Phosphatidylinositol 3,4,5-trisphosphate regulates Ca2+ entry via Btk in platelets and megakaryocytes without increasing phospholipase C activity

• Published in: The EMBO journal 2000-06, Vol.19 (12), p.2793-2802
• Main Authors: Pasquet, Jean-Max, Quek, Lynn, Stevens, Christiaan, Bobe, Régis, Huber, Michael, Duronio, Vincent, Krystal, Gerald, Watson, Steve P.
• Format: Article
• Language: English
• Subjects: 5-trisphosphate; Btk; phosphatidylinositol 3; phosphatidylinositol 3,4,5‐trisphosphate; phospholipase C; platelets; X-linked immunodeficiency
• ISSN: 0261-4189; 1460-2075; 1460-2075
• DOI: 10.1093/emboj/19.12.2793

The role of phosphatidylinositol 3,4,5‐trisphosphate (PI3,4,5P 3 ) and Btk in signalling by the collagen receptor glycoprotein VI was investigated. PI3,4,5P 3 was increased in platelets from mice deficient in the SH2 domain‐containing inositol 5‐phosphatase (SHIP), in response to collagen related peptide (CRP). Tyrosine phosphorylation and activation of phospholipase Cγ2 (PLCγ2) were unaltered in SHIP −/− platelets, whereas Btk was heavily tyrosine phosphorylated under basal conditions and maximally phosphorylated by low concentrations of CRP. There was an increase in basal Ca 2+ , maximal expression of P‐selectin, and potentiation of Ca 2+ and aminophospholipid exposure to CRP in SHIP −/− platelets in the presence of Ca 2+ (1 mM). Microinjection of PI3,4,5P 3 into megakaryocytes caused a 3‐fold increase in Ca 2+ in response to CRP, which was absent in X‐linked immunodeficiency (Xid) mice, which have a mutation in the PH domain of Btk. There was a corresponding partial reduction in the sustained level of intracellular Ca 2+ in response to CRP in Xid mice but no change in PLC activity. These results demonstrate a novel pathway of Ca 2+ entry that involves PI3,4,5P 3 and Btk, and which is independent of increased PLC activity.