Argininosuccinate synthetase regulates hepatic AMPK linking protein catabolism and ureagenesis to hepatic lipid metabolism

Abstract

A key sensor of cellular energy status, AMP-activated protein kinase (AMPK), interacts allosterically with AMP to maintain an active state. When active, AMPK triggers a metabolic switch, decreasing the activity of anabolic pathways and enhancing catabolic processes such as lipid oxidation to restore the energy balance. Unlike oxidative tissues, in which AMP is generated from adenylate kinase during states of high energy demand, the ornithine cycle enzyme argininosuccinate synthetase (ASS) is a principle site of AMP generation in the liver. Here we show that ASS regulates hepatic AMPK, revealing a central role for ureagenesis flux in the regulation of metabolism via AMPK. Treatment of primary rat hepatocytes with amino acids increased gluconeogenesis and ureagenesis and, despite nutrient excess, induced both AMPK and acetyl-CoA carboxylase (ACC) phosphorylation. Antisense oligonucleotide knockdown of hepatic ASS1 expression in vivo decreased liver AMPK activation, phosphorylation of ACC, and plasma β-hydroxybutyrate concentrations. Taken together these studies demonstrate that increased amino acid flux can activate AMPK through increased AMP generated by ASS, thus providing a novel link between protein catabolism, ureagenesis, and hepatic lipid metabolism.argininosuccinate synthetase | AMPK | ureagenesis | amino acids | lipid metabolism M aintaining appropriate nitrogen balance is crucial to the fitness of an organism. The decision to deaminate amino acids to use their carbon skeleton for oxidative and/or synthetic purposes is tightly regulated. Excessive amino acid catabolism not only generates toxic nitrogenous waste but also could jeopardize the ability to generate or maintain protein levels.The liver is unique in its capacity to detoxify nitrogenous waste through the energy-requiring synthesis of urea. Accounting for nearly a third of hepatic metabolism, ureagenesis consumes three ATPs per urea produced and has an overall flux comparable to that of gluconeogenesis (1, 2). Importantly, the ligation of citrulline to aspartate via argininosuccinate synthetase (ASS) hydrolyzes one ATP into AMP per turn of the ornithine cycle, making ASS a predominant hepatocellular contributor of AMP. Hence, a metabolic signal generated through the AMP-generating ornithine cycle could be intrinsic to preserving nitrogen balance.In oxidative tissues such as muscle, AMPK helps balance energy homeostasis by detecting fluctuations in the [AMP]:[ATP] ratio. Phosphorylation of AMPK by upstream kinases such as liver kinase B1 enhance AMPK activity, and AMP binding to an allosteric site stabilizes a conformation that supports phosphorylation and protects AMPK from dephosphorylation (3). The canonical view of AMPK activation is that, following global shifts in energy balance (e.g., excessive energy demand during exercise), adenylate kinase (AK) increases AMP synthesis to salvage ATP from ADP (4). As a consequence, the AMPK-mediated phosphorylation of sterol regulatory element-binding protein 1 (SREBP-1), acetyl-CoA carboxylase 1 (ACC1), a...

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