Orai1-induced store-operated Ca(2+) entry enhances phospholipase activity and modulates canonical transient receptor potential channel 6 function in murine platelets

• Published in: Journal of thrombosis and haemostasis 2014-04, Vol.12 (4), p.528-539
• Main Authors: Chen, W, Thielmann, I, Gupta, S, Subramanian, H, Stegner, D, van Kruchten, R, Dietrich, A, Gambaryan, S, Heemskerk, J W M, Hermanns, H M, Nieswandt, B, Braun, A
• Format: Article
• Language: English
• Subjects: Animals; Blood Platelets - cytology; Calcium - metabolism; Calcium Channels - metabolism; Diglycerides - chemistry; Glycoproteins - chemistry; Liver - metabolism; Mice; Mice, Inbred C57BL; Mice, Knockout; Mice, Transgenic; ORAI1 Protein; Phospholipases - metabolism; Platelet Aggregation; Signal Transduction; Thapsigargin - metabolism; Thromboxane A2 - metabolism; Time Factors; TRPC Cation Channels - metabolism
• ISSN: 1538-7836
• DOI: 10.1111/jth.12525

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. We aimed to elucidate the role of Orai1 in diacylglycerol (DAG)-mediated ROCE. Trpc6(-/-) , Orai1(-/-) and Orai1(-/-) /Trpc6(-/-) mice were generated, and their platelets were analyzed. 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. 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.