The Arl4 Family of Small G Proteins Can Recruit the Cytohesin Arf6 Exchange Factors to the Plasma Membrane

SummaryThe small GTPase Arf6 regulates endocytosis, actin dynamics, and cell adhesion, and one of its major activators is the exchange factor Arf nucleotide-binding site opener (ARNO), also called cytohesin-2 1 and 2. ARNO must be recruited from the cytosol to the plasma membrane in order to activate Arf6, and in addition to a Sec7 nucleotide-exchange domain it contains a C-terminal pleckstrin homology (PH) domain that binds phosphoinositides 3 and 4. ARNO and its three relatives, cytohesin-1, Grp1/cytohesin-3, and cytohesin-4, are expressed as two splice variants, with either two or three glycines in a loop in the phosphoinositide-binding pocket of the PH domain 5 and 6. The diglycine form binds PtdIns(3,4,5)P3 with high affinity and mediates recruitment of cytohesins to the plasma membrane in response to insulin and growth factors 7 and 8. However, the triglycine form has only micromolar affinity for both PtdIns(3,4,5)P3 and PtdIns(4,5)P2, affinities that are insufficient to confer membrane recruitment, raising the question of how the triglycine forms of cytohesins are regulated 5 and 9. Here we show that three related Arf-like GTPases of unknown function, Arl4a, Arl4c, and Arl4d, are able to recruit ARNO and other cytohesins to the plasma membrane by binding to their PH domains irrespective of whether they are in the diglycine or triglycine form. The Arl4 family thus defines a signal-transduction pathway that can mediate the plasma-membrane recruitment of cytohesins independently of a requirement for the generation of PtdIns(3,4,5)P3.