RESUMEN
RNA-binding proteins (RBPs) regulate gene expression at the post-transcriptional level. They play major roles in the tissue- and stage-specific expression of protein isoforms as well as in the maintenance of protein homeostasis. The inner ear is a bi-functional organ, with the cochlea and the vestibular system required for hearing and for maintaining balance, respectively. It is relatively well documented that transcription factors and signaling pathways are critically involved in the formation of inner ear structures and in the development of hair cells. Accumulating evidence highlights emerging functions of RBPs in the post-transcriptional regulation of inner ear development and hair cell function. Importantly, mutations of splicing factors of the RBP family and defective alternative splicing, which result in inappropriate expression of protein isoforms, lead to deafness in both animal models and humans. Because RBPs are critical regulators of cell proliferation and differentiation, they present the potential to promote hair cell regeneration following noise- or ototoxin-induced damage through mitotic and non-mitotic mechanisms. Therefore, deciphering RBP-regulated events during inner ear development and hair cell regeneration can help define therapeutic strategies for treatment of hearing loss. In this review, we outline our evolving understanding of the implications of RBPs in hair cell formation and hearing disease with the aim of promoting future research in this field.
Asunto(s)
Oído Interno , Animales , Humanos , Oído Interno/metabolismo , Factores de Transcripción/metabolismo , Proteínas de Unión al ARN/genética , Proteínas de Unión al ARN/metabolismo , Factores de Empalme de ARN/metabolismo , Cabello/metabolismoRESUMEN
BACKGROUND: RNA-binding proteins plays critical roles in several post-transcriptional regulatory processes. The RNA-binding protein, Rbm24, has been shown to be involved in the development of the heart and skeletal muscles by regulating different post-transcriptional processes such as splicing and stabilization of specific target mRNAs. Here, by performing a detailed expression and localization analysis in mice embryos, we show that Rbm24 protein is not only expressed in heart and skeletal muscles as previously reported, but it is also strongly and specifically detected in specific regions of all the head sensory organs during mouse development. RESULTS: Rbm24 expression is indeed found to be activated in the lens, in the sensory olfactory epithelium and in mechanosensory cells of the auditory and vestibular systems. Within these territories, Rbm24 is shown to be restricted to distinct subdomains, potentially regulating cell specificity and proliferation. Moreover, Rbm24 protein is found to be restricted to the cytoplasmic compartment in all these organs, thus providing clues to the posttranscriptional activity that it may exert in these cells. CONCLUSIONS: Altogether, these results highlight that Rbm24 may potentially function as a novel key regulator for the development of the eye, nasal epithelium, and inner ear in vertebrates. Developmental Dynamics 247:1160-1169, 2018. © 2018 Wiley Periodicals, Inc.
Asunto(s)
Oído Interno/metabolismo , Desarrollo Embrionario , Cristalino/metabolismo , Proteínas de Unión al ARN/metabolismo , Animales , Proliferación Celular , Citoplasma/química , Oído Interno/embriología , Embrión de Mamíferos , Cristalino/embriología , Ratones , Mucosa Nasal/metabolismoRESUMEN
Mesoderm and embryonic axis formation in vertebrates is mediated by maternal and zygotic factors that activate the expression of target genes. Transcriptional derepression plays an important role in the regulation of expression in different contexts; however, its involvement and possible mechanism in mesoderm and embryonic axis formation are largely unknown. Here we demonstrate that XDSCR6, a Xenopus homologue of human Down syndrome critical region protein 6 (DSCR6, or RIPPLY3), regulates mesoderm and embryonic axis formation through derepression of polycomb group (PcG) proteins. Xdscr6 maternal mRNA is enriched in the endoderm of the early gastrula and potently triggers the formation of dorsal mesoderm and neural tissues in ectoderm explants; it also dorsalises ventral mesoderm during gastrulation and induces a secondary embryonic axis. A WRPW motif, which is present in all DSCR6 homologues, is necessary and sufficient for the dorsal mesoderm- and axis-inducing activity. Knockdown of Xdscr6 inhibits dorsal mesoderm gene expression and results in head deficiency. We further show that XDSCR6 physically interacts with PcG proteins through the WRPW motif, preventing the formation of PcG bodies and antagonising their repressor activity in embryonic axis formation. By chromatin immunoprecipitation, we demonstrate that XDSCR6 releases PcG proteins from chromatin and allows dorsal mesoderm gene transcription. Our studies suggest that XDSCR6 might function to sequester PcG proteins and identify a novel derepression mechanism implicated in embryonic induction and axis formation.
