title: Orai1-induced store-operated Ca(2+) entry enhances phospholipase activity and modulates canonical transient receptor potential channel 6 function in murine platelets.
abstract: BACKGROUND
  Orai1, the major store-operated Ca(2+) entry (SOCE) channel in platelets, is not only critical for enhancing diverse signaling pathways, but may also regulate receptor-operated Ca(2+) entry (ROCE). Dynamic coupling of the Orai1 signalosome to canonical transient receptor potential channels (TRPCs) has been suggested as an essential step in the activation of SOCE and ROCE. However, the functional significance of the biochemical interaction between Orai and TRPC isoforms remains controversial.
  
  
  OBJECTIVE
  We aimed to elucidate the role of Orai1 in diacylglycerol (DAG)-mediated ROCE.
  
  
  METHODS
  Trpc6(-/-) , Orai1(-/-) and Orai1(-/-) /Trpc6(-/-) mice were generated, and their platelets were analyzed.
  
  
  RESULTS
  Thapsigargin (TG)-induced SOCE was further reduced in Orai1(-/-) /Trpc6(-/-) platelets as compared with Orai1(-/-) platelets, thus revealing that TG-induced signaling pathways can activate TRPC6. Thapsigargin-induced SOCE leads to enhanced phospholipase C and D activity in wild-type platelets. The activity of both enzymes was significantly reduced in Orai1(-/-) platelets upon TG stimulation, whereas receptor-induced phospholipase activity was not affected. Furthermore, TG-induced and glycoprotein VI-mediated thromboxane A2 release was strongly dependent on Orai1-mediated SOCE.
  
  
  CONCLUSIONS
  The regulation of TRPC6 activity can occur independently of the physical interaction with Orai1. TRPC6 operates in crosstalk with Orai1 through Orai1-induced DAG production via phospholipase activation. Orai1-induced DAG production and thromboxane release amplify the second phase of Ca(2+) signaling in platelets.
authors: University Hospital and Rudolf Virchow Center for Experimental Biomedicine, University of Würzburg, Würzburg, Germany.
authors: W
authors: W
authors: Chen
authors: I
authors: I
authors: Thielmann
authors: S
authors: S
authors: Gupta
authors: H
authors: H
authors: Subramanian
authors: D
authors: D
authors: Stegner
authors: R
authors: R
authors: van Kruchten
authors: A
authors: A
authors: Dietrich
authors: S
authors: S
authors: Gambaryan
authors: J W M
authors: JW
authors: Heemskerk
authors: H M
authors: HM
authors: Hermanns
authors: B
authors: B
authors: Nieswandt
authors: A
authors: A
authors: Braun
mesh_terms: D000818:Animals; D001792:Blood Platelets / Q000166:cytology; D002118:Calcium / Q000378:metabolism; D015220:Calcium Channels / Q000378:metabolism*; D004075:Diglycerides / Q000737:chemistry; D006023:Glycoproteins / Q000737:chemistry; D008099:Liver / Q000378:metabolism; D051379:Mice; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D008822:Mice, Transgenic; D000071740:ORAI1 Protein; D010740:Phospholipases / Q000378:metabolism*; D010974:Platelet Aggregation; D015398:Signal Transduction; D050052:TRPC Cation Channels / Q000378:metabolism*; D000074287:TRPC6 Cation Channel; D019284:Thapsigargin / Q000378:metabolism; D013928:Thromboxane A2 / Q000378:metabolism; D013997:Time Factors
chemical_list: C011439:1,2-diacylglycerol; D015220:Calcium Channels; D004075:Diglycerides; D006023:Glycoproteins; D000071740:ORAI1 Protein; C520430:Orai1 protein, mouse; D050052:TRPC Cation Channels; D000074287:TRPC6 Cation Channel; C492146:Trpc6 protein, mouse; D013928:Thromboxane A2; D019284:Thapsigargin; D010740:Phospholipases; D002118:Calcium
keywords: blood platelets; calcium channels; calcium signaling; phospholipases; platelet activation