Regulation of thromboxane receptor signaling at multiple levels by oxidative stress-induced stabilization, relocation and enhanced responsiveness

• Published in: PloS one Vol. 5; no. 9; p. e12798
• Main Authors: Ball, Stephen K, Field, Mark C, Tippins, John R
• Format: Journal Article
• Language: English
• Published: United States Public Library of Science 15.09.2010; Public Library of Science (PLoS)
• Subjects: Amino acids; Apoptosis; Arachidonic acid; Biotinylation; Brefeldin A; Calcium; Calcium (intracellular); Calcium content; Cardiovascular disease; Cardiovascular Disorders/Hypertension; Cardiovascular Disorders/Vascular Biology; Cell Biology; Cell Biology/Membranes and Sorting; Cell Line, Tumor; Cell surface; Cellular signal transduction; Confocal; Confocal microscopy; Coronary vessels; Endoplasmic reticulum; Exposure; Golgi apparatus; HEK293 Cells; Humans; Hydrogen; Hydrogen peroxide; Hydrogen Peroxide - metabolism; Internalization; Kinases; Metabolites; Microscopy; Molecular biology; Oxidative Stress; Pathology; Penicillin; Physiological aspects; Plasma; Polyclonal antibodies; Potassium; Potentiation; Protein Stability; Protein Transport; Proteins; Receptors; Receptors, Thromboxane - chemistry; Receptors, Thromboxane - genetics; Receptors, Thromboxane - metabolism; Relocation; Signal Transduction; Signaling; Smooth muscle; Stabilization; Thromboxane A2; Transduction; Translocation; United Kingdom > UK
• ISSN: 1932-6203; 1932-6203
• DOI: 10.1371/journal.pone.0012798

Thromboxane A(2) (TxA(2)) is a major, unstable arachidonic acid metabolite, and plays a key role in normal physiology and control of vascular tone. The human thromboxane receptor (TPβ), expressed in COS-7 cells, is located predominantly in the endoplasmic reticulum (ER). Brief hydrogen peroxide exposure increases the efficiency of translocation of TPβ from the ER into the Golgi complex, inducing maturation and stabilization of TPβ. However, the ultimate fate of this post-ER TPβ pool is not known, nor is its capacity to initiate signal transduction. Here we specifically assessed if functional TPβ was transported to the plasma membrane following H(2)O(2) exposure. We demonstrate, by biotinylation and confocal microscopy, that exposure to H(2)O(2) results in rapid delivery of a cohort of TPβ to the cell surface, which is stable for at least eight hours. Surface delivery is brefeldin A-sensitive, indicating that translocation of this receptor cohort is from internal pools and via the Golgi complex. H(2)O(2) treatment results in potentiation of the increase to intracellular calcium concentrations in response to TPβ agonists U46619 and 8-iso PGF(2α) and also in the loss of ligand-dependent receptor internalization. Further there is increased responsiveness to a second application of the agonist. Finally we demonstrate that the effect of H(2)O(2) on stimulating surface delivery is shared with the FP prostanoid receptor but not the EP3 or EP4 receptors. In summary, brief exposure to H(2)O(2) results in an immediate and sustained increase in the surface pool of thromboxane receptor that is capable of mediating a persistent hyper-responsiveness of the cell and suggests a highly sophisticated mechanism for rapidly regulating thromboxane signaling.