Asunto(s)
Tipificación del Cuerpo/fisiología , Mesodermo/metabolismo , Proteínas del Grupo Polycomb/metabolismo , Proteínas de Xenopus/metabolismo , Xenopus/embriología , Animales , Cromatina/metabolismo , Síndrome de Down/genética , Ectodermo/embriología , Ectodermo/metabolismo , Embrión no Mamífero/embriología , Embrión no Mamífero/metabolismo , Inducción Embrionaria , Gástrula/citología , Gastrulación , Regulación del Desarrollo de la Expresión Génica , Humanos , Mesodermo/embriología , Proteínas del Grupo Polycomb/biosíntesis , ARN Mensajero , Somitos/embriología , Factores de Transcripción , Transcripción Genética , Xenopus/genética , Xenopus/metabolismo , Proteínas de Xenopus/genéticaRESUMEN
The cranial base is essential for integrated craniofacial development and growth. It develops as a cartilaginous template that is replaced by bone through the process of endochondral ossification. Here, we describe a novel and specific role for the homeoprotein Six2 in the growth and elongation of the cranial base. Six2-null newborn mice display premature fusion of the bones in the cranial base. Chondrocyte differentiation is abnormal in the Six2-null cranial base, with reduced proliferation and increased terminal differentiation. Gain-of-function experiments indicate that Six2 promotes cartilage development and growth in other body areas and appears therefore to control general regulators of chondrocyte differentiation. Our data indicate that the main factors restricting Six2 function to the cranial base are tissue-specific transcription of the gene and compensatory effects of other Six family members. The comparable expression during human embryogenesis and the high protein conservation from mouse to human implicate SIX2 loss-of-function as a potential congenital cause of anterior cranial base defects in humans.
Asunto(s)
Base del Cráneo/crecimiento & desarrollo , Animales , Animales Recién Nacidos , Huesos , Cartílago/crecimiento & desarrollo , Cartílago/metabolismo , Diferenciación Celular , Condrogénesis , Proteínas de Homeodominio/metabolismo , Humanos , Ratones , Ratones Noqueados , Ratones Transgénicos , Proteínas del Tejido Nervioso , Osteogénesis/fisiología , Proteínas/metabolismoRESUMEN
Embryonic myogenesis is a temporally and spatially regulated process that generates skeletal muscle of the trunk and limbs. During this process, mononucleated myoblasts derived from myogenic progenitor cells within the somites undergo proliferation, migration and differentiation to elongate and fuse into multinucleated functional myofibers. Skeletal muscle is the most abundant tissue of the body and has the remarkable ability to self-repair by re-activating the myogenic program in muscle stem cells, known as satellite cells. Post-transcriptional regulation of gene expression mediated by RNA-binding proteins is critically required for muscle development during embryogenesis and for muscle homeostasis in the adult. Differential subcellular localization and activity of RNA-binding proteins orchestrates target gene expression at multiple levels to regulate different steps of myogenesis. Dysfunctions of these post-transcriptional regulators impair muscle development and homeostasis, but also cause defects in motor neurons or the neuromuscular junction, resulting in muscle degeneration and neuromuscular disease. Many RNA-binding proteins, such as members of the muscle blind-like (MBNL) and CUG-BP and ETR-3-like factors (CELF) families, display both overlapping and distinct targets in muscle cells. Thus they function either cooperatively or antagonistically to coordinate myoblast proliferation and differentiation. Evidence is accumulating that the dynamic interplay of their regulatory activity may control the progression of myogenic program as well as stem cell quiescence and activation. Moreover, the role of RNA-binding proteins that regulate post-transcriptional modification in the myogenic program is far less understood as compared with transcription factors involved in myogenic specification and differentiation. Here we review past achievements and recent advances in understanding the functions of RNA-binding proteins during skeletal muscle development, regeneration and disease, with the aim to identify the fundamental questions that are still open for further investigations.
