriado/crecimiento & desarrollo , Fosforilación Oxidativa , Transducción de Señal/fisiologíaRESUMEN
Low-density lipoprotein receptor-related protein 6 (LRP6) serves as a Wnt coreceptor. Although Wnt/LRP6 signalling is best known for the ß-catenin-dependent regulation of target genes in tissue development and homeostasis, emerging evidence demonstrates the biological aspects of LRP6 beyond a Wnt coreceptor. Whether LRP6 modulates tissue development in a Wnt/ß-catenin signalling-independent manner remains unknown. Using a model of striated muscle development, we observed that LRP6 was almost undetectable in proliferating myoblasts, whereas its expression gradually increased in the nucleus of myodifferentiating cells. During myodifferentiation, LRP6 modulated the muscle-specific splicing of integrin-ß1D and consequent myotube maturation independently of the ß-catenin-dependent Wnt signalling. Furthermore, we identified that the carboxy-terminal serine-rich region in LRP6 bond to the adenine-rich sequence within alternative exon D (AED) of integrin-ß1 pre-mRNA, and therefore, elicited AED inclusion when the spliceosome was recruited to the splice site. The interaction of LRP6 with the adenine-rich sequence was sufficient to overcome AED exclusion by a splicing repressor, polypyrimidine tract binding protein-1. Besides the integrin-ß1, deep RNA sequencing in different types of cells revealed that the LRP6-mediated splicing regulation was widespread. Thus, our findings implicate LRP6 as a potential regulator for alternative pre-mRNA splicing.
Asunto(s)
Empalme Alternativo , Regulación del Desarrollo de la Expresión Génica , Proteína-6 Relacionada a Receptor de Lipoproteína de Baja Densidad/genética , Desarrollo de Músculos/genética , Músculo Estriado/metabolismo , Precursores del ARN/genética , Animales , Animales Recién Nacidos , Secuencia de Bases , Diferenciación Celular , Línea Celular , Núcleo Celular/metabolismo , Proliferación Celular , Citosol/metabolismo , Exones , Ribonucleoproteínas Nucleares Heterogéneas/genética , Ribonucleoproteínas Nucleares Heterogéneas/metabolismo , Células Endoteliales de la Vena Umbilical Humana/citología , Células Endoteliales de la Vena Umbilical Humana/metabolismo , Humanos , Integrina beta1/genética , Integrina beta1/metabolismo , Proteína-6 Relacionada a Receptor de Lipoproteína de Baja Densidad/metabolismo , Ratones , Músculo Estriado/citología , Músculo Estriado/crecimiento & desarrollo , Mioblastos/citología , Mioblastos/metabolismo , Miocitos Cardíacos/citología , Miocitos Cardíacos/metabolismo , Proteína de Unión al Tracto de Polipirimidina/genética , Proteína de Unión al Tracto de Polipirimidina/metabolismo , Cultivo Primario de Células , Precursores del ARN/metabolismo , Ratas , Vía de Señalización Wnt , beta Catenina/genética , beta Catenina/metabolismoRESUMEN
Muscle Lim Protein (MLP) is a protein with multiple functional roles in striated muscle physiology and pathophysiology. Herein, we demonstrate that MLP directly binds to slow, fast, and cardiac myosin-binding protein C (MyBP-C) during myogenesis, as shown by yeast two-hybrid and a range of protein-protein interaction assays. The minimal interacting domains involve MLP inter-LIM and MyBP-C [C4]. The interaction is sensitive to cytosolic Ca2+ concentrations changes and to MyBP-C phosphorylation by PKA or CaMKII. Confocal microscopy of differentiating myoblasts showed MLP and MyBP-C colocalization during myoblast differentiation. Suppression of the complex formation with recombinant MyBP-C [C4] peptide overexpression, inhibited myoblast differentiation by 65%. Suppression of both MLP and MyBP-C expression in myoblasts by siRNA revealed negative synergistic effects on differentiation. The MLP/MyBP-C complex modulates the actin activated myosin II ATPase activity in vitro, which could interfere with sarcomerogenesis and myofilaments assembly during differentiation. Our data demonstrate a critical role of the MLP/MyBP-C complex during early myoblast differentiation. Its absence in muscles with mutations or aberrant expression of MLP or MyBP-C could be directly implicated in the development of cardiac and skeletal myopathies.
