Regulation of a truncated isoform of AMP-activated protein kinase α (AMPKα) in response to hypoxia in the muscle of Pacific oyster Crassostrea gigas

Abstract

AMP-activated protein kinase α (AMPKα) is a key regulator of energy balance in many model species during hypoxia. In a marine bivalve, the Pacific oyster Crassostrea gigas, we analyzed the protein content of adductor muscle in response to hypoxia during 6 h. In both smooth and striated muscles, the amount of full-length AMP-activated protein kinase α (AMPKα) remained unchanged during hypoxia. However, hypoxia induced a rapid and muscle-specific response concerning truncated isoforms of AMPKα. In the smooth muscle, a truncated isoform of AMPKα was increased from 1 to 6 h of hypoxia, and was linked with accumulation of AKT kinase, a key enzyme of the insulin signaling pathway which controls intracellular glucose metabolism. In this muscle, aerobic metabolism was maintained over the 6 h of hypoxia, as mitochondrial citrate synthase activity remained constant. In contrast, in striated muscle, hypoxia did not induce any significant modification of neither truncated AMPKα nor AKT protein content, and citrate synthase activity was altered after 6 h of hypoxia. Together, our results demonstrate that hypoxia response is specific to muscle type in Pacific oyster, and that truncated AMPKα and AKT proteins might be involved in maintaining aerobic metabolism in smooth muscle. Such regulation might occur in vivo during tidal intervals that cause up to 6 h of hypoxia.

Seascape genomics reveals adaptive divergence in a connected and commercially important mollusc, the greenlip abalone (Haliotis laevigata), along a longitudinal environmental gradient

“…Additionally, four candidate loci were mapped to genes involved in the glycolysis process (production of ATP in anaerobic or aerobic conditions), whose efficiency plays a fundamental role in the response of mammals to low levels of oxygen and high energy demands (Robin, Murphy, & Theodore, ). Moreover, four enriched gene functions (nitrogen compound metabolic process, CoA‐ligase activity, citrate metabolic process, dicarboxylic acid metabolic process, see Appendix : Table S8) are involved in glycolytic and metabolic pathways which have important roles in adaptation to hypoxia and anoxia in different animals, including molluscs (Guévélou et al., ; Hermes‐Lima et al., ; Ivanina, Nesmelova, Leamy, Sokolov, & Sokolova, ).…”

Sex-Specific Regulation of AMP-Activated Protein Kinase (AMPK) in the Pacific Oyster Crassostrea gigas1

“…In the Pacific oyster, we characterized the two noncatalytic subunits of the AMPK heterotrimer, AMPK beta and AMPK gamma, to make a complete characterization of the AMPK system with the three subunits, including the AMPK alpha previously characterized [64]. When examining the phylogenetic tree generated, the C. gigas AMPK beta subunit seems to be slightly closer to the AMPK beta1 group sequence.…”

“…In C. gigas, both the entire AMPK alpha (62 kDa) and the truncated AMPK alpha (45 kDa) were already detected in some tissues [64]. The amount of truncated AMPK alpha was upregulated under hypoxia in the smooth muscle and was supposed to play a role in maintaining aerobic metabolism in this tissue [56].…”

“…Immunodetections were performed with a rabbit monoclonal antibody anti-AMPK alpha (dilution 1:1000; CST #2603; Ozyme [the Cell Signaling Technology distributor in France]), a mouse monoclonal antibody anti-AMPK alpha (dilution 1:1000; CST #2793; Ozyme), and a rabbit monoclonal antibody anti-phospho-threonine 172 AMPK alpha (dilution 1:1000; CST #2535; Ozyme); heterologous antibodies CST #2603 and CST #2793 directed against AMPK alpha had already been validated for the recognition of the truncated AMPK alpha isoforms or the entire AMPK alpha, respectively [64]. Moreoever, Cell Signaling Technology technical services blasted and confirmed that the antigenic region of the commercially available anti-phospho-threonine 172 AMPK alpha antibody was present at 100% in the C. gigas protein (Cell Signaling Technology, personal communication).…”

Active and passive biomonitoring suggest metabolic adaptation in blue mussels ( Mytilus spp.) chronically exposed to a moderate contamination in Brest harbor (France)

“…Then, total protein extractions were performed on 200 mg of pool powder mixed in 1 ml of lysis buffer [pH 7.4; NaCl 150 mM, Tris HCl 10 mM, EDTA 1 mM, EGTA 1 mM, phosphatase inhibitor cocktail II (Sigma-Aldrich) 1%, Triton X-100 1%, CA-Igepal ® (Sigma-Aldrich) 0.5%, sodium pyrophosphatase decahydrate 5 mM and 1 tablets/25 ml of complete EDTA free protease inhibitor cocktail (Roche)] (Guévélou et al, 2013) using a Precellys ® 24 grinder coupled to a Cryolys ® cooling system (Bertin technologies). Samples were subsequently centrifuged at 10,000 × g for 45 min at 4 • C prior to supernatant collection, aliquoting and storage at −80 • C.…”