RESUMEN
Skeletal muscle has a remarkable capacity of regeneration after injury, but the regulatory network underlying this repair process remains elusive. RNA-binding proteins play key roles in the post-transcriptional regulation of gene expression and the maintenance of tissue homeostasis and plasticity. Rbm24 regulates myogenic differentiation during early development, but its implication in adult muscle is poorly understood. Here we show that it exerts multiple functions in muscle regeneration. Consistent with its dynamic subcellular localization during embryonic muscle development, Rbm24 also displays cytoplasm to nucleus translocation during C2C12 myoblast differentiation. In adult mice, Rbm24 mRNA is enriched in slow-twitch muscles along with myogenin mRNA. The protein displays nuclear localization in both slow and fast myofibers. Upon injury, Rbm24 is rapidly upregulated in regenerating myofibers and accumulates in the myonucleus of nascent myofibers. Through satellite cell transplantation, we demonstrate that Rbm24 functions sequentially to regulate myogenic differentiation and muscle regeneration. It is required for myogenin expression at early stages of muscle injury and for muscle-specific pre-mRNA alternative splicing at late stages of regeneration. These results identify Rbm24 as a multifaceted regulator of myoblast differentiation. They provide insights into the molecular pathway orchestrating the expression of myogenic factors and muscle functional proteins during regeneration.
Asunto(s)
Diferenciación Celular/genética , Desarrollo de Músculos/fisiología , Músculo Esquelético/crecimiento & desarrollo , Proteínas de Unión al ARN/metabolismo , Regeneración/fisiología , Animales , Regulación de la Expresión Génica/genética , Ratones , Desarrollo de Músculos/genética , Músculo Esquelético/metabolismo , Procesamiento Postranscripcional del ARN , Proteínas de Unión al ARN/genética , Células Madre/citologíaRESUMEN
Branchiomeric craniofacial muscles differ from all other skeletal muscles with respect to embryological origin, motor innervation and upstream activators of myogenesis. A series of recent studies has revealed a striking juxtaposition and overlapping genetic program of craniofacial skeletal muscle progenitor cells with a population of cells giving rise to cardiac muscle. The divergent myogenic fates of adjacent progenitor cells revealed by these data provide a new framework for the study of craniofacial myogenesis.
Asunto(s)
Región Branquial/embriología , Cabeza/embriología , Desarrollo de Músculos , Músculo Esquelético/embriología , Animales , Región Branquial/metabolismo , Regulación del Desarrollo de la Expresión Génica , Cabeza/fisiología , Corazón/embriología , Humanos , Modelos Biológicos , Desarrollo de Músculos/genética , Miocardio/metabolismo , Transducción de SeñalRESUMEN
RNA-binding proteins control the metabolism of RNAs at all stages of their lifetime. They are critically required for the post-transcriptional regulation of gene expression in a wide variety of physiological and pathological processes. Rbm24 is a highly conserved RNA-binding protein that displays strongly regionalized expression patterns and exhibits dynamic changes in subcellular localization during early development. There is increasing evidence that it acts as a multifunctional regulator to switch cell fate determination and to maintain tissue homeostasis. Dysfunction of Rbm24 disrupts cell differentiation in nearly every tissue where it is expressed, such as skeletal and cardiac muscles, and different head sensory organs, but the molecular events that are affected may vary in a tissue-specific, or even a stage-specific manner. Recent works using different animal models have uncovered multiple post-transcriptional regulatory mechanisms by which Rbm24 functions in key developmental processes. In particular, it represents a major splicing factor in muscle cell development, and plays an essential role in cytoplasmic polyadenylation during lens fiber cell terminal differentiation. Here we review the advances in understanding the implication of Rbm24 during development and disease, by focusing on its regulatory roles in physiological and pathological conditions.
