Angiotensin II inhibits insulin-stimulated phosphorylation of eukaryotic initiation factor 4E-binding protein-1 in proximal tubular epithelial cells

Abstract

Interaction between angiotensin II, which binds a G-protein-coupled receptor, and insulin, a ligand for receptor tyrosine kinase, was examined in renal proximal tubular epithelial cells. Augmented protein translation by insulin involves activation of eukaryotic initiation factor 4E (eIF4E) which follows the release of the factor from a heterodimeric complex by phosphorylation of its binding protein, 4E-BP1. Angiotensin II (1 nM) or insulin (1 nM) individually stimulated 4E-BP1 phosphorylation. However, pre-incubation with angiotensin II abrogated insulin-induced phosphorylation of 4E-BP1, resulting in persistent binding to eIF4E. Although angiotensin II and insulin individually activated phosphoinositide 3-kinase and extracellular signal-regulated kinase (ERK)-1/-2-type mitogen-activated protein (MAP) kinase, pre-incubation with angiotensin II abolished insulin-induced stimulation of these kinases, suggesting more proximal events in insulin signalling may be intercepted. Pretreatment with angiotensin II markedly inhibited insulin-stimulated tyrosine phosphorylation of insulin-receptor beta-chain and insulin-receptor substrate 1. Losartan prevented angiotensin II inhibition of insulin-induced ERK-1/-2-type MAP kinase activation and 4E-BP1 phosphorylation, suggesting mediation of the effect of angiotensin II by its type 1 receptor. Insulin-stimulated de novo protein synthesis was also abolished by pre-incubation with angiotensin II. These data show that angiotensin II inhibits 4E-BP1 phosphorylation and stimulation of protein synthesis induced by insulin by interfering with proximal events in insulin signalling. Our data provide a mechanistic basis for insulin insensitivity induced by angiotensin II.

Translational regulation of vascular endothelial growth factor expression in renal epithelial cells by angiotensin II

“…In agreement with the above schema, ANG II stimulated important events in capdependent translation including phosphorylation of 4E-BP1 and eIF4E. Our studies in MCT cells have shown that phosphorylation of 4E-BP1 by ANG II is associated with dissolution of the eIF4E-4E-BP1 complex (43). The inability of ANG II to induce VEGF expression in cells expressing 4E-BP1 phosphorylation mutant further confirmed cap-dependent translation as the underlying mechanism.…”

“…5C, P Ͻ 0.01). We previously showed that ANG II promotes dissociation between eIF4E and 4E-BP1 under these experimental conditions in MCT cells (43). These data demonstrate that ANG II regulates events of regulatory significance in the initiation phase of translation.…”

“…To this end, we examined the role of phosphorylation of Thr 37 and Thr 46 residues on 4E-BP1 by employing MCT cells stably expressing a Thr 37,46 3 Ala 37,46 mutant (mut. 4E-BP1) (42,43). In vitro translation of total RNA isolated from heavy ribosomal fraction demonstrated increase in synthesis of the 23-and 30-kDa proteins in MCT overexpressing the empty vector, but not mut.…”

“…SV40-immortalized murine proximal tubular epithelial cells (MCT cells; kindly provided by Dr. E. Neilson, Vanderbilt University, Nashville, TN) were grown in DMEM containing 7% FBS, 5 mM glucose, and no insulin, as recently described (43,44). Confluent monolayers of cells were serum deprived in DMEM for 18 h before treatment.…”

A role for AMP-activated protein kinase in diabetes-induced renal hypertrophy

“…Protein synthesis measurement was as previously described (49). Serum-starved cells were labeled with 10 Ci/ml of [ 35 S]methionine for the terminal 2 h of incubation.…”