Asunto(s)
Proteínas Portadoras/genética , Diferenciación Celular/genética , Proteínas con Dominio LIM/genética , Desarrollo de Músculos/genética , Proteínas Musculares/genética , Adenosina Trifosfatasas/genética , Animales , Proteínas Portadoras/metabolismo , Regulación del Desarrollo de la Expresión Génica , Humanos , Proteínas con Dominio LIM/metabolismo , Proteínas Musculares/metabolismo , Músculo Estriado/crecimiento & desarrollo , Músculo Estriado/metabolismo , Enfermedades Musculares/genética , Enfermedades Musculares/patología , Mioblastos/metabolismo , Fosforilación , Sarcómeros/genéticaRESUMEN
The cyclic 3',5'-adenosine monophosphate (cAMP) signalling pathway constitutes an ancient signal transduction pathway present in prokaryotes and eukaryotes. Previously, it was thought that in eukaryotes three effector proteins mediate cAMP signalling, namely protein kinase A (PKA), exchange factor directly activated by cAMP (EPAC) and the cyclic-nucleotide gated channels. However, recently a novel family of cAMP effector proteins emerged and was termed the Popeye domain containing (POPDC) family, which consists of three members POPDC1, POPDC2 and POPDC3. POPDC proteins are transmembrane proteins, which are abundantly present in striated and smooth muscle cells. POPDC proteins bind cAMP with high affinity comparable to PKA. Presently, their biochemical activity is poorly understood. However, mutational analysis in animal models as well as the disease phenotype observed in patients carrying missense mutations suggests that POPDC proteins are acting by modulating membrane trafficking of interacting proteins. In this review, we will describe the current knowledge about this gene family and also outline the apparent gaps in our understanding of their role in cAMP signalling and beyond.
Asunto(s)
Moléculas de Adhesión Celular/genética , AMP Cíclico/genética , Proteínas de la Membrana/genética , Proteínas Musculares/genética , Proteínas Quinasas Dependientes de AMP Cíclico/genética , Humanos , Familia de Multigenes/genética , Músculo Estriado/crecimiento & desarrollo , Músculo Estriado/metabolismo , Dominios Proteicos/genética , Transducción de Señal/genéticaRESUMEN
The major sperm protein domain (MSPd) has an extracellular signaling function implicated in amyotrophic lateral sclerosis. Secreted MSPds derived from the C. elegans VAPB homolog VPR-1 promote mitochondrial localization to actin-rich I-bands in body wall muscle. Here we show that the nervous system and germ line are key MSPd secretion tissues. MSPd signals are transduced through the CLR-1 Lar-like tyrosine phosphatase receptor. We show that CLR-1 is expressed throughout the muscle plasma membrane, where it is accessible to MSPd within the pseudocoelomic fluid. MSPd signaling is sufficient to remodel the muscle mitochondrial reticulum during adulthood. An RNAi suppressor screen identified survival of motor neuron 1 (SMN-1) as a downstream effector. SMN-1 acts in muscle, where it colocalizes at myofilaments with ARX-2, a component of the Arp2/3 actin-nucleation complex. Genetic studies suggest that SMN-1 promotes Arp2/3 activity important for localizing mitochondria to I-bands. Our results support the model that VAPB homologs are circulating hormones that pattern the striated muscle mitochondrial reticulum. This function is crucial in adults and requires SMN-1 in muscle, likely independent of its role in pre-mRNA splicing.