Asunto(s)
Diferenciación Celular/genética , Linaje de la Célula/genética , Desarrollo Embrionario/genética , Regulación de la Expresión Génica , Homeostasis/genética , Poliadenilación/genética , Proteínas de Unión al ARN/genética , Empalme Alternativo/genética , Animales , Citoplasma/metabolismo , Humanos , Células Musculares/metabolismo , Proteínas de Unión al ARN/metabolismoRESUMEN
Muscle fibers show great differences in their contractile and metabolic properties. This diversity enables skeletal muscles to fulfill and adapt to different tasks. In this report, we show that the Six/Eya pathway is implicated in the establishment and maintenance of the fast-twitch skeletal muscle phenotype. We demonstrate that the MEF3/Six DNA binding element present in the aldolase A pM promoter mediates the high level of activation of this promoter in fast-twitch glycolytic (but not in slow-twitch) muscle fibers. We also show that among the Six and Eya gene products expressed in mouse skeletal muscle, Six1 and Eya1 proteins accumulate preferentially in the nuclei of fast-twitch muscles. The forced expression of Six1 and Eya1 together in the slow-twitch soleus muscle induced a fiber-type transition characterized by the replacement of myosin heavy chain I and IIA isoforms by the faster IIB and/or IIX isoforms, the activation of fast-twitch fiber-specific genes, and a switch toward glycolytic metabolism. Collectively, these data identify Six1 and Eya1 as the first transcriptional complex that is able to reprogram adult slow-twitch oxidative fibers toward a fast-twitch glycolytic phenotype.
Asunto(s)
Regulación de la Expresión Génica , Proteínas de Homeodominio/metabolismo , Fibras Musculares de Contracción Rápida/fisiología , Fibras Musculares de Contracción Lenta/fisiología , Regiones Promotoras Genéticas , Transactivadores/metabolismo , Animales , Núcleo Celular/metabolismo , Femenino , Fructosa-Bifosfato Aldolasa/genética , Genes Reporteros , Proteínas de Homeodominio/genética , Péptidos y Proteínas de Señalización Intracelular , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , Fibras Musculares de Contracción Rápida/citología , Fibras Musculares de Contracción Lenta/citología , Proteínas Nucleares , Fenotipo , Unión Proteica , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , Proteínas Tirosina Fosfatasas , Tacrolimus/metabolismo , Transactivadores/genética , TransgenesRESUMEN
MATERIALS AND METHODS: To investigate the genetic alterations that occur during the development of renal cell carcinomas (RCC), we used 20 microsatellite markers to examine 48 renal cell carcinomas for allelic losses of chromosome arm 14q. RESULTS: We identified 14q LOH in 31% of cases. Twelve tumors were entirely lacking the 14q arm and three were partially deleted. For the first time on fresh tumors, these findings led to the delineation of a 17.9 Mb region between markers D14S281 and D14S277 that is commonly deleted. Interestingly, this segment overlaps with the previously reported 37.8 Mb commonly deleted region. CONCLUSION: Taken together these results allowed us to define a new 2.8 Mb segment between markers D14S588 and D14S277 that potentially harbors a tumor suppressor gene involved in the development of RCC which can be reached by positional cloning.
Asunto(s)
Carcinoma de Células Renales/genética , Cromosomas Humanos Par 14 , Genes Supresores de Tumor , Neoplasias Renales/genética , Pérdida de Heterocigocidad/genética , Adulto , Anciano , Femenino , Marcadores Genéticos , Humanos , Masculino , Persona de Mediana EdadRESUMEN
RNA-binding proteins (RBP) contribute to gene regulation through post-transcriptional events. Despite the important roles demonstrated for several RBP in regulating skeletal myogenesis in vitro, very few RBP coding genes have been characterized during skeletal myogenesis in vertebrate embryo. In the present study we report that Rbm24, which encodes the RNA-binding motif protein 24, is required for skeletal muscle differentiation in vivo. We show that Rbm24 transcripts are expressed at all sites of skeletal muscle formation during embryogenesis of different vertebrates, including axial, limb and head muscles. Interestingly, we find that Rbm24 protein starts to accumulate in MyoD-positive myoblasts and is transiently expressed at the onset of muscle cell differentiation. It accumulates in myotomal and limb myogenic cells, but not in Pax3-positive progenitor cells. Rbm24 expression is under the direct regulation by MyoD, as demonstrated by in vivo chromatin immunoprecipitation assay. Using morpholino knockdown approach, we further show that Rbm24 is required for somitic myogenic progenitor cells to differentiate into muscle cells during chick somitic myogenesis. Altogether, these results highlight Rbm24 as a novel key regulator of the myogenic differentiation program during vertebrate development.
