Effects of acid phospholipids on ARF activities: Potential roles in membrane traffic

ADP-ribosylation factors are a family of ∼ 21 kDa GTP binding proteins which have been implicated as ubiquitous regulators of multiple steps in both exocytic and endocytic membrane traffic in mammals and yeast. Reversible membrane associations are thought to be an essential component in the physiological actions of ARF and are regulated by GTP binding. ARFs are unique among the superfamily of GTP binding proteins in having a strict dependence on phospholipids for nucleotide exchange. In addition, ARF proteins were found to bind phospatidylinositol 4,5-bisphosphate (PIP₂) specifically. PIP₂ was found to increase the rate of GDP dissociation and stabilize the nucleotide-free form of the protein. The previously described requirements for PIP₂ in the ARF stimulated phospholipase D (PLD) activity and ARF GTPase activating protein (ARF GAP) assays provide the basis for a model in which PIP₂ acts as a cofactor in one or more ARF pathways. There are potentially two distinct phospholipid binding sites each of which are coupled to the nucleotide binding site of ARFs.