Assembly of Myotendinous Junctions in the Chick Embryo: Deposition of P68 Is an Early Event in Myotendinous Junction Formation

doi:10.1006/dbio.1994.1161

Developmental Biology

Volume 163, Issue 2, June 1994, Pages 447-456

https://doi.org/10.1006/dbio.1994.1161 Get rights and content

Abstract

Myotendinous junctions (MTJs) are specialized sites at the surface of muscle fibers where force is transmitted between muscle and tendon. Morphogenesis of MTJs in the latter half of embryonic chick development is characterized by formation of basement membrane, appearance of subsarcolemmal densities, association of myofibrils with the membrane, and then membrane folding. In the present investigation, the time of appearance of proteins involved in force transmission at MTJs is compared to those previously described structural specializations occurring during MTJ development. In addition, P68, a recently discovered collagen-binding protein that is shown to be identical or closely related to laminin-binding lectin is also demonstrated at MTJs, and an improved technique for P68 purification is presented. P68 accumulates on the surface of collagen fibers that lie near the ends of myotubes in chick hindlimb muscle prior to the appearance of ultrastructurally discernible basement membrane. At Day 10, the stage when MTJ basement membrane is first discernible by electron microscopy, laminin and the β1 subunit of integrin are detectable at the MTJ. At this stage, laminin distribution coincides with P68 distribution and no laminin is detectable at sites other than the forming MTJs. Fibronectin is detectable at the MTJ at Day 12, but is not enriched at the MTJ relative to other sites on the myotubes. At Day 13, the stage at which subsarcolemmal densities first appear, talin is demonstrable at the MTJ and fibronectin concentration at the MTJ is enriched. Collagen type IV cannot be discerned at the MTJ until Day 15. Thus, the earliest demonstrable specializations at sites of MTJ formation are: (1) the appearance of P68 on collagen fibers that lie at the ends of myotubes, which occurs prior to ultrastructurally discernible specializations at the MTJ and (2) the accumulation of laminin at the MTJ with a distribution identical to that of P68 at the stage of MTJ formation in which basement membrane first appears. These findings indicate that P68 and laminin may mediate early events in MTJ assembly.

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Cited by (16)