Asunto(s)
Desarrollo Embrionario/fisiología , Regulación del Desarrollo de la Expresión Génica , Desarrollo de Músculos/fisiología , Músculo Esquelético/embriología , Mioblastos/metabolismo , Proteínas de Unión al ARN/metabolismo , Animales , Diferenciación Celular , Embrión de Pollo , Músculo Esquelético/metabolismo , Factores Reguladores Miogénicos/genética , Factores Reguladores Miogénicos/metabolismo , Proteínas de Unión al ARN/genética , Xenopus , Proteínas de Xenopus/genética , Proteínas de Xenopus/metabolismoRESUMEN
Satellite cells (SCs) are stem cells that mediate skeletal muscle growth and regeneration. Here, we observe that adult quiescent SCs and their activated descendants expressed the homeodomain transcription factor Six1. Genetic disruption of Six1 specifically in adult SCs impaired myogenic cell differentiation, impaired myofiber repair during regeneration, and perturbed homeostasis of the stem cell niche, as indicated by an increase in SC self-renewal. Six1 regulated the expression of the myogenic regulatory factors MyoD and Myogenin, but not Myf5, which suggests that Six1 acts on divergent genetic networks in the embryo and in the adult. Moreover, we demonstrate that Six1 regulates the extracellular signal-regulated kinase 1/2 (ERK1/2) pathway during regeneration via direct control of Dusp6 transcription. Muscles lacking Dusp6 were able to regenerate properly but showed a marked increase in SC number after regeneration. We conclude that Six1 homeoproteins act as a rheostat system to ensure proper regeneration of the tissue and replenishment of the stem cell pool during the events that follow skeletal muscle trauma.
Asunto(s)
Proteínas de Homeodominio/metabolismo , Músculo Esquelético/fisiología , Regeneración/fisiología , Células Satélite del Músculo Esquelético/fisiología , Células Madre/fisiología , Cicatrización de Heridas/fisiología , Animales , Diferenciación Celular/genética , Diferenciación Celular/fisiología , Fosfatasa 6 de Especificidad Dual/genética , Fosfatasa 6 de Especificidad Dual/metabolismo , Proteínas de Homeodominio/genética , Homeostasis , Sistema de Señalización de MAP Quinasas/genética , Sistema de Señalización de MAP Quinasas/fisiología , Ratones , Ratones Endogámicos C57BL , Músculo Esquelético/metabolismo , Proteína MioD/genética , Proteína MioD/metabolismo , Miofibrillas/metabolismo , Miofibrillas/fisiología , Factor 5 Regulador Miogénico/genética , Factor 5 Regulador Miogénico/metabolismo , Factores Reguladores Miogénicos/genética , Factores Reguladores Miogénicos/metabolismo , Miogenina/genética , Miogenina/metabolismo , Regeneración/genética , Células Satélite del Músculo Esquelético/metabolismo , Células Madre/metabolismo , Factores de Transcripción/genética , Factores de Transcripción/metabolismo , Cicatrización de Heridas/genéticaRESUMEN
BACKGROUND: In vertebrates, the skeletal elements of the jaw, together with the connective tissues and tendons, originate from neural crest cells, while the associated muscles derive mainly from cranial mesoderm. Previous studies have shown that neural crest cells migrate in close association with cranial mesoderm and then circumscribe but do not penetrate the core of muscle precursor cells of the branchial arches at early stages of development, thus defining a sharp boundary between neural crest cells and mesodermal muscle progenitor cells. Tendons constitute one of the neural crest derivatives likely to interact with muscle formation. However, head tendon formation has not been studied, nor have tendon and muscle interactions in the head. METHODOLOGY/PRINCIPAL FINDINGS: Reinvestigation of the relationship between cranial neural crest cells and muscle precursor cells during development of the first branchial arch, using quail/chick chimeras and molecular markers revealed several novel features concerning the interface between neural crest cells and mesoderm. We observed that neural crest cells migrate into the cephalic mesoderm containing myogenic precursor cells, leading to the presence of neural crest cells inside the mesodermal core of the first branchial arch. We have also established that all the forming tendons associated with branchiomeric and eye muscles are of neural crest origin and express the Scleraxis marker in chick and mouse embryos. Moreover, analysis of Scleraxis expression in the absence of branchiomeric muscles in Tbx1(-/-) mutant mice, showed that muscles are not necessary for the initiation of tendon formation but are required for further tendon development. CONCLUSIONS/SIGNIFICANCE: This results show that neural crest cells and muscle progenitor cells are more extensively mixed than previously believed during arch development. In addition, our results show that interactions between muscles and tendons during craniofacial development are similar to those observed in the limb, despite the distinct embryological origin of these cell types in the head.
