Mechanical stimuli play a major role in the regulation of skeletal muscle mass, and the maintenance of muscle mass contributes significantly to disease prevention and the quality of life. Although the link between mechanical stimulation and the regulation of muscle mass has been recognized for decades, the molecular mechanisms underlying this process are not known. Hence, the long-term goal of our research is to define the molecular events through which mechanical stimuli regulate skeletal muscle mass. The primary objective of this project is to define the role of the mammalian target of rapamycin (mTOR) in regulating skeletal muscle mass, and determine how mechanical stimuli activate mTOR signaling. Our rationale for focusing on mTOR comes from our preliminary studies which suggest that: i) the activation of mTOR plays a critical role in mechanically-induced growth, and ii) mechanical stimuli activate mTOR signaling through a unique PI3K/PKB- independent mechanism involving phosphatidic acid (PA). Given that mechanical stimuli activate mTOR, it follows that a molecular mechanism (i.e. mechanotransduction pathway) exists for converting mechanical signals into mTOR activation. Thus, our plan is to identify the critical events in this pathway and determine if mimicking these events can induce muscle growth and attenuate disuse atrophy. Our current hypothesis is that mechanical stimuli promote an increase in phosphatidic acid (PA) which subsequently activates mTOR signaling and ultimately growth. To test this hypothesis we will pursue the following four specific aims: 1) Determine if the activation of mTOR is sufficient to induce growth and attenuate disuse atrophy;2) Define the role of mTOR in mechanically-induced growth;3) Determine if an increase in [PA] is sufficient to induce growth and attenuate disuse atrophy and 4) Identify the upstream molecules that regulate the mechanical activation of mTOR. In the first aim, over-expression of Rheb will be used to induce a PI3K/PKB-independent activation of mTOR in mouse skeletal muscles, and the resulting effect on muscle mass during normal use and disuse will be determined. In the second aim, transgenic mice expressing various mutants of mTOR will be used to define the muscle specific role of mTOR, and mTOR kinase activity, in mechanically-induced growth. In the third aim, over-expression of PA synthesizing enzymes will be used to determine if an increase in [PA] is sufficient to induce growth and attenuate disuse atrophy. In the fourth aim, activity assays will be used to identify the enzymes that regulate mechanically-induced changes in PA, and then pharmacological and molecular interventions will be used to further define the role that these enzymes play in the mechanical activation of mTOR. The proposed studies are significant because the outcomes will fill major gaps in our current knowledge of how mechanical stimuli regulate mTOR signaling and skeletal muscle mass. Furthermore, the outcomes could lead to the identification of targets for therapies that mimic the effects of mechanical stimuli and, in-turn, prevent atrophy during periods of disuse such as bedrest, immobilization and aging.

Public Health Relevance

The proposed studies have broad application to health-related research and could lead to the development of therapies aimed at preventing skeletal muscle atrophy during conditions such as bedrest, immobilization, spaceflight, aging, cachexia and dystrophy.

Agency
National Institute of Health (NIH)
Institute
National Institute of Arthritis and Musculoskeletal and Skin Diseases (NIAMS)
Type
Research Project (R01)
Project #
5R01AR057347-03
Application #
8230788
Study Section
Skeletal Muscle and Exercise Physiology Study Section (SMEP)
Program Officer
Boyce, Amanda T
Project Start
2010-04-01
Project End
2015-02-28
Budget Start
2012-03-01
Budget End
2013-02-28
Support Year
3
Fiscal Year
2012
Total Cost
$288,684
Indirect Cost
$94,284
Name
University of Wisconsin Madison
Department
Biology
Type
Schools of Veterinary Medicine
DUNS #
161202122
City
Madison
State
WI
Country
United States
Zip Code
53715
White, James P; Wrann, Christiane D; Rao, Rajesh R et al. (2014) G protein-coupled receptor 56 regulates mechanical overload-induced muscle hypertrophy. Proc Natl Acad Sci U S A 111:15756-61
You, Jae-Sung; Lincoln, Hannah C; Kim, Chan-Ran et al. (2014) The role of diacylglycerol kinase ýý and phosphatidic acid in the mechanical activation of mammalian target of rapamycin (mTOR) signaling and skeletal muscle hypertrophy. J Biol Chem 289:1551-63
Vaughan, Emily M; You, Jae-Sung; Elsie Yu, Hoi-Ying et al. (2014) Lipid domain-dependent regulation of single-cell wound repair. Mol Biol Cell 25:1867-76
Jacobs, Brittany L; Goodman, Craig A; Hornberger, Troy A (2014) The mechanical activation of mTOR signaling: an emerging role for late endosome/lysosomal targeting. J Muscle Res Cell Motil 35:11-21
Frey, John W; Jacobs, Brittany L; Goodman, Craig A et al. (2014) A role for Raptor phosphorylation in the mechanical activation of mTOR signaling. Cell Signal 26:313-22
Goodman, Craig A; Hornberger, Troy A (2013) Measuring protein synthesis with SUnSET: a valid alternative to traditional techniques? Exerc Sport Sci Rev 41:107-15
Jacobs, Brittany L; You, Jae-Sung; Frey, John W et al. (2013) Eccentric contractions increase the phosphorylation of tuberous sclerosis complex-2 (TSC2) and alter the targeting of TSC2 and the mechanistic target of rapamycin to the lysosome. J Physiol 591:4611-20
Goodman, Craig A; McNally, Rachel M; Hoffmann, F Michael et al. (2013) Smad3 induces atrogin-1, inhibits mTOR and protein synthesis, and promotes muscle atrophy in vivo. Mol Endocrinol 27:1946-57
Goodman, Craig A; Pierre, Philippe; Hornberger, Troy A (2012) Imaging of protein synthesis with puromycin. Proc Natl Acad Sci U S A 109:E989; author reply E990
Goodman, Craig A; Mayhew, David L; Hornberger, Troy A (2011) Recent progress toward understanding the molecular mechanisms that regulate skeletal muscle mass. Cell Signal 23:1896-906

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