Fine structure of myotendinous junction between the anterior belly of the digastric muscle and intermediate tendon in adults rats
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Citation Excerpt :
Analyzing the MTJ interface, terminal myofibrils bundles presented at their extremity, elongated sarcoplasmic projections with finger-like processes, surrounded by interdigitations that represent sarcoplasmic invaginations (sarcolemma folds) from the extracellular matrix, with shearing aspect that characterizes the MTJ region (Ciena et al., 2010). According to Tidball (1994), invaginations promote the folds formation, which are the extracellular matrix projections named as interdigitations, placed between the projections of sarcoplasmic-shaped “fingers” that provide functionally, a better relationship between the myofibrils bundles to connective tissue, increasing the contact area in the muscle-tendon interface. However, it was observed in the examined samples, beyond the traditional MTJ components, atypical ultrastructural characteristics of MTJ, not yet reported in the literature.
This study analyzed the ultrastructural characteristics of the myotendinous junction (MTJ) between anterior belly of digastrics muscle and the intermediate tendon in adult rats. Six male Wistar rats were used and were anesthetized with an overdose of urethane and sacrificed by intracardiac perfusion with modified Karnovsky solution, postfixed in 1% osmium tetroxide, dehydrated in increasing series of alcohols and embedded in Spurr resin for transmission electron microscopic analysis. Ultrastructural analysis showed conical shape of the fiber extremity in MTJ region, highlighting the presence of numerous mitochondria arranged in groups in the subsarcolemmal and intermyofibrillary regions. Atypical MTJ characteristics were seen interspersed with bundles of collagen fibers. Classic characteristics such as finger-like processes by means of sarcoplasmic projections were observed among interdigitations. Terminals and periphericals bundles of myofibrils showed close relationship with the adjacent muscle fiber's endomysium through lateral junctions. In the distal portion, it was observed that the communication region of microtendons forming the intermediate tendon of digastric muscle, and it can highlight the columns disposition of tenocytes. In conclusion, the MTJ ultrastructure between the anterior belly of digastric muscle and intermediate tendon of adult rats showed classical morphologic descriptions and presented an atypical region revealed by the subspecialization between the myofibrils bundles and collagen fibers in the MTJ region.
Zipper nonmuscle myosin-II functions downstream of PS2 integrin in Drosophila myogenesis and is necessary for myofibril formation
2001, Developmental Biology
Nonmuscle myosin-II is a key motor protein that drives cell shape change and cell movement. Here, we analyze the function of nonmuscle myosin-II during Drosophila embryonic myogenesis. We find that nonmuscle myosin-II and the adhesion molecule, PS2 integrin, colocalize at the developing muscle termini. In the paradigm emerging from cultured fibroblasts, nonmuscle actomyosin-II contractility, mediated by the small GTPase Rho, is required to cluster integrins at focal adhesions. In direct opposition to this model, we find that neither nonmuscle myosin-II nor RhoA appear to function in PS2 clustering. Instead, PS2 integrin is required for the maintenance of nonmuscle myosin-II localization and we show that the cytoplasmic tail of the β_PS integrin subunit is capable of mediating this PS2 integrin function. We show that embryos that lack zygotic expression of nonmuscle myosin-II fail to form striated myofibrils. In keeping with this, we demonstrate that a PS2 mutant that specifically disrupts myofibril formation is unable to mediate proper localization of nonmuscle myosin-II at the muscle termini. In contrast, embryos that lack RhoA function do generate striated muscles. Finally, we find that nonmuscle myosin-II localizes to the Z-line in mature larval muscle. We suggest that nonmuscle myosin-II functions at the muscle termini and the Z-line as an actin crosslinker and acts to maintain the structural integrity of the sarcomere.
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Tetranectin, a plasminogen-binding protein with a C-type lectin domain, is found in both serum and the extracellular matrix. In the present study we report that tetranectin is closely associated with myogenesis during embryonic development, skeletal muscle regeneration, and muscle cell differentiationin vitro.We find that tetranectin expression coincides with muscle differentiation and maturation in the second half of gestation and further that tetranectin is enriched at the myotendinous and myofascial junctions. The tetranectin immunostaining declines after birth and no immunostaining is observed in normal adult muscle. However, during skeletal muscle regeneration induced by the intramuscular injection of the myotoxic anesthetic Marcaine, myoblasts, myotubes, and the stumps of damaged myofibers exhibit intense tetranectin immunostaining. Tetranectin is also present in regenerating muscle cells in dystrophicmdxmice. Murine C2C12 myogenic cells and pluripotent embryonic stem cells can undergo muscle cell differentiationin vitro.Tetranectin is not expressed in the undifferentiated myogenic cells, but during the progression of muscle differentiation, tetranectin mRNA is induced, and both cytoplasmic and cell surface tetranectin immunostaining become apparent. Finally, we demonstrate that while tetranectin mRNA is translated to a similar degree in developing limbs and lung, the protein does not seem to be tissue associated in the lung as it is in the limbs. This indicates that in some tissues, such as the limbs, tetranectin may function locally, whereas in other tissues, such as the lung, tetranectin production may be destined for body fluids. In summary, these results suggest that tetranectin is a matricellular protein and plays a role in myogenesis.
Platelet-derived growth factor-stimulated secretion of basement membrane proteins by skeletal muscle occurs by tyrosine kinase-dependent and - independent pathways
1997, Journal of Biological Chemistry
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Full PapersAssembly of Myotendinous Junctions in the Chick Embryo: Deposition of P68 Is an Early Event in Myotendinous Junction Formation

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Assembly of Myotendinous Junctions in the Chick Embryo: Deposition of P68 Is an Early Event in Myotendinous Junction Formation