Asunto(s)
Proteínas de Caenorhabditis elegans/metabolismo , Caenorhabditis elegans/crecimiento & desarrollo , Caenorhabditis elegans/metabolismo , Proteínas de la Membrana/metabolismo , Músculo Estriado/crecimiento & desarrollo , Músculo Estriado/metabolismo , Proteínas del Complejo SMN/metabolismo , Proteína 2 Relacionada con la Actina/metabolismo , Complejo 2-3 Proteico Relacionado con la Actina/metabolismo , Esclerosis Amiotrófica Lateral/metabolismo , Animales , Animales Modificados Genéticamente , Caenorhabditis elegans/genética , Proteínas de Caenorhabditis elegans/química , Proteínas de Caenorhabditis elegans/genética , Genes de Helminto , Células Germinativas/metabolismo , Humanos , Larva/crecimiento & desarrollo , Larva/metabolismo , Masculino , Proteínas de la Membrana/química , Proteínas de la Membrana/genética , Mitocondrias Musculares/metabolismo , Neuronas Motoras/metabolismo , Mutación , Dominios Proteicos , Interferencia de ARN , Proteínas Tirosina Fosfatasas Similares a Receptores/genética , Proteínas Tirosina Fosfatasas Similares a Receptores/metabolismo , Proteínas del Complejo SMN/antagonistas & inhibidores , Proteínas del Complejo SMN/genética , Sarcolema/metabolismo , Transducción de SeñalRESUMEN
The esophagus functions to transport food from the oropharyngeal region to the stomach via waves of peristalsis and transient relaxation of the lower esophageal sphincter. The gastrointestinal tract, including the esophagus, is ensheathed by the muscularis externa (ME). However, while the ME of the gastrointestinal tract distal to the esophagus is exclusively smooth muscle, the esophageal ME of many vertebrate species comprises a variable amount of striated muscle. The esophageal ME is initially composed only of smooth muscle, but its developmental maturation involves proximal-to-distal replacement of smooth muscle with striated muscle. This fascinating phenomenon raises two important questions: what is the developmental origin of the striated muscle precursor cells, and what are the cellular and morphogenetic mechanisms underlying the process? Studies addressing these questions have provided controversial answers. In this review, we discuss the development of ideas in this area and recent work that has shed light on these issues. A working model has emerged that should permit deeper understanding of the role of ME development and maturation in esophageal disorders and in the functional and evolutionary underpinnings of the variable degree of esophageal striated myogenesis in vertebrate species.
Asunto(s)
Esófago/crecimiento & desarrollo , Desarrollo de Músculos , Músculo Liso/crecimiento & desarrollo , Músculo Estriado/crecimiento & desarrollo , Animales , Esófago/embriología , Esófago/metabolismo , Humanos , Modelos Biológicos , Fibras Musculares Esqueléticas/fisiología , Músculo Liso/embriología , Músculo Liso/metabolismo , Músculo Estriado/embriología , Músculo Estriado/metabolismo , Mioblastos/fisiología , Miocitos del Músculo Liso/fisiologíaRESUMEN
In some rare and striking cases, striated muscle fibers of the skeleton or body wall, which consist of terminally differentiated syncytia with complex ultrastructures, were found to be capable of dedifferentiating and fragmenting into mononucleate cells. Examples of such events will be discussed in which the dedifferentiated cells reenter the cell cycle, proliferate, and rebuilt damaged muscle fibers during limb regeneration or transdifferentiate to generate new types of muscles during normal development.
Asunto(s)
Músculo Estriado/citología , Músculo Estriado/fisiología , Regeneración , Anfibios/fisiología , Animales , Ciclo Celular/fisiología , Transdiferenciación Celular , Cnidarios/fisiología , Drosophila/fisiología , Peces/crecimiento & desarrollo , Metamorfosis Biológica , Músculo Estriado/crecimiento & desarrollo , Células Madre/citología , Células Madre/fisiologíaRESUMEN
Troponin T (TnT) is a central player in the calcium regulation of actin thin filament function and is essential for the contraction of striated muscles. Three homologous genes have evolved in vertebrates to encode three muscle type-specific TnT isoforms: TNNT1 for slow skeletal muscle TnT, TNNT2 for cardiac muscle TnT, and TNNT3 for fast skeletal muscle TnT. Alternative splicing and posttranslational modifications confer additional structural and functional variations of TnT during development and muscle adaptation to various physiological and pathological conditions. This review focuses on the TnT isoform genes and their molecular evolution, alternative splicing, developmental regulation, structure-function relationships of TnT proteins, posttranslational modifications, and myopathic mutations and abnormal splicing. The goal is to provide a concise summary of the current knowledge and some perspectives for future research and translational applications.
