King JD et al. (2009), AMP-activated protein kinase phosphorylation of...

XB-ART-39648

Am J Physiol Cell Physiol 2009 Jul 01;2971:C94-101. doi: 10.1152/ajpcell.00677.2008.

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AMP-activated protein kinase phosphorylation of the R domain inhibits PKA stimulation of CFTR.

King JD , Fitch AC , Lee JK , McCane JE , Mak DO , Foskett JK , Hallows KR .

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The metabolic sensor AMP-activated protein kinase (AMPK) has emerged as an important link between cellular metabolic status and ion transport activity. We previously found that AMPK binds to and phosphorylates CFTR in vitro and inhibits PKA-dependent stimulation of CFTR channel gating in Calu-3 bronchial serous gland epithelial cells. To further characterize the mechanism of AMPK-dependent regulation of CFTR, whole cell patch-clamp measurements were performed with PKA activation in Calu-3 cells expressing either constitutively active or dominant-negative AMPK mutants (AMPK-CA or AMPK-DN). Baseline CFTR conductance in cells expressing AMPK-DN was substantially greater than controls, suggesting that tonic AMPK activity in these cells inhibits CFTR under basal conditions. Although baseline CFTR conductance in cells expressing AMPK-CA was comparable to that of controls, PKA stimulation of CFTR was completely blocked in AMPK-CA-expressing cells, suggesting that AMPK activation renders CFTR resistant to PKA activation in vivo. Phosphorylation studies of CFTR in human embryonic kidney-293 cells using tetracycline-inducible expression of AMPK-DN demonstrated AMPK-dependent phosphorylation of CFTR in vivo. However, AMPK activity modulation had no effect on CFTR in vivo phosphorylation in response to graded doses of PKA or PKC agonists. Thus, AMPK-dependent CFTR phosphorylation renders the channel resistant to activation by PKA and PKC without preventing phosphorylation by these kinases. We found that Ser768, a CFTR R domain residue considered to be an inhibitory PKA site, is the dominant site of AMPK phosphorylation in vitro. Ser-to-Ala mutation at this site enhanced baseline CFTR activity and rendered CFTR resistant to inhibition by AMPK, suggesting that AMPK phosphorylation at Ser768 is required for its inhibition of CFTR. In summary, our findings indicate that AMPK-dependent phosphorylation of CFTR inhibits CFTR activation by PKA, thereby tuning the PKA-responsiveness of CFTR to metabolic and other stresses in the cell.

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Species referenced: Xenopus laevis
Genes referenced: calu cftr prkaa1

References [+] :

Bhalla, AMP-activated kinase inhibits the epithelial Na+ channel through functional regulation of the ubiquitin ligase Nedd4-2. 2006, Pubmed, Xenbase

Bhalla, AMP-activated kinase inhibits the epithelial Na+ channel through functional regulation of the ubiquitin ligase Nedd4-2. 2006, Pubmed , Xenbase
Carattino, Epithelial sodium channel inhibition by AMP-activated protein kinase in oocytes and polarized renal epithelial cells. 2005, Pubmed , Xenbase
Carson, The two nucleotide-binding domains of cystic fibrosis transmembrane conductance regulator (CFTR) have distinct functions in controlling channel activity. 1995, Pubmed
Chang, Protein kinase A (PKA) still activates CFTR chloride channel after mutagenesis of all 10 PKA consensus phosphorylation sites. 1993, Pubmed
Chappe, Phosphorylation of protein kinase C sites in NBD1 and the R domain control CFTR channel activation by PKA. 2003, Pubmed
Cheng, Phosphorylation of the R domain by cAMP-dependent protein kinase regulates the CFTR chloride channel. 1991, Pubmed
Csanády, Preferential phosphorylation of R-domain Serine 768 dampens activation of CFTR channels by PKA. 2005, Pubmed , Xenbase
Hallows, Regulation of channel gating by AMP-activated protein kinase modulates cystic fibrosis transmembrane conductance regulator activity in lung submucosal cells. 2003, Pubmed , Xenbase
Hallows, Physiological modulation of CFTR activity by AMP-activated protein kinase in polarized T84 cells. 2003, Pubmed , Xenbase
Hallows, Emerging role of AMP-activated protein kinase in coupling membrane transport to cellular metabolism. 2005, Pubmed
Hallows, AMP-activated protein kinase inhibits alkaline pH- and PKA-induced apical vacuolar H+-ATPase accumulation in epididymal clear cells. 2009, Pubmed
Hallows, Inhibition of cystic fibrosis transmembrane conductance regulator by novel interaction with the metabolic sensor AMP-activated protein kinase. 2000, Pubmed , Xenbase
Hamilton, An activating mutation in the gamma1 subunit of the AMP-activated protein kinase. 2001, Pubmed
Hardie, AMP-activated protein kinase: the energy charge hypothesis revisited. 2001, Pubmed
Hardie, Management of cellular energy by the AMP-activated protein kinase system. 2003, Pubmed
Huang, PKA holoenzyme is functionally coupled to CFTR by AKAPs. 2000, Pubmed
Hurley, Regulation of AMP-activated protein kinase by multisite phosphorylation in response to agents that elevate cellular cAMP. 2006, Pubmed
Jia, Phosphorylation by protein kinase C is required for acute activation of cystic fibrosis transmembrane conductance regulator by protein kinase A. 1997, Pubmed
Kongsuphol, Mechanistic insight into control of CFTR by AMPK. 2009, Pubmed
Liedtke, Protein kinase C epsilon-dependent regulation of cystic fibrosis transmembrane regulator involves binding to a receptor for activated C kinase (RACK1) and RACK1 binding to Na+/H+ exchange regulatory factor. 2002, Pubmed
Neumann, Mammalian AMP-activated protein kinase: functional, heterotrimeric complexes by co-expression of subunits in Escherichia coli. 2003, Pubmed
Pastor-Soler, Alkaline pH- and cAMP-induced V-ATPase membrane accumulation is mediated by protein kinase A in epididymal clear cells. 2008, Pubmed
Quinton, Control of CFTR chloride conductance by ATP levels through non-hydrolytic binding. 1992, Pubmed
Riordan, Identification of the cystic fibrosis gene: cloning and characterization of complementary DNA. 1989, Pubmed
Rommens, Identification of the cystic fibrosis gene: chromosome walking and jumping. 1989, Pubmed
Treharne, The phosphorylation status of membrane-bound nucleoside diphosphate kinase in epithelia and the role of AMP. 2009, Pubmed
Vais, Dibasic phosphorylation sites in the R domain of CFTR have stimulatory and inhibitory effects on channel activation. 2004, Pubmed
Wilkinson, CFTR activation: additive effects of stimulatory and inhibitory phosphorylation sites in the R domain. 1997, Pubmed , Xenbase
Wine, Cystic fibrosis: How do CFTR mutations cause cystic fibrosis? 1995, Pubmed
Winter, Stimulation of CFTR activity by its phosphorylated R domain. 1997, Pubmed