GAP1(IP4BP) contains a novel group I pleckstrin homology domain that directs constitutive plasma membrane association

The group I family of pleckstrin homology (PH) domains are characterized by their inherent ability to specifically bind phosphatidylinositol 3,4,5-tri sphosphate (PtdIns(3,4,5)P-3) and its corresponding inositol headgroup inos itol 1,3,4,5-tetrakisphosphate (Ins(1,3,4,5)P-4). In vivo this interaction results in the regulated plasma membrane recruitment of cytosolic group I P H domain-containing proteins following agonist-stimulated PtdIns(3,4,5)P-3 production. Among group I PH domain-containing proteins, the Pas GTPase-act ivating protein GAP1(IP4BP) is unique in being constitutively associated wi th the plasma membrane. Here we show that, although the GAP1(IP4BP) PH doma in interacts with PtdIns(3,4,5)P-3, it also binds, with a comparable affini ty, phosphatidylinositol 4,5-bisphosphate (PtdIns(4,5)P-2) (K-d values of 0 .5 +/- 0.2 and 0.8 +/- 0.5 mu M, respectively). Intriguingly, whereas this binding site overlaps with that for Ins(1,3,4,5)P-4, consistent with the co nstitutive plasma membrane association of GAP1(IP4BP) resulting from its PH domain-binding PtdIns(4,5)P-2, we show that in vivo depletion of PtdIns(4, 5)P-2, but not PtdIns(3,4,5)P-3, results in dissociation of GAP1(IP4BP) fro m this membrane. Thus, the Ins(1,3,4,5)P-4-binding PH domain from GAP1(IP4B P) defines a novel class of group I PH domains that constitutively targets the protein to the plasma membrane and may allow GAP1(IP4BP) to be regulate d in vivo by Ins(1,3,4,5)P-4 rather than PtdIns(3,4,5)P-3.