title: Phospholipase C, Ca2+, and calmodulin signaling are required for 5-HT2A receptor-mediated transamidation of Rac1 by transglutaminase.
abstract: BACKGROUND
  Serotonin and especially serotonin 2A (5-HT(2A)) receptor signaling are important in the etiology and treatment of schizophrenia and affective disorders. We previously reported a novel 5-HT(2A) receptor effector, increased transglutaminase (TGase)-catalyzed transamidation, and activation of the small G protein Rac1 in A1A1v cells, a rat embryonic cortical cell line.
  
  
  OBJECTIVE
  In this study, we explore the signaling pathway involved in 5-HT(2A) receptor-mediated Rac1 transamidation.
  
  
  METHODS
  A1A1v cells were pretreated with pharmacological inhibitors of phospholipase C (PLC) or calmodulin (CaM), and then stimulated by the 5-HT(2A) receptor agonist, 2,5-dimethoxy-4-iodoamphetamine (DOI). Intracellular Ca(2+) concentration and TGase-modified Rac1 transamidation were monitored. The effect of manipulation of intracellular Ca(2+) by a Ca(2+) ionophore or a chelating agent on Rac1 transamidation was also evaluated.
  
  
  RESULTS
  In cells pretreated with a PLC inhibitor U73122, DOI-stimulated increases in the intracellular Ca(2+) concentration and TGase-modified Rac1 were significantly attenuated as compared to those pretreated with U73343, an inactive analog. The membrane-permeant Ca(2+) chelator, BAPTA-AM strongly reduced TGase-catalyzed Rac1 transamidation upon DOI stimulation. Conversely, the Ca(2+) ionophore ionomycin, at a concentration that induced an elevation of cytosolic Ca(2+) to a level comparable to cells treated with DOI, produced an increase in TGase-modified Rac1 without 5-HT(2A) receptor activation. Moreover, the CaM inhibitor W-7, significantly decreased Rac1 transamidation in a dose-dependent manner in DOI-treated cells.
  
  
  CONCLUSIONS
  These results indicate that 5-HT(2A) receptor-coupled PLC activation and subsequent Ca(2+) and CaM signaling are necessary for TGase-catalyzed Rac1 transamidation, and an increase in intracellular Ca(2+) is sufficient to induce Rac1 transamidation.
authors: Neuroscience Program, Loyola University, Maywood, IL, USA.
authors: Ying
authors: Y
authors: Dai
authors: Nichole L
authors: NL
authors: Dudek
authors: Qian
authors: Q
authors: Li
authors: Nancy A
authors: NA
authors: Muma
mesh_terms: D000662:Amphetamines / Q000494:pharmacology; D000818:Animals; D002118:Calcium / Q000378:metabolism*; D002147:Calmodulin / Q000378:metabolism; D002460:Cell Line; D002540:Cerebral Cortex / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D004963:Estrenes / Q000494:pharmacology; D015759:Ionomycin / Q000494:pharmacology; D011760:Pyrrolidinones / Q000494:pharmacology; D051381:Rats; D044402:Receptor, Serotonin, 5-HT2A / Q000378:metabolism*; D015398:Signal Transduction / Q000187:drug effects; D013449:Sulfonamides / Q000008:administration & dosage / Q000494:pharmacology; D011503:Transglutaminases / Q000378:metabolism*; D010738:Type C Phospholipases / Q000378:metabolism; D020830:rac1 GTP-Binding Protein / Q000187:drug effects / Q000378:metabolism*
chemical_list: D000662:Amphetamines; D002147:Calmodulin; D004963:Estrenes; D011760:Pyrrolidinones; D044402:Receptor, Serotonin, 5-HT2A; D013449:Sulfonamides; C060229:1-(6-((3-methoxyestra-1,3,5(10)-trien-17-yl)amino)hexyl)-1H-pyrrole-2,5-dione; D015759:Ionomycin; C017967:W 7; D011503:Transglutaminases; D010738:Type C Phospholipases; D020830:rac1 GTP-Binding Protein; C015952:4-iodo-2,5-dimethoxyphenylisopropylamine; D002118:Calcium