Loss of PIP5KIbeta demonstrates that PIP5KI isoform-specific PIP2 synthesis is required for IP3 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 properly in vivo. Together, these data demonstrate that PIP5KIβ is required for rapid PIP₂ synthesis, second-messenger production, and stable platelet adhesion under shear in 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.