Asunto(s)
Evolución Molecular , Troponina T/genética , Citoesqueleto de Actina/genética , Citoesqueleto de Actina/metabolismo , Empalme Alternativo/genética , Calcio/metabolismo , Regulación del Desarrollo de la Expresión Génica , Humanos , Contracción Muscular/genética , Músculo Esquelético/crecimiento & desarrollo , Músculo Esquelético/metabolismo , Músculo Estriado/crecimiento & desarrollo , Músculo Estriado/metabolismo , Isoformas de Proteínas/biosíntesis , Isoformas de Proteínas/genética , Relación Estructura-Actividad , Troponina T/biosíntesis , Troponina T/química , Troponina T/metabolismoRESUMEN
BACKGROUND: The striated muscle Z-line, a multiprotein complex at the boundary between sarcomeres, plays an integral role in maintaining striated muscle structure and function. Multiple Z-line-associated proteins have been identified and shown to play an increasingly important role in the pathogenesis of human cardiomyopathy. Cypher and its close homologue, Enigma homolog protein (ENH), are 2 Z-line proteins previously shown to be individually essential for maintenance of postnatal cardiac function and stability of the Z-line during muscle contraction, but dispensable for cardiac myofibrillogenesis and development. METHODS AND RESULTS: The current studies were designed to test whether Cypher and ENH play redundant roles during embryonic development. Here, we demonstrated that mice lacking both ENH and Cypher exhibited embryonic lethality and growth retardation. Lethality in double knockout embryos was associated with cardiac dilation and abnormal Z-line structure. In addition, when ENH was ablated in conjunction with selective ablation of either Cypher short isoforms (CypherS), or Cypher long isoforms (CypherL), only the latter resulted in embryonic lethality. CONCLUSIONS: Cypher and ENH redundantly play an essential role in sustaining Z-line structure from the earliest stages of cardiac function, and are redundantly required to maintain normal embryonic heart function and embryonic viability.
Asunto(s)
Proteínas Adaptadoras Transductoras de Señales/genética , Cardiomiopatías/genética , Desarrollo Embrionario/genética , Proteínas con Dominio LIM/genética , Proteínas de Microfilamentos/genética , Músculo Estriado/anomalías , Músculo Estriado/crecimiento & desarrollo , Proteínas Adaptadoras Transductoras de Señales/deficiencia , Animales , Proteínas con Dominio LIM/deficiencia , Ratones , Ratones Noqueados , Proteínas Musculares/genética , Proteínas Musculares/metabolismo , Miocardio/patología , Isoformas de Proteínas/genética , Factores de RiesgoRESUMEN
Transcriptome analysis allowed the identification of new long noncoding RNAs differentially expressed during murine myoblast differentiation. These transcripts were classified on the basis of their expression under proliferating versus differentiated conditions, muscle-restricted activation, and subcellular localization. Several species displayed preferential expression in dystrophic (mdx) versus wild-type muscles, indicating their possible link with regenerative processes. One of the identified transcripts, lnc-31, even if originating from the same nuclear precursor of miR-31, is produced by a pathway mutually exclusive. We show that lnc-31 and its human homologue hsa-lnc-31 are expressed in proliferating myoblasts, where they counteract differentiation. In line with this, both species are more abundant in mdx muscles and in human Duchenne muscular dystrophy (DMD) myoblasts, than in their normal counterparts. Altogether, these data suggest a crucial role for lnc-31 in controlling the differentiation commitment of precursor myoblasts and indicate that its function is maintained in evolution despite the poor sequence conservation with the human counterpart.
