Loss of PIP5KIβ demonstrates that PIP5KI isoform-specific PIP 2 synthesis is required for IP 3 formation

The three isoforms of PIP5KI (α, β, and γ) synthesize PI4,5P2 (PIP2 ) by phosphorylating PI4P. Therefore, it is not clear why platelets, like all eukaryotic cells, have more than one isoform. To test the hypothesis that PIP5KI isoforms have nonoverlapping functions, we generated a murine line containing a null mutation of PIP5KIβ and analyzed the effect on platelet signaling. PIP5KIβ-null mice had normal platelet counts. In contrast to platelets lacking PIP5KIα, platelets lacking PIP5KIβ exhibited impaired aggregation accompanied by disaggregation. Although platelets lacking PIP5KIβ had only a moderate deficiency of PIP2 under basal conditions, they had a striking deficiency in PIP2 synthesis and IP3 formation after thrombin stimulation. We have also observed that platelets lacking both PIP5KIα and PIP5KIβ have a complete loss of thrombin-induced IP3 synthesis even though they still contain PIP5KIγ, the predominant PIP5KI isoform in platelets. These results demonstrate that PIP5KIβ, like PIP5KIα, contributes to the rapid synthesis of a pool of PIP2 that is required for second-messenger formation, whereas the pool of PIP2 synthesized by PIP5KIγ does not contribute to this process. Additionally, we found that PIP5KIβ-null platelets failed to form arterial thrombi properlyin vivo . Together, these data demonstrate that PIP5KIβ is required for rapid PIP2 synthesis, second-messenger production, and stable platelet adhesion under shearin vivo . These results also demonstrate that after stimulation of a G protein-coupled receptor, IP3 is completely derived from a rapidly synthesized discrete pool of PIP2 synthesized by PIP5KIα and PIP5KIβ.