Muscle-Derived Extracellular Signal-Regulated Kinases 1 and 2 Are Required for the Maintenance of Adult Myofibers and Their Neuromuscular Junctions

Abstract

The Ras-extracellular signal-regulated kinase 1 and 2 (ERK1/2) pathway appears to be important for the development, maintenance, aging, and pathology of mammalian skeletal muscle. Yet no gene targeting of Erk1/2 in muscle fibers in vivo has been reported to date. We combined a germ line Erk1 mutation with Cre-loxP Erk2 inactivation in skeletal muscle to produce, for the first time, mice lacking ERK1/2 selectively in skeletal myofibers. Animals lacking muscle ERK1/2 displayed stunted postnatal growth, muscle weakness, and a shorter life span. Their muscles examined in this study, sternomastoid and tibialis anterior, displayed fragmented neuromuscular synapses and a mixture of modest fiber atrophy and loss but failed to show major changes in fiber type composition or absence of cell surface dystrophin. Whereas the lack of only ERK1 had no effects on the phenotypes studied, the lack of myofiber ERK2 explained synaptic fragmentation in the sternomastoid but not the tibialis anterior and a decrease in the expression of the acetylcholine receptor (AChR) epsilon subunit gene mRNA in both muscles. A reduction in AChR protein was documented in line with the above mRNA results. Evidence of partial denervation was found in the sternomastoid but not the tibialis anterior. Thus, myofiber ERK1/2 are differentially required for the maintenance of myofibers and neuromuscular synapses in adult mice. Mitogen-activated protein kinases (MAPKs) are components of intracellular signaling modules that control a myriad of cellular processes. MAPK modules consist of 3 core protein kinase components. The most downstream is the actual MAPK, an S/T kinase that phosphorylates the transcription factors, cytoskeletal elements, or other kinases that are the targets of regulation by signaling cascades started at the cell surface. A MAPK is activated by an upstream MAPK kinase (MAP2K), which, in turn, is activated by a MAP2K kinase (MAP3K). MAP3Ks are usually at the receiving end of signals derived from small, monomeric GTPases such as the Ras family or by other more intricate mechanisms (1). In mammalian cells, the prototypical MAPK module is composed of the MAPKs extracellular signal-regulated kinases 1 and 2 (ERK1/2), the MAP2Ks MEK1/2, and the MAP3K Raf. ERK1/2 regulate normal cellular responses to multiple growth factors and cytokines in proliferation, differentiation, and apoptosis (2, 3).Multiple studies suggest an important role for the Ras-ERK1/2 pathway in the development, normal maintenance, aging, and pathology of mammalian skeletal muscle. Thus, ERK1/2 activity has both stimulatory and inhibitory roles in the differentiation of cultured skeletal myotubes that vary with the stage of this protracted process (4-8). ERK1/2 have been implicated in the maintenance of adult skeletal muscle mass (9) and, seemingly paradoxically, in the control of both the fast-twitch (10) and the slow-twitch (11) fiber type phenotypes. Alterations in levels of ERK1/2 activity in aging rodent muscle correlate with sarcopenia (12), the loss of muscle mass and ...

Defective Acetylcholine Receptor Subunit Switch Precedes Atrophy of Slow-Twitch Skeletal Muscle Fibers Lacking ERK1/2 Kinases in Soleus Muscle

“…Cre was driven by the human α-skeletal muscle actin ( Hsa ) promoter, which is expressed only in myofibers, and not in other cells in the muscle tissue, starting around embryonic day 9.5131415. In the fast-twitch STN and TA we measured a ~90% reduction in ERK2 levels in mutants relative to control10. In DKO SOL, ERK2 levels were ~70% lower than in control (Fig.…”

“…1). In our initial characterization of these animals, we found they could be divided into two groups according to how fast they lost weight10. “Severe” animals lost weight quickly starting at about 7 weeks of age, while “mild” double mutants were able to keep their weight at that age but clearly failed to keep up with controls.…”

“…These results were consistent with a role for ERK1/2 in the maintenance of muscle mass, but not of the fast-twitch fiber phenotype, and demonstrated an important role for ERK1/2 in keeping the structural integrity of the mature NMJ in vivo . In our previous experiments10, we did not study the effects of lack of ERK1/2 on predominantly slow-twitch muscles, nor could we discern whether the phenotypes observed were primarily derived from synaptic or extrasynaptic functions of myofiber ERK1/2.…”

“…ERK1/2 have been implicated in the maintenance of adult skeletal muscle mass7 and, seemingly paradoxically, in the control of both the fast-twitch (type 2)8 and the slow-twitch (type 1)9 fiber type phenotypes. Previously10, we generated mice genetically deficient in myofiber ERK1/2. These animals survived development but displayed stunted postnatal growth, muscle weakness and shorter lifespan.…”

Schwann cells participate in synapse elimination at the developing neuromuscular junction

“…These observations suggest that the level of expression of NRG1-III by motor neurons might be a mechanism for controlling the behavior of tSCs even in the adult. Since the same fragmented morphology seen in these adult junctions occurs in a variety of experimental and pathological conditions, including alterations in transcription factors, cell adhesion molecules, and in various dystrophies and myopathies [38–40], it is tempting to speculate that tSC behaviors could be responsible for the remodeling of NMJs in various pathologies.…”