title: GAP1IP4BP contains a novel group I pleckstrin homology domain that directs constitutive plasma membrane association.
abstract: The group I family of pleckstrin homology (PH) domains are characterized by their inherent ability to specifically bind phosphatidylinositol 3,4,5-trisphosphate (PtdIns(3,4,5)P(3)) and its corresponding inositol head-group inositol 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 PH domain-containing proteins following agonist-stimulated PtdIns(3,4,5)P(3) production. Among group I PH domain-containing proteins, the Ras GTPase-activating protein GAP1(IP4BP) is unique in being constitutively associated with the plasma membrane. Here we show that, although the GAP1(IP4BP) PH domain interacts with PtdIns(3,4, 5)P(3), it also binds, with a comparable affinity, phosphatidylinositol 4,5-bisphosphate (PtdIns(4,5)P(2)) (K(d) values of 0.5 +/- 0.2 and 0.8 +/- 0.5 microm, respectively). Intriguingly, whereas this binding site overlaps with that for Ins(1,3,4,5)P(4), consistent with the constitutive 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) from this membrane. Thus, the Ins(1,3,4,5)P(4)-binding PH domain from GAP1(IP4BP) 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 regulated in vivo by Ins(1,3,4,5)P(4) rather than PtdIns(3,4,5)P(3).
authors.affiliation: Department of Biochemistry, School of Medical Sciences, University of Bristol, Bristol BS8 1TD, United Kingdom.
authors.forename: G E
authors.initials: GE
authors.lastname: Cozier
authors.forename: P J
authors.initials: PJ
authors.lastname: Lockyer
authors.forename: J S
authors.initials: JS
authors.lastname: Reynolds
authors.forename: S
authors.initials: S
authors.lastname: Kupzig
authors.forename: J R
authors.initials: JR
authors.lastname: Bottomley
authors.forename: T H
authors.initials: TH
authors.lastname: Millard
authors.forename: G
authors.initials: G
authors.lastname: Banting
authors.forename: P J
authors.initials: PJ
authors.lastname: Cullen
mesh_terms: D019943:Amino Acid Substitution; D000818:Animals; D001665:Binding Sites; D019556:COS Cells; D002462:Cell Membrane / Q000378:metabolism*; D002467:Cell Nucleus / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D007295:Inositol Phosphates / Q000378:metabolism*; D008081:Liposomes; D016297:Mutagenesis, Site-Directed; D018129:Phosphatidylinositol Phosphates / Q000378:metabolism*; D018160:Receptors, Cytoplasmic and Nuclear / Q000737:chemistry* / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000378:metabolism; D013347:Subcellular Fractions / Q000378:metabolism; D013395:Sucrose; D014162:Transfection
chemical_list: D007295:Inositol Phosphates; D008081:Liposomes; D018129:Phosphatidylinositol Phosphates; D018160:Receptors, Cytoplasmic and Nuclear; D011993:Recombinant Fusion Proteins; C061107:inositol-1,3,4,5-tetrakisphosphate receptor; C060974:phosphatidylinositol 3,4,5-triphosphate; C081655:inositol-1,3,4,5-tetrakisphosphate; D013395:Sucrose