Identification of ARAP3, a novel PI3K effector regulating both Arf and Rho GTPases, by selective capture on phosphoinositide affinity matrices

S Krugmann; K E Anderson; S H Ridley; N Risso; A McGregor; J Coadwell; K Davidson; A Eguinoa; C D Ellson; P Lipp; M Manifava; N Ktistakis; G Painter; J W Thuring; M A Cooper; Z-Y Lim; A B Holmes; S K Dove; R H Michell; A Grewal; A Nazarian; H Erdjument-Bromage; P Tempst; L R Stephens; P T Hawkins

doi:10.1016/s1097-2765(02)00434-3

Identification of ARAP3, a novel PI3K effector regulating both Arf and Rho GTPases, by selective capture on phosphoinositide affinity matrices

Mol Cell. 2002 Jan;9(1):95-108. doi: 10.1016/s1097-2765(02)00434-3.

Affiliation

¹ Inositide Laboratory, The Babraham Institute, Cambridge, CB2 4AT, United Kingdom.

PMID: 11804589
DOI: 10.1016/s1097-2765(02)00434-3

Abstract

We show that matrices carrying the tethered homologs of natural phosphoinositides can be used to capture and display multiple phosphoinositide binding proteins in cell and tissue extracts. We present the mass spectrometric identification of over 20 proteins isolated by this method, mostly from leukocyte extracts: they include known and novel proteins with established phosphoinositide binding domains and also known proteins with surprising and unusual phosphoinositide binding properties. One of the novel PtdIns(3,4,5)P3 binding proteins, ARAP3, has an unusual domain structure, including five predicted PH domains. We show that it is a specific PtdIns(3,4,5)P3/PtdIns(3,4)P2-stimulated Arf6 GAP both in vitro and in vivo, and both its Arf GAP and Rho GAP domains cooperate in mediating PI3K-dependent rearrangements in the cell cytoskeleton and cell shape.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

ADP-Ribosylation Factor 6
ADP-Ribosylation Factors / metabolism*
Adaptor Proteins, Signal Transducing*
Animals
COS Cells
Carrier Proteins / genetics
Carrier Proteins / metabolism*
Cloning, Molecular
Cytosol / metabolism
GTPase-Activating Proteins / genetics
GTPase-Activating Proteins / metabolism*
Leukocytes / metabolism*
Leukocytes / ultrastructure
Mass Spectrometry
Molecular Sequence Data
Phosphatidylinositol 3-Kinases / metabolism*
Phosphatidylinositol Phosphates / metabolism*
Protein Binding
Proteins / genetics
Proteins / metabolism*
Recombinant Proteins / metabolism
Signal Transduction
Swine
rho GTP-Binding Proteins / metabolism*

Substances

ADP-Ribosylation Factor 6
ARAP3 protein, human
Adaptor Proteins, Signal Transducing
Carrier Proteins
GTPase-Activating Proteins
Phosphatidylinositol Phosphates
Proteins
Recombinant Proteins
phosphatidylinositol 3,4,5-triphosphate
ADP-Ribosylation Factors
ARF6 protein, human
rho GTP-Binding Proteins

Associated data

GENBANK/AJ310567