Asunto(s)
Mesodermo/citología , Desarrollo de Músculos/fisiología , Músculo Esquelético/embriología , Cresta Neural/citología , Cráneo/embriología , Animales , Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/genética , Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/metabolismo , Embrión de Pollo , Embrión de Mamíferos/metabolismo , Embrión no Mamífero/metabolismo , Regulación del Desarrollo de la Expresión Génica , Mesodermo/metabolismo , Ratones , Ratones Endogámicos , Músculo Esquelético/metabolismo , Proteína MioD/genética , Proteína MioD/metabolismo , Cresta Neural/metabolismo , Cráneo/metabolismo , Factores de Transcripción/genética , Factores de Transcripción/metabolismoRESUMEN
In mammals, Pax3, Six4, Six1 and Six5 genes are co-expressed with Eya1, Eya2 and Eya4 genes during mouse somitogenesis. To unravel the functions of Eya genes during muscle development, we analyzed myogenesis in Eya2-/- and in Eya1-/- embryos. A delay in limb myogenesis was observed between E10 and E13 in Eya1-/- embryos only, that is later compensated. Compound E18 Eya1-/-Eya2-/+ fetuses present a muscle phenotype comparable with that of Six1-/- fetuses; lacking a diaphragm and with a specific absence of limb muscles, suggesting either genetic epistasis between Six and Eya genes, or biochemical interactions between Six and Eya proteins. We tested these two non-exclusive possibilities. First, we show that Six proteins recruit Eya proteins to drive transcription during embryogenesis in the dermomyotomal epaxial and hypaxial lips of the somites by binding MEF3 DNA sites. Second, we show that Pax3 expression is lost in the ventrolateral (hypaxial) dermomyotomes of the somite in both Eya1-/-Eya2-/- embryos and in Six1-/-Six4-/- embryos, precluding hypaxial lip formation. This structure, from which myogenic cells delaminate to invade the limb does not form in these double mutant embryos, leading to limb buds without myogenic progenitor cells. Eya1 and Eya2, however, are still expressed in the somites of Six1Six4 double mutant and in splotch embryos, and Six1 is expressed in the somites of Eya1Eya2 double mutant embryos and in splotch embryos. Altogether these results show that Six and Eya genes lie genetically upstream of Pax3 gene in the formation of ventrolateral dermomyotome hypaxial lips. No genetic links have been characterized between Six and Eya genes, but corresponding proteins activate key muscle determination genes (Myod, Myogenin and Mrf4). These results establish a new hierarchy of genes controlling early steps of hypaxial myogenic commitment in the mouse embryo.
Asunto(s)
Péptidos y Proteínas de Señalización Intracelular/fisiología , Músculo Esquelético/fisiología , Proteínas Nucleares/fisiología , Proteínas Tirosina Fosfatasas/fisiología , Somitos/fisiología , Animales , Tipificación del Cuerpo/fisiología , Movimiento Celular/fisiología , Elementos de Facilitación Genéticos , Extremidades/embriología , Extremidades/fisiología , Proteínas de Homeodominio/genética , Proteínas de Homeodominio/metabolismo , Péptidos y Proteínas de Señalización Intracelular/genética , Ratones , Ratones Noqueados , Desarrollo de Músculos , Músculo Esquelético/embriología , Proteína MioD/metabolismo , Factores Reguladores Miogénicos/metabolismo , Miogenina/metabolismo , Proteínas Nucleares/genética , Factor de Transcripción PAX3 , Factores de Transcripción Paired Box/metabolismo , Regiones Promotoras Genéticas , Proteínas Tirosina Fosfatasas/genéticaRESUMEN
Previous studies have suggested a role of the homeodomain Six family proteins in patterning the developing vertebrate head that involves appropriate segmentation of three tissue layers, the endoderm, the paraxial mesoderm and the neural crest cells; however, the developmental programs and mechanisms by which the Six genes act in the pharyngeal endoderm remain largely unknown. Here, we examined their roles in pharyngeal pouch development. Six1-/- mice lack thymus and parathyroid and analysis of Six1-/- third pouch endoderm demonstrated that the patterning of the third pouch into thymus/parathyroid primordia is initiated. However, the endodermal cells of the thymus/parathyroid