Extracellular Nucleotides Inhibit Insulin Receptor Signaling, Stimulate Autophagy and Control Lipoprotein Secretion

Abstract

Hyperglycemia is associated with abnormal plasma lipoprotein metabolism and with an elevation in circulating nucleotide levels. We evaluated how extracellular nucleotides may act to perturb hepatic lipoprotein secretion. Adenosine diphosphate (ADP) (>10 µM) acts like a proteasomal inhibitor to stimulate apoB100 secretion and inhibit apoA-I secretion from human liver cells at 4 h and 24 h. ADP blocks apoA-I secretion by stimulating autophagy. The nucleotide increases cellular levels of the autophagosome marker, LC3-II, and increases co-localization of LC3 with apoA-I in punctate autophagosomes. ADP affects autophagy and apoA-I secretion through P2Y13. Overexpression of P2Y13 increases cellular LC3-II levels by ∼50% and blocks induction of apoA-I secretion. Conversely, a siRNA-induced reduction in P2Y13 protein expression of 50% causes a similar reduction in cellular LC3-II levels and a 3-fold stimulation in apoA-I secretion. P2Y13 gene silencing blocks the effects of ADP on autophagy and apoA-I secretion. A reduction in P2Y13 expression suppresses ERK1/2 phosphorylation, increases the phosphorylation of IR-β and protein kinase B (Akt) >3-fold, and blocks the inhibition of Akt phosphorylation by TNFα and ADP. Conversely, increasing P2Y13 expression significantly inhibits insulin-induced phosphorylation of insulin receptor (IR-β) and Akt, similar to that observed after treatment with ADP. Nucleotides therefore act through P2Y13, ERK1/2 and insulin receptor signaling to stimulate autophagy and affect hepatic lipoprotein secretion.

P2X receptors regulate adenosine diphosphate release from hepatic cells

“…We have previously shown that extracellular ADP blocks insulin receptor signaling and thereby stimulates cellular autophagy and perturbs protein secretion [1]. This work may therefore illustrate how nutrient deprivation initiates a cellular response to starvation through an autocrine and paracrine control of purinergic signaling.…”

“…The extracellular nucleotide milieu thereby acts through specific P2X and P2Y receptors to promote a purinergic signaling response. Acute and short-lived nucleotide signaling is required for normal cellular function, while chronic purinergic signaling appears to have negative effects on insulin receptor (IR-β) signaling [1,33] and cellular inflammatory pathways [7,8].…”

“…We have shown that extracellular nucleotides play an important regulatory role in both insulin receptor signaling and cellular autophagy and thereby regulate protein secretion from human hepatic carcinoma cells [1]. The factors that regulate nucleotide release and metabolism by hepatic cells were therefore evaluated.…”

“…Our research has shown that adenosine diphosphate (ADP) signaling through specific G-protein-coupled purinergic receptors (P2Y) perturbs both lipid and protein metabolism in the liver [1,2]. Extracellular nucleotides are known to activate nuclear factor kappa B [3,4] and trigger the release of proinflammatory cytokines [5,6].…”

The PRKAA1/AMPKα1 pathway triggers autophagy during CSF1-induced human monocyte differentiation and is a potential target in CMML

“…17,[33][34][35] Moreover, it has been previously reported that P2RY13 activation by ADP induces autophagy in hepatoma cell lines. 36 However to the best of our knowledge, this is the first description of purinergic receptor-mediated differentiation being dependent on autophagy induction.…”