Asunto(s)
MicroARNs/genética , Desarrollo de Músculos/genética , Distrofia Muscular de Duchenne/genética , Mioblastos/metabolismo , ARN Largo no Codificante/genética , Animales , Evolución Biológica , Diferenciación Celular , Proliferación Celular , Perfilación de la Expresión Génica , Regulación de la Expresión Génica , Humanos , Ratones , Ratones Transgénicos , MicroARNs/metabolismo , Músculo Liso/crecimiento & desarrollo , Músculo Liso/metabolismo , Músculo Liso/patología , Músculo Estriado/crecimiento & desarrollo , Músculo Estriado/metabolismo , Músculo Estriado/patología , Distrofia Muscular de Duchenne/metabolismo , Distrofia Muscular de Duchenne/patología , Mioblastos/patología , Precursores del ARN/genética , Precursores del ARN/metabolismo , ARN Largo no Codificante/metabolismo , TranscriptomaRESUMEN
The nematode Caenorhabditis elegans has been used as a valuable system to study structure and function of striated muscle. The body wall muscle of C. elegans is obliquely striated muscle with highly organized sarcomeric assembly of actin, myosin, and other accessory proteins. Genetic and molecular biological studies in C. elegans have identified a number of genes encoding structural and regulatory components for the muscle contractile apparatuses, and many of them have counterparts in mammalian cardiac and skeletal muscles or striated muscles in other invertebrates. Applicability of genetics, cell biology, and biochemistry has made C. elegans an excellent system to study mechanisms of muscle contractility and assembly and maintenance of myofibrils. This review focuses on the regulatory mechanisms of structure and function of actin filaments in the C. elegans body wall muscle. Sarcomeric actin filaments in C. elegans muscle are associated with the troponin-tropomyosin system that regulates the actin-myosin interaction. Proteins that bind to the side and ends of actin filaments support ordered assembly of thin filaments. Furthermore, regulators of actin dynamics play important roles in initial assembly, growth, and maintenance of sarcomeres. The knowledge acquired in C. elegans can serve as bases to understand the basic mechanisms of muscle structure and function.
Asunto(s)
Citoesqueleto de Actina/fisiología , Actinas/metabolismo , Proteínas de Caenorhabditis elegans/metabolismo , Caenorhabditis elegans/fisiología , Contracción Muscular , Músculo Estriado/fisiología , Sarcómeros/fisiología , Citoesqueleto de Actina/metabolismo , Animales , Caenorhabditis elegans/anatomía & histología , Caenorhabditis elegans/crecimiento & desarrollo , Caenorhabditis elegans/metabolismo , Desarrollo de Músculos , Músculo Estriado/anatomía & histología , Músculo Estriado/crecimiento & desarrollo , Músculo Estriado/metabolismo , Sarcómeros/metabolismo , Transducción de SeñalRESUMEN
The sarcomeres of striated muscle are among the most elaborate and dynamic eukaryotic cellular protein machinery, and the mechanisms by which these semicrystalline filament networks are initially patterned and assembled remain contentious. In addition to the acto-myosin filaments that provide motor function, the sarcomere contains titin filaments, comprised of individual molecules of the giant Ig- and fibronectin domain-rich protein titin. Titin is the largest known protein, containing many structurally distinct domains with a variety of proposed functions, including sarcomere stabilization, the prevention of over-stretching, and returning to resting length after contraction. One molecule of titin, which binds to both the Z-disk and the M-line, spans a half-sarcomere, and is proposed to serve as a "molecular ruler" that dictates the spacing of sarcomeres. The semirigid rod-like A-band region of titin has also been proposed to act as a scaffold for thick filament formation during muscle development, but despite decades of research, this hypothesis has not been rigorously tested. Recent studies in zebrafish have brought into question the necessity for the A-band region of titin during the early stages of sarcomere patterning. In this review, we give an overview of the many different roles of titin in the development and function of striated muscle, and address the validity of the "molecular ruler" model of myofibrillogenesis in light of the current literature.
