Date of Award

2015

Degree Type

Dissertation

Degree Name

Doctor of Philosophy (PhD)

Graduate Group

Cell & Molecular Biology

First Advisor

Elisabeth R. Barton

Second Advisor

H. Lee Sweeney

Abstract

Loss of gamma-sarcoglycan (g-SG) is common to both Duchenne and Limb Girdle muscular dystrophies, and is sufficient to induce severe muscle degeneration and signaling defects in response to mechanical load without causing susceptibility to contractile damage. This suggests that disease occurs not by structural deficits, but through aberrant signaling, namely, disruption of normal mechanotransduction signaling through the SG complex. However, the mechanisms of g-SG-mediated mechanical signaling are poorly understood. Here, we identified the muscle-specific protein archvillin as a g-SG and dystrophic interacting protein. Archvillin expression is dependent on g-SG, where levels are significantly unregulated in human LGMD2C patient muscle and at the sarcolemma of murine g-SG-null (gsg-/-) muscle. However, archvillin is delocalized in mdx muscle, indicating dystrophin-dependent sarcolemmal localization. In situ eccentric contraction (ECC) of C57 mouse tibialis anterior (TA) muscles causes ERK1/2 phosphorylation, nuclear activation of P-ERK1/2, and stimulus-dependent archvillin association with P-ERK1/2, whereas TA muscles from gsg-/- and mdx mice exhibit heightened P-ERK1/2 and increased nuclear P-ERK1/2 localization following ECCs, but the archvillin-P-ERK1/2 association is completely ablated. These results position archvillin as a mechanically sensitive component of the dystrophin complex, and demonstrate that signaling defects caused by loss of g-SG occur both at the sarcolemma and in the nucleus. We also extended our previous studies on mechanosensitive, g-SG dependent ERK1/2 phosphorylation to determine whether additional pathways are altered with loss of g-SG. Using a passive stretching protocol to isolate the effects of externally applied tension, we found that in isolated EDL muscles, Akt, S6RP, and p70S6K activation increases with stretch in both C57 and gsg-/- isolated muscles. Treatment of muscles with or without rapamycin during stretch blocks p70S6K activation in stretched C57 muscles, and reduces downstream S6RP phosphorylation, indicating mTOR dependence. However, even though rapamycin treatment decreases p70S6K activation in stretched gsg-/- muscles, S6RP phosphorylation remains elevated. These results suggest that p70S6K is an important component of g-SG-dependent mechanotransduction in skeletal muscle, where loss of g-SG uncouples the response of p70S6K to stretch and implies that g-SG is important for inactivation of this pathway. In sum, this study demonstrates altered load-sensing mechanisms in muscular dystrophies where the sarcoglycans are absent.

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