title: Creatine kinase, an ATP-generating enzyme, is required for thrombin receptor signaling to the cytoskeleton.
abstract: Thrombin orchestrates cellular events after injury to the vascular system and extravasation of blood into surrounding tissues. The pathophysiological response to thrombin is mediated by protease-activated receptor-1 (PAR-1), a seven-transmembrane G protein-coupled receptor expressed in the nervous system that is identical to the thrombin receptor in platelets, fibroblasts, and endothelial cells. Once activated by thrombin, PAR-1 induces rapid and dramatic changes in cell morphology, notably the retraction of growth cones, axons, and dendrites in neurons and processes in astrocytes. The signal is conveyed by a series of localized ATP-dependent reactions directed to the actin cytoskeleton. How cells meet the dynamic and localized energy demands during signal transmission is unknown. Using the yeast two-hybrid system, we identified an interaction between PAR-1 cytoplasmic tail and the brain isoform of creatine kinase, a key ATP-generating enzyme that regulates ATP within subcellular compartments. The interaction was confirmed in vitro and in vivo. Reducing creatine kinase levels or its ATP-generating potential inhibited PAR-1-mediated cellular shape changes as well as a PAR-1 signaling pathway involving the activation of RhoA, a small G protein that relays signals to the cytoskeleton. Thrombin-stimulated intracellular calcium release was not affected. Our results suggest that creatine kinase is bound to PAR-1 where it may be poised to provide bursts of site-specific high-energy phosphate necessary for efficient receptor signal transduction during cytoskeletal reorganization.
authors.affiliation: Department of Microbiology and Molecular Genetics, College of Medicine University of California, Irvine, CA 92697-4025, USA.
authors.forename: V B
authors.initials: VB
authors.lastname: Mahajan
authors.forename: K S
authors.initials: KS
authors.lastname: Pai
authors.forename: A
authors.initials: A
authors.lastname: Lau
authors.forename: D D
authors.initials: DD
authors.lastname: Cunningham
mesh_terms: D000255:Adenosine Triphosphate / Q000096:biosynthesis*; D000818:Animals; D001483:Base Sequence; D002118:Calcium / Q000378:metabolism; D003402:Creatine Kinase / Q000378:metabolism*; D003599:Cytoskeleton / Q000378:metabolism*; D016376:Oligonucleotides, Antisense; D051381:Rats; D044463:Receptor, PAR-1; D018179:Receptors, Thrombin / Q000378:metabolism*; D015398:Signal Transduction; D020798:Two-Hybrid System Techniques
chemical_list: D016376:Oligonucleotides, Antisense; D044463:Receptor, PAR-1; D018179:Receptors, Thrombin; D000255:Adenosine Triphosphate; D003402:Creatine Kinase; D002118:Calcium