Asunto(s)
Conectina/metabolismo , Desarrollo de Músculos , Músculo Estriado/fisiología , Sarcómeros/fisiología , Animales , Humanos , Músculo Estriado/crecimiento & desarrollo , Músculo Estriado/metabolismo , Sarcómeros/metabolismo , Transducción de SeñalRESUMEN
Muscle contraction depends on interactions between actin and myosin filaments organized into sarcomeres, but the mechanism by which actin filaments incorporate into sarcomeres remains unclear. We have found that, during larval development in Caenorhabditis elegans, two members of the actin-assembling formin family, CYK-1 and FHOD-1, are present in striated body wall muscles near or on sarcomere Z lines, where barbed ends of actin filaments are anchored. Depletion of either formin during this period stunted growth of the striated contractile lattice, whereas their simultaneous reduction profoundly diminished lattice size and number of striations per muscle cell. CYK-1 persisted at Z lines in adulthood, and its near complete depletion from adults triggered phenotypes ranging from partial loss of Z line-associated filamentous actin to collapse of the contractile lattice. These results are, to our knowledge, the first genetic evidence implicating sarcomere-associated formins in the in vivo organization of the muscle cytoskeleton.
Asunto(s)
Proteínas de Caenorhabditis elegans/metabolismo , Caenorhabditis elegans/crecimiento & desarrollo , Proteínas de Microfilamentos/metabolismo , Músculo Estriado/crecimiento & desarrollo , Músculo Estriado/metabolismo , Sarcómeros/metabolismo , Animales , Caenorhabditis elegans/metabolismo , Proteínas de Caenorhabditis elegans/genética , Forminas , Proteínas de Microfilamentos/genéticaRESUMEN
The adult musculature in D. melanogaster forms during metamorphosis. Much is known about the flight and leg musculature, but not about the muscles surrounding the male reproductive tract. The inner genitalia of males consist of the testes, which emerge from the gonads; the remaining genital organs, i.e., paragonia (or accessory glands), ejaculatory duct, sperm pump, and seminal vesicles, develop out of the genital imaginal disc. We analyzed the myoblasts forming the muscle layers of these organs. In myoblasts derived from the genital imaginal disc, the regulatory region of the transcription factor DMef2 is active. DMef2 is also needed for specification and differentiation of embryonic and adult myoblasts. We could discriminate three different muscle types: (i) multinucleated muscles that resemble vertebrate smooth muscles surround the testes, (ii) multinucleated muscles that resemble striated muscles comprises seminal vesicles and the sperm pump, and (iii) mononucleated striated musculature encloses the paragonia and ejaculatory duct. Members of the immunoglobulin superfamily involved in embryonic myogenesis, Dumbfounded (Duf) and Sticks and Stones (Sns), were also expressed in the genital imaginal disc, in the muscle sheath of the testes during muscle differentiation and in the secretory secondary cells, which are part of the binucleated epithelia enclosing the paragonia.
Asunto(s)
Drosophila melanogaster/crecimiento & desarrollo , Regulación del Desarrollo de la Expresión Génica/fisiología , Genitales Masculinos/crecimiento & desarrollo , Metamorfosis Biológica/fisiología , Desarrollo de Músculos/fisiología , Músculo Liso/crecimiento & desarrollo , Músculo Estriado/crecimiento & desarrollo , Animales , Diferenciación Celular/fisiología , Proteínas de Drosophila/metabolismo , Regulación del Desarrollo de la Expresión Génica/genética , Genitales Masculinos/ultraestructura , Inmunoglobulinas/metabolismo , Inmunohistoquímica , Masculino , Proteínas de la Membrana/metabolismo , Microscopía Electrónica de Rastreo , Proteínas Musculares/metabolismo , Mioblastos/citología , Mioblastos/ultraestructuraRESUMEN
The ENH (PDLIM5) protein acts as a scaffold to tether various functional proteins at subcellular sites via PDZ and three LIM domains. Splicing of the ENH primary transcript generates various products with different repertories of protein interaction modules. Three LIM-containing ENH predominates in neonatal cardiac tissue, whereas LIM-less ENHs are abundant in adult hearts, as well as skeletal muscles. Here we examine the timing of splicing transitions of ENH gene products during postnatal heart development and C2C12 myoblast differentiation. Real-time PCR analysis shows that LIM-containing ENH1 mRNA is gradually decreased during postnatal heart development, whereas transcripts with the short exon 5 appear in the late postnatal period and continues to increase until at least one month after birth. The splicing transition from LIM-containing ENH1 to LIM-less ENHs is also observed during the early period of C2C12 differentiation. This transition correlates with the emergence of ENH transcripts with the short exon 5, as well as the expression of myogenin mRNA. In contrast, the shift from the short exon 5 to the exon 7 occurs in the late differentiation period. The timing of this late event corresponds to the appearance of mRNA for the skeletal myosin heavy chain MYH4. Thus, coordinated and stepwise splicing transitions result in the production of specific ENH transcripts in mature striated muscles.
