Arf Proteins and Their Regulators: At the Interface Between Membrane Lipids and the Protein Trafficking Machinery - Ras Superfamily Small G Proteins: Biology and Mechanisms 2 - page 156

curved membranes of COPI vesicles (Bigay et al. 2003 , 2005 ; Mesmin et al. 2007 ).
In an elegant self-organizing mechanism, COPI vesicles are generated through
activation of Arf1 at the Golgi, which recruits COPI to deform the membrane
into a highly curved vesicle, which then recruits ArfGAP1 through its ALPS
motif to hydrolyse the GTP on Arf1, releasing the coat and allowing fusion of the
vesicle with its target membrane (Bigay et al. 2003 ). ArfGAP2 and 3 also bind to
COPI vesicles, but do so through direct interaction with the COPI coat, and hence
can also bind to COPI-coated regions that are not highly curved (Kliouchnikov
et al. 2009 ; Weimer et al. 2008 ). ArfGAP1, 2, and 3 and their yeast orthologues
Gcs1p and Glo3p function at the early Golgi (Spang et al. 2010 ), whereas most of
the other Arf GAPs function at the cell periphery in TGN-endosomal-plasma
membrane trafficking and actin cytoskeleton remodelling (Inoue and Randazzo
2007 ; Sabe et al. 2006 ). The following chapter in this volume will describe the
latter class of Arf GAPs in depth.
8.2 Localization and Functions of Arf Proteins
A major distinguishing feature of the Arf proteins is the presence of a myristoylated
amino-terminal amphipathic helix that is necessary for membrane binding
(Fig. 8.1a, b, c ). Myristoylation is a cotranslational modification required for the
essential functions of Arf1 in vivo (Kahn et al. 1995 ). In cells, myristoylation is
required for correct localization of Arf proteins, including Golgi localization of
Arf1 and PM localization of Arf6 (Donaldson and Jackson 2011 ). In vitro, the
myristoyl group of the GDP-bound form of Arf1 is available for membrane
insertion, permitting a weak association of the inactive form of Arf1 with mem-
branes (Franco et al. 1995 ). Upon GTP binding, the N-terminal amphipathic helix
of Arf1 is released and inserts into the membrane, resulting in tight membrane
association (Antonny et al. 1997 ) (Fig. 8.1c ). Structural studies revealed the details
of this change in conformation, showing that the interswitch region changes
position to occlude the hydrophobic pocket that harbours the amphipathic
N-terminal helix in the GDP-bound form of Arf1 (Goldberg 1998 ). NMR studies
of N-myristoylated Arf1-GTP further confirmed this mechanism (Liu et al. 2010 ).
Thus, in addition to changes in the effector binding regions upon exchange of GDP
for GTP, Arf proteins undergo a second change in conformation that brings them
into very close contact with the membrane (Fig. 8.1c ) (Antonny et al. 1997 ;
Chavrier and Menetrey 2010 ). This property distinguishes them from other small
G proteins of the Ras superfamily, including the Ras, Rho, and Rab Families, which
have a long C-terminal linker to which their lipid membrane anchor is attached
(Gillingham and Munro 2007b ). Arf effectors are thus constrained to a position
close to the membrane, in contrast to those of Rab and Rho, which can be located at
a distance from the membrane surface (Gillingham and Munro 2007b ; Khan and
Menetrey 2013 ).