Date on Master's Thesis/Doctoral Dissertation

12-2014

Document Type

Doctoral Dissertation

Degree Name

Ph. D.

Department

Anatomical Sciences and Neurobiology

Committee Chair

Kumar, Ashok

Committee Co-Chair (if applicable)

Friedland, Robert P.

Committee Member

Lundy, Robert F.

Committee Member

Moore, J. Patrick

Committee Member

Petruska, Jeffrey C.

Subject

Myogenesis; Cellular signal transduction

Abstract

Skeletal muscle constitutes a highly plastic and malleable tissue that rapidly and profoundly adapts to various environmental and physiological stimuli. This response takes place by means of regulated signaling transduction which elicits changes in gene expression, biochemical, and metabolic properties. Adult muscle is composed by alignment of multinucleated, post mitotic muscle fibers formed during embryonic and neonatal development. Residing in the niche of mature muscle fibers is a small population of undifferentiated muscle progenitors termed satellite cells. Satellite cells are muscle stem cells that carry out regenerative response in adult myofibers. Coordinated signaling emitting from terminally differentiated muscle fibers and un-differentiated muscle progenitors sustains muscle homeostasis. De-regulation in the components of this synchronized signaling machinery manifests in deleterious disease states and myopathies. The main focus of my research has been towards understanding the role of the adaptor protein, tumor necrosis factor-associated factor 6 (TRAF6) in regulation of regenerative myogenesis. Utilizing the Cre-loxP system, we generated TRAF6 musclespecific knock-out mice: TRAF6mko, and TRAF6 satellite cell-specific knock-out mice: TRAF6scko. Our findings displayed distinct roles of TRAF6 in differentiated myofibers vs. satellite cells. Deletion of TRAF6 under the control of muscle creatine kinase (MCK) promoter (differentiated muscle) led to improvement in muscle regeneration in wildtype (WT) mice following induced muscle injury. This improvement was brought about by prevalence of an anti-inflammatory environment characterized by reduced signaling from inflammatory pathways such as NF-B and a domination of pro-regeneration M2c over proinflammatory M1 macrophages. Regenerating muscle of TRAF6mko mice also displayed a higher up-regulation in Notch signaling family proteins and target genes which in turn enhanced the activation of satellite cells leading to accelerated regeneration. Similar results were also observed upon deletion of TRAF6 in differentiated muscle of mdx mice (modelfor Duchenne muscular dystrophy). However, mdx;TRAF6mko muscle later displayed exacerbated signs of myopathy possibly due to diminished autophagy which has been associated with progressive myopathy in mdx mice. Employing the same models of muscle injury, our analysis revealed an absolute requirement of TRAF6 in satellite cell function during regenerative myogenesis. Contrary to its role in differentiated muscle, injured muscle of TRAF6scko mice displayed impaired regeneration due to a cell-autonomous defect in TRAF6-deficient satellite cells. Deletion of TRAF6 in satellite cells dramatically repressed the levels of Pax7 resulting in precocious differentiation and progressive depletion of the satellite cell reservoir. Myopathy was further exaggerated in dystrophic muscle of mdx;TRAF6scko due to satellite cell dysfunction. Our results further demonstrate that TRAF6 regulates proliferation and self-renewal of satellite cells through activation of mitogen-activated protein kinases (MAPKs) such as ERK1/2 and JNK and through modulating the levels of muscle specific microRNA’s: miR-1, miR-206, and mir-133a. The findings of this work highlight the importance of implementing a holistic approach upon development of target-based therapies. Oversight of the fundamental roles of targeted molecules has been associated with virulent outcomes. Developing a universal understanding of the various implications of individual molecules maximizes therapeutic potential.

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