TRIB3 R84 Variant Is Associated With Impaired Insulin-Mediated Nitric Oxide Production in Human Endothelial Cells

Abstract

Background-In the endothelium, insulin promotes nitric oxide (NO) production, through the insulin receptor/IRS-1/PI3-Kinase/Akt/eNOS signaling pathway. An inhibitor of insulin action, TRIB3, has recently been identified which affects insulin action by binding to and inhibiting Akt phosphorylation. We have recently described a Q84R gain-of-function polymorphism of TRIB3 with the R84 variant being associated with insulin resistance and an earlier age at myocardial infarction. Methods and Results-To investigate the TRIB3 R84 variant impact on endothelial insulin action, we cultured human umbilical vein endothelial cells (HUVECs) naturally carrying different TRIB3 genotypes (QQ-, QR-, or RR-HUVECs). TRIB3 inhibitory activity on insulin-stimulated Akt phosphorylation and the amount of protein which was coimmunoprecipitable with Akt were significantly greater in QR-and RR-as compared to QQ-HUVECs. After insulin stimulation, Akt and eNOS activation as well as NO production were markedly decreased in QR-and RR-as compared to QQ-HUVECs. TRIB3 molecular modeling analysis provided insights into the structural changes related to the polymorphisms potentially determining differences in protein-protein interaction with Akt. Conclusions-Our data demonstrate that the TRIB3 R84 variant impairs insulin signaling and NO production in human endothelial cells. This finding provides a plausible biological background for the deleterious role of TRIB3 R84 on genetic susceptibility to coronary artery disease.

TRIB3 R84 variant affects glucose homeostasis by altering the interplay between insulin sensitivity and secretion

“…In functional studies on transfected cells, an inhibitory role of R84 variant on insulin-stimulated Akt phosphorylation has been observed in vitro in both cells from peripheral insulin target tissues [6,7] and insulin-secreting beta cells (C. Wee Liew, J. Bochenski, J. Hu, A.S. Krowleski and R.N. Kulkarni, unpublished data), making plausible a direct role of this variant on both in vivo insulin sensitivity and insulin secretion.…”

“…Thus, an altered interplay between insulin sensitivity and secretion is instrumental for the development of abnormal glucose homeostasis [3,4]. The mammalian tribbles homologue 3 (TRIB3) is an insulin signalling inhibitor which binds Akt [5] and inhibits [5][6][7] insulin-stimulated Akt phosphorylation and subsequent insulin action.…”

“…rs2295490, where arginine replaces glutamine at position 84), resulting in a gain-of-function variant [6,7]. In vitro studies have revealed that the R84 variant is more effective than the Q84 variant in engaging Akt and reducing insulin-mediated Ser473 Akt phosphorylation in human insulin target tissues [6,7] as well as in insulin-secreting beta cells (C. Wee Liew, J. Bochenski, J. Hu, A.S. Krowleski and R.N. Kulkarni, unpublished data), thus suggesting a deleterious role of this variant on both insulin action and secretion.…”

Tribbles-1 regulates hepatic lipogenesis through posttranscriptional regulation of C/EBPα

“…This function is critical for tribbles-mediated leukemogenesis (16,17) and also coordinates TRIB1 regulation of macrophage polarization and differentiation (18). Earlier work has identified partially defined roles for other tribbles proteins in human metabolism, particularly TRIB3, which is involved in regulation of insulin signaling through interactions with AKT/Protein Kinase B (19,20) and as a binding partner for ATF4 in pancreatic β cells (21). Adipose TRIB3 can also facilitate the turnover of the protein acetylcoenzyme A carboxylase (ACACA), the rate-limiting enzyme in fatty acid synthesis (22).…”

Aberrant hepatic TRIB3 gene expression in insulin-resistant obese humans

“…In humans, associations of a gain-of-function TRIB3 Glu84Arg missense polymorphism with phenotypes related to insulin resistance have been described [7]. Furthermore, overexpression of the variant Arg84 allele enhanced inhibition of insulin-mediated AKT1 phosphorylation in HepG2 cells [7] and was associated with impaired insulin signalling and nitric oxide production in human endothelial cells [8]. Because of a possible role of TRIB3 in insulin resistance in humans, we measured hepatic mRNA expression of TRIB3 in obese patients with varying degrees of insulin resistance and determined associations with biochemical variables and hepatic transcript levels of genes involved in transcriptional regulation and glucose homeostasis.…”