Ventral axial organs regulate expression of myotomal Fgf-8 that influences rib development

Fgf-8 encodes a secreted signaling molecule mediating key roles in embryonic patterning. This study analyzes the expression pattern, regulation, and function of this growth factor in the paraxial mesoderm of the avian embryo. In the mature somite, expression of Fgf-8 is restricted to a subpopulation of myotome cells, comprising most, but not all, epaxial and hypaxial muscle precursors. Following ablation of the notochord and floor plate, Fgf-8 expression is not activated in the somites, in either the epaxial or the hypaxial domain, while ablation of the dorsal neural tube does not affect Fgf-8 expression in paraxial mesoderm. Contrary to the view that hypaxial muscle precursors are independent of regulatory influences from axial structures, these findings provide the first evidence for a regulatory influence of ventral, but not dorsal axial structures on the hypaxial muscle domain. Sonic hedgehog can substitute for the ventral neural tube and notochord in the initiation of Fgf-8 expression in the myotome. It is also shown that Fgf-8 protein leads to an increase in sclerotomal cell proliferation and enhances rib cartilage development in mature somites, whereas inhibition of Fgf signaling by SU 5402 causes deletions in developing ribs. These observations demonstrate: (1) a regulatory influence of the ventral axial organs on the hypaxial muscle compartment; (2) regulation of epaxial and hypaxial expression of Fgf-8 by Sonic hedgehog; and (3) independent regulation of Fgf-8 and MyoD in the hypaxial myotome by ventral axial organs. It is postulated that the notochord and ventral neural tube influence hypaxial expression of Fgf-8 in the myotome and that, in turn, Fgf-8 has a functional role in rib formation.     

The NLRR gene family and mouse development: Modified differential display PCR identifies NLRR-1 as a gene expressed in early somitic myoblasts

During vertebrate embryogenesis, the somites form by segmentation of the trunk mesoderm, lateral to the neural tube, in an anterior to posterior direction. Analysis of differential gene expression during somitogenesis has been problematic due to the limited amount of tissue available from early mouse embryos. To circumvent these problems, we developed a modified differential display PCR technique that is highly sensitive and yields products that can be used directly as in situ hybridisation probes. Using this technique, we isolated NLRR-1 as a gene expressed in the myotome of developing somites but not in the presomitic mesoderm. Detailed expression analysis showed that this gene was expressed in the skeletal muscle precursors of the myotome, branchial arches and limbs as well as in the developing nervous system. Somitic expression occurs in the earliest myoblasts that originate from the dorsal lip in a pattern reminiscent of the muscle determination gene Myf5, but not at the ventral lip, indicating that NLRR-1 is expressed in a subset of myotome cells. The NLRR genes comprise a three-gene family encoding glycosylated transmembrane proteins with external leucine-rich repeats, a fibronectin domain, an immunoglobulin domain and short intracellular tails capable of mediating protein-protein interaction. Analysis of NLRR-3 expression revealed regulated expression in the neural system in developing ganglia and motor neurons. NLRR-2 expression appears to be predominately confined to the adult. The regulated embryonic expression and cellular location of these proteins suggest important roles during mouse development in the control of cell adhesion, movement or signalling.     

Control of morphogenetic cell movements in the early zebrafish myotome

As the vertebrate myotome is generated, myogenic precursor cells undergo extensive and coordinated movements as they differentiate into properly positioned embryonic muscle fibers. In the zebrafish, the "adaxial" cells adjacent to the notochord are the first muscle precursors to be specified. After initially differentiating into slow-twitch myosin-expressing muscle fibers, these cells have been shown to undergo a remarkable radial migration through the lateral somite, to populate the superficial layer of slow-twitch muscle of the mature myotome. Here we characterize an earlier set of adaxial cell behaviors; the transition from a roughly 4x5 array of cuboidal cells to a 1x20 stack of elongated cells, prior to the migration event. We find that adaxial cells display a highly stereotypical series of behaviors as they undergo this rearrangement. Furthermore, we show that the actin regulatory molecule, Cap1, is specifically expressed in adaxial cells and is required for the progression of these behaviors. The requirement of Cap1 for a cellular apical constriction step is reminiscent of similar requirements of Cap during apical constriction in Drosophila development, suggesting a conservation of gene function for a cell biological event critical to many developmental processes.     

Medial pioneer fibers pattern the morphogenesis of early myoblasts derived from the lateral somite

The first wave of myoblasts which constitutes the post-mitotic myotome stems from the medial epithelial somite. Whereas medial pioneers extend throughout the entire mediolateral myotome at cervical and limb levels, at flank regions they are complemented laterally by a population of early myoblasts emerging from the lateral epithelial somite. These myoblasts delaminate underneath the nascent dermomyotome and become post-mitotic. They are Myf5-positive but express MyoD and desmin only a day later while differentiating into fibers. Overexpression of Noggin in the lateral somite triggers their premature differentiation suggesting that lateral plate-BMP4 maintains them in an undifferentiated state. Moreover, directly accelerating their differentiation by MyoD overexpression prior to arrival of medial fibers, generates a severely mispatterned lateral myotome. This is in contrast to medial pioneers that have the capacity for self-organization. Furthermore, inhibiting differentiation of medial pioneers with dominant-negative MyoD also disrupts lateral myoblast patterning and differentiation. Thus, we propose that medial pioneers are needed for proper morphogenesis of the lateral population which is kept as undifferentiated mesenchyme by BMP4 until their arrival. In addition, medial pioneers also organize dermomyotome lip-derived fibers suggesting that they have a general role in patterning myotome development.     

Interactions between muscle fibers and segment boundaries in zebrafish

The most obvious segmental structures in the vertebrate embryo are somites: transient structures that give rise to vertebrae and much of the musculature. In zebrafish, most somitic cells give rise to long muscle fibers that are anchored to intersegmental boundaries. Therefore, this boundary is analogous to the mammalian tendon in that it transduces muscle-generated force to the skeletal system. We have investigated interactions between somite boundaries and muscle fibers. We define three stages of segment boundary formation. The first stage is the formation of the initial epithelial somite boundary. The second "transition" stage involves both the elongation of initially round muscle precursor cells and somite boundary maturation. The third stage is myotome boundary formation, where the boundary becomes rich in extracellular matrix and all muscle precursor cells have elongated to form long muscle fibers. It is known that formation of the initial epithelial somite boundary requires Notch signaling; vertebrate Notch pathway mutants show severe defects in somitogenesis. However, many zebrafish Notch pathway mutants are homozygous viable suggesting that segmentation of their larval and adult body plans at least partially recovers. We show that epithelial somite boundary formation and slow-twitch muscle morphogenesis are initially disrupted in after eight (aei) mutant embryos (which lack function of the Notch ligand, DeltaD); however, myotome boundaries form later ("recover") in a Hedgehog-dependent fashion. Inhibition of Hedgehog-induced slow muscle induction in aei/deltaD and deadly seven (des)/notch1a mutant embryos suggests that slow muscle is necessary for myotome boundary recovery in the absence of initial epithelial somite boundary formation. Because we have previously demonstrated that slow muscle migration triggers fast muscle cell elongation in zebrafish, we hypothesize that migrating slow muscle facilitates myotome boundary formation in aei/deltaD mutant embryos by patterning coordinated fast muscle cell elongation. In addition, we utilized genetic mosaic analysis to show that somite boundaries also function to limit the extent to which fast muscle cells can elongate. Combined, our results indicate that multiple interactions between somite boundaries and muscle fibers mediate zebrafish segmentation.