Thioredoxin-Interacting Protein (TXNIP) is a Biomechanical Regulator of Src Activity: Key Role in Endothelial Cell Stress Fiber Formation.
ABSTRACT Fluid shear stress (FSS) differentially regulates endothelial cell (EC) stress fiber formation with decreased stress fibers in areas of disturbed-flow (d-flow) compared to steady-flow (s-flow) areas. Importantly, stress fibers are critical for several EC functions including cell shape, mechano-signal transduction, and EC cell-cell junction integrity. A key mediator of s-flow induced stress fiber formation is Src, which regulates downstream signaling mediators such as phosphorylation of cortactin, activity of focal adhesion kinase and small GTPases.
Previously we showed that thioredoxin-interacting protein (TXNIP, also VDUP1 and TBP-2) was regulated by FSS; TXNIP expression was increased in d-flow compared to s-flow areas. While TXNIP was originally characterized for its role in redox and metabolic cellular functions, recent reports show important scaffold functions related to its α-arrestin structure. Based on these findings, we hypothesized that TXNIP acts as a biomechanical sensor that regulates Src kinase activity and stress fiber formation.
Using en face immunohistochemistry of the aorta and cultured EC, we show inverse relationship between TXNIP expression and Src activity. Specifically, s-flow increased Src activity and stress fiber formation, while it decreased TXNIP expression. In contrast, d-flow had opposite effects. We studied the role of TXNIP in regulating SHP2 plasma membrane localization and VE-cadherin binding, because SHP2 indirectly regulates dephosphorylation of Src tyrosine 527 that inhibits Src activity. Using immunohistochemistry and immunoprecipitation we found that TXNIP prevented SHP2-VE-cadherin interaction.
In summary, these data characterize a FSS mediated mechanism for stress fiber formation that involves a TXNIP-dependent VE-cadherin-SHP2-Src pathway.
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ABSTRACT: The thioredoxin system, which consists of thioredoxin (Trx), nicotinamide adenine dinucleotide phosphate (NADPH) and thioredoxin reductase (TrxR), has emerged as a major anti-oxidant involved in the maintenance of cellular physiology and survival. Dysregulation in this system has been associated with metabolic, cardiovascular, and malignant disorders. Thioredoxin-interacting protein (TXNIP), also known as vitamin D-upregulated protein or thioredoxin-binding-protein-2, functions as a physiological inhibitor of Trx, and pathological suppression of Trx by TXNIP has been demonstrated in diabetes and cardiovascular diseases. Furthermore, TXNIP effects are partially Trx-independent; these include direct activation of inflammation and inhibition of glucose uptake. Many of the effects of TXNIP are initiated by its dissociation from intra-nuclear binding with Trx or other SH-containing proteins: these effects include its migration to cytoplasm, modulating stress responses in mitochondria and endoplasmic reticulum, and also potentially activating apoptotic pathways. TXNIP also interacts with the nitric oxide (NO) signaling system, with apparent suppression of NO effect. TXNIP production is modulated by redox stress, glucose levels, hypoxia and several inflammatory activators. In recent studies, it has been shown that therapeutic agents including insulin, metformin, angiotensin converting enzyme inhibitors and calcium channel blockers reduce TXNIP expression, although it is uncertain to what extent TXNIP suppression contributes to their clinical efficacy. This review addresses the role of TXNIP in health and in cardiovascular and metabolic disorders. Finally, the potential advantages (and disadvantages) of pharmacological suppression of TXNIP in cardiovascular disease and diabetes are summarized.Cardiovascular Drugs and Therapy 08/2014; 28(4). DOI:10.1007/s10557-014-6538-5 · 3.19 Impact Factor
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ABSTRACT: Mitogen-activated protein kinases (MAPKs) are key signal transducers involved in various cellular events such as growth, proliferation, and differentiation. Previous studies have reported that H2O2 leads to phosphorylation of extracellular signal-regulated kinase (ERK), one of the MAPKs in endothelial cells. The current study shows that H2O2 suppressed ERK1/2 activation and phosphorylation at specific concentrations and times in human umbilical vein endothelial cells but not in immortalized mouse aortic endothelial cells or human astrocytoma cell line CRT-MG. Phosphorylation of other MAPK family members (i.e., p38 and JNK) was not suppressed by H2O2. The decrease in ERK1/2 phosphorylation induced by H2O2 was inversely correlated with the level of phosphorylation of Src tyrosine 530. Using siRNA, it was found that H2O2-induced suppression of ERK1/2 was dependent on Csk. Physiological laminar flow abrogated, but oscillatory flow did not affect, the H2O2-induced suppression of ERK1/2 phosphorylation. In conclusion, H2O2-induced Csk translocation to the plasma membrane leads to phosphorylation of Src at the tyrosine 530 residue resulting in a reduction of ERK1/2 phosphorylation. Physiological laminar flow abrogates this effect of H2O2 by inducing phosphorylation of Src tyrosine 419. These findings broaden our understanding of signal transduction mechanisms in the endothelial cells against oxidative stress.Scientific Reports 08/2015; 5:12725. DOI:10.1038/srep12725 · 5.58 Impact Factor