Ca2+/calmodulin-dependent protein kinase kinase-β acts upstream of AMP-activated protein kinase in mammalian cells
ABSTRACT AMP-activated protein kinase (AMPK) is the downstream component of a kinase cascade that plays a pivotal role in energy homeostasis. Activation of AMPK requires phosphorylation of threonine 172 (T172) within the T loop region of the catalytic alpha subunit. Recently, LKB1 was shown to activate AMPK. Here we show that AMPK is also activated by Ca(2+)/calmodulin-dependent protein kinase kinase (CaMKK). Overexpression of CaMKKbeta in mammalian cells increases AMPK activity, whereas pharmacological inhibition of CaMKK, or downregulation of CaMKKbeta using RNA interference, almost completely abolishes AMPK activation. CaMKKbeta isolated from rat brain or expressed in E. coli phosphorylates and activates AMPK in vitro. In yeast, CaMKKbeta expression rescues a mutant strain lacking the three kinases upstream of Snf1, the yeast homolog of AMPK. These results demonstrate that AMPK is regulated by at least two upstream kinases and suggest that AMPK may play a role in Ca(2+)-mediated signal transduction pathways.
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ABSTRACT: Myocardial glucose and long-chain fatty acid uptake are regulated by specific membrane transport proteins, i.e., GLUT4 and CD36, respectively. Upon hormonal (insulin) or mechanical stimuli (muscle contraction) GLUT4 and CD36 move from endosomal stores to the plasma membrane to facilitate substrate uptake. Contraction-mediated substrate uptake is known to require AMP-dependent protein kinase (AMPK) activation.11/2015; 10(3):127-127. DOI:10.1007/s12467-012-0075-2
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ABSTRACT: AMP-activated protein kinase (AMPK), whose activity is a critical determinant of cell health, serves a fundamental role in integrating extracellular and intracellular nutrient information into signals that regulate various metabolic processes. Despite the importance of AMPK, its specific roles within the different intracellular spaces remain unresolved, largely due to the lack of real-time, organelle-specific AMPK activity probes. Here, we present a series of molecular tools that allows for the measurement of AMPK activity at the different subcellular localizations and that allows for the rapid induction of AMPK inhibition. We discovered that AMPKα1, not AMPKα2, was the subunit that preferentially conferred spatial specificity to AMPK, and that inhibition of AMPK activity at the mitochondria was sufficient for triggering cytosolic ATP increase. These findings suggest that genetically encoded molecular probes represent a powerful approach for revealing the basic principles of the spatiotemporal nature of AMPK regulation. Copyright © 2015 The Authors. Published by Elsevier Inc. All rights reserved.Cell Reports 04/2015; 12(4). DOI:10.1016/j.celrep.2015.03.057 · 7.21 Impact Factor
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ABSTRACT: The AMP-activated protein kinase, a key regulator of energy homeostasis, has a critical role in metabolic disorders and cancers. AMPK is mainly regulated by cellular AMP and phosphorylation by upstream kinases. Here, we show that PIKE-A binds to AMPK and blocks its tumor suppressive actions, which are mediated by tyrosine kinase Fyn. PIKE-A directly interacts with AMPK catalytic alpha subunit and impairs T172 phosphorylation, leading to repression of its kinase activity on the downstream targets. Mutation of Fyn phosphorylation sites on PIKE-A, depletion of Fyn, or pharmacological inhibition of Fyn blunts the association between PIKE-A and AMPK, resulting in loss of its inhibitory effect on AMPK. Cell proliferation and oncogenic assays demonstrate that PIKE-A antagonizes tumor suppressive actions of AMPK. In human glioblastoma samples, PIKE-A expression inversely correlates with the p-AMPK levels, supporting that PIKE-A negatively regulates AMPK activity in cancers. Thus, our findings provide additional layer of molecular regulation of the AMPK signaling pathway in cancer progression.Cell Death and Differentiation advance online publication, 22 May 2015; doi:10.1038/cdd.2015.66.Cell death and differentiation 05/2015; DOI:10.1038/cdd.2015.66 · 8.39 Impact Factor