Role of AMP-activated protein kinase (AMPK) in the control of skeletal muscle protein synthesis and epithelial cell architecture
(2010)
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- Rider, Mark
- Abstract
- (en) The AMP-activated protein kinase (AMPK) system has been principally considered as a regulator of cellular and systemic energy balance. However, it is now recognized that its role extends beyond energy sensing to the control of cell division, cell polarity and cell migration. In this thesis, a study of the AMPK system on the control of one energy consuming process (protein synthesis) is presented as well as a study of one non-metabolic function of AMPK, namely control of the actin cytoskeleton. The mechanisms of control of skeletal muscle protein synthesis by contraction/AMPK and insulin were first investigated. In incubated rat epitrochlearis skeletal muscles, insulin stimulated protein synthesis. Contraction induced by electrical stimulation on the other hand led to AMPK activation and inhibition of the basal and insulin-stimulated rates of protein synthesis. Electrical stimulation had no effect on basal mammalian target of rapamycin complex 1 (mTORC1) signaling, but did antagonize mTORC1 signaling after stimulation of the pathway by insulin. Phosphorylation of eukaryotic elongation factor-2 (eEF2), a known target of AMPK, increased rapidly upon electrical stimulation, but could not be related to AMPK activation, which occurred more slowly to reach a 3-fold activation after 30 min. Therefore, in electrically stimulated skeletal muscle, the contraction-induced inhibition of protein synthesis could not be attributed to inhibition of mTORC1 signaling but could be due to increased eEF2 phosphorylation, independent of AMPK activation. The non-metabolic role of AMPK in controlling reorganization of the actin cytoskeleton was studied in epithelial Madin-Darby canine kidney (MDCK) cells. The effects of hyperosmolarity and treatment with pharmacological AMPK activators (AICAR and A-769662) on cytoskeletal organization and signaling were investigated. AMPK activation by A-769662 induced peripheral F-actin accumulation, disappearance of focal adhesions and disassembly of stress fibers. In parallel, RhoA was activated, associated with an increase in phosphorylation of the Rho-kinase (ROCK) downstream targets, myosin regulatory light chain (MLC) and cofilin. Accordingly, the A769662-induced increase in MLC and cofilin phosphorylation was prevented by H1152, a ROCK inhibitor. In MDCK cells hyperosmotic shock activated AMPK via the calmodulin-dependent protein kinase kinase-b pathway. Cell shrinkage induced similar changes in cell architecture and increased MLC phosphorylation to those observed with pharmacological AMPK activators. However, AMPK activation due to ATP-depletion by 2-deoxyglucose treatment did not induce cytoskeletal reorganization or increase MLC phosphorylation in these cells. We propose that AMPK participates via the RhoA/ROCK pathway, in the reorganization of the actin cytoskeleton induced by osmotic stress.
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Citations
Miranda Miranda, L. (2010). Role of AMP-activated protein kinase (AMPK) in the control of skeletal muscle protein synthesis and epithelial cell architecture. https://hdl.handle.net/2078.5/130216