rudiments fail to maintain the expression of the parathyroid-specific gene Gcm2 and the thymus-specific gene Foxn1 and subsequently undergo abnormal apoptosis, leading to a complete disappearance of organ primordia by E12.5. This thus defines the thymus/parathyroid defects present in the Six1 mutant. Analyses of the thymus/parathyroid development in Six1-/-;Six4-/- double mutant show that both Six1 and Six4 act synergistically to control morphogenetic movements of early thymus/parathyroid tissues, and the threshold of Six1/Six4 appears to be crucial for the regulation of the organ primordia-specific gene expression. Previous studies in flies and mice suggested that Eya and Six genes may function downstream of Pax genes. Our data clearly show that Eya1 and Six1 expression in the pouches does not require Pax1/Pax9 function, suggesting that they may function independently from Pax1/Pax9. In contrast, Pax1 expression in all pharyngeal pouches requires both Eya1 and Six1 function. Moreover, we show that the expression of Tbx1, Fgf8 and Wnt5b in the pouch endoderm was normal in Six1-/- embryos and slightly reduced in Six1-/-;Six4-/- double mutant, but was largely reduced in Eya1-/- embryos. These results indicate that Eya1 appears to be upstream of very early events in the initiation of thymus/parathyroid organogenesis, while Six genes appear to act in an early differentiation step during thymus/parathyroid morphogenesis. Together, these analyses establish an essential role for Eya1 and Six genes in patterning the third pouch into organ-specific primordia.
Asunto(s)
Región Branquial/embriología , Proteínas de Homeodominio/genética , Péptidos y Proteínas de Señalización Intracelular/genética , Proteínas Nucleares/genética , Glándulas Paratiroides/embriología , Proteínas Tirosina Fosfatasas/genética , Timo/embriología , Transactivadores/genética , Animales , Tipificación del Cuerpo/genética , Región Branquial/metabolismo , Factor 8 de Crecimiento de Fibroblastos/genética , Factores de Transcripción Forkhead/genética , Edad Gestacional , Glicoproteínas/genética , Proteínas de Homeodominio/metabolismo , Péptidos y Proteínas de Señalización Intracelular/deficiencia , Péptidos y Proteínas de Señalización Intracelular/metabolismo , Ratones , Ratones Noqueados , Ratones Mutantes , Modelos Biológicos , Proteínas Nucleares/deficiencia , Proteínas Nucleares/metabolismo , Factores de Transcripción Paired Box/genética , Glándulas Paratiroides/metabolismo , Proteínas Tirosina Fosfatasas/deficiencia , Proteínas Tirosina Fosfatasas/metabolismo , Proteínas de Dominio T Box/genética , Timo/metabolismo , Transactivadores/deficiencia , Transactivadores/metabolismo , Factores de Transcripción/genética , Proteínas Wnt/genéticaRESUMEN
In mammals, Six5, Six4 and Six1 genes are co-expressed during mouse myogenesis. Six4 and Six5 single knockout (KO) mice have no developmental defects, while Six1 KO mice die at birth and show multiple organ developmental defects. We have generated Six1Six4 double KO mice and show an aggravation of the phenotype previously reported for the single Six1 KO. Six1Six4 double KO mice are characterized by severe craniofacial and rib defects, and general muscle hypoplasia. At the limb bud level, Six1 and Six4 homeogenes control early steps of myogenic cell delamination and migration from the somite through the control of Pax3 gene expression. Impaired in their migratory pathway, cells of the somitic ventrolateral dermomyotome are rerouted, lose their identity and die by apoptosis. At the interlimb level, epaxial Met expression is abolished, while it is preserved in Pax3-deficient embryos. Within the myotome, absence of Six1 and Six4 impairs the expression of the myogenic regulatory factors myogenin and Myod1, and Mrf4 expression becomes undetectable. Myf5 expression is correctly initiated but becomes restricted to the caudal region of each somite. Early syndetomal expression of scleraxis is reduced in the Six1Six4 embryo, while the myotomal expression of Fgfr4 and Fgf8 but not Fgf4 and Fgf6 is maintained. These results highlight the different roles played by Six proteins during skeletal myogenesis.