Asunto(s)
Proteínas Adaptadoras Transductoras de Señales/genética , Regulación del Desarrollo de la Expresión Génica , Corazón/crecimiento & desarrollo , Proteínas de Microfilamentos/genética , Desarrollo de Músculos/genética , Músculo Estriado/crecimiento & desarrollo , Empalme del ARN , Animales , Diferenciación Celular/genética , Línea Celular , Ratones , Músculo Estriado/citología , Mioblastos Cardíacos/citología , Ratas , Ratas Sprague-DawleyRESUMEN
A precise organization of gene expression is required for developing neural and muscular systems. Steroid hormones can control the expression of genes that are critical for development. In this study we test the hypothesis that the steroid hormone ecdysone regulates gene expression of the voltage-gated calcium-activated potassium ion channel, Slowpoke or KCNMA1. Late in adult development of the tobacco hawkmoth Manduca sexta, slowpoke (msslo) levels increased contributing to the maturation of the dorsal longitudinal flight muscles (DLMs) and CNS. We show that critical components of ecdysteroid gene regulation were present during upreglation of msslo in late adult DLM and CNS development. Ecdysteroid receptor complex heterodimeric partner proteins, the ecdysteroid receptor (EcR) and ultraspiracle (USP), and the ecdysone-induced early gene, msE75B, were expressed at key developmental time points, suggesting that ecdysteroids direct aspects of gene expression in the DLMs during these late developmental stages. We provide evidence that ecdysteroids suppress msslo transcription in the DLMs; when titers decline msslo transcript levels increase. These results are consistent with msslo being a downstream gene in an ecdysteroid-mediated gene cascade during DLM development. We also show that the ecdysteroids regulate msslo transcript levels in the developing CNS. These results will contribute to our understanding of how the spatiotemporal regulation of slowpoke transcription contributes to tailoring cell excitability to the differing physiological and behavioral demands during development.
Asunto(s)
Músculo Estriado/crecimiento & desarrollo , Sistema Nervioso/crecimiento & desarrollo , Canales de Potasio con Entrada de Voltaje/biosíntesis , Canales de Potasio con Entrada de Voltaje/genética , Esteroides/fisiología , Animales , Ecdisona/fisiología , Regulación del Desarrollo de la Expresión Génica/fisiología , Manduca , Músculo Estriado/citología , Músculo Estriado/metabolismo , Sistema Nervioso/citología , Sistema Nervioso/metabolismoRESUMEN
The N-terminus of telethonin forms a unique structure linking two titin N-termini at the Z-disc. While a specific role for the C-terminus has not been established, several studies indicate it may have a regulatory function. Using a morpholino approach in Xenopus, we show that telethonin knockdown leads to embryonic paralysis, myocyte defects, and sarcomeric disruption. These myopathic defects can be rescued by expressing full-length telethonin mRNA in morpholino background, indicating that telethonin is required for myofibrillogenesis. However, a construct missing C-terminal residues is incapable of rescuing motility or sarcomere assembly in cultured myocytes. We, therefore, tested two additional constructs: one where four C-terminal phosphorylatable residues were mutated to alanines and another where terminal residues were randomly replaced. Data from these experiments support that the telethonin C-terminus is required for assembly, but in a context-dependent manner, indicating that factors and forces present in vivo can compensate for C-terminal truncation or mutation.