The structural basis of Arfaptin-mediated cross-talk between Rac and Arf signalling pathways.
Small G proteins are GTP-dependent molecular switches that regulate numerous cellular functions. They can be classified into homologous subfamilies that are broadly associated with specific biological processes. Cross-talk between small G-protein families has an important role in signalling, but the mechanism by which it occurs is poorly understood. The coordinated action of Arf and Rho family GTPases is required to regulate many cellular processes including lipid signalling, cell motility and Golgi function. Arfaptin is a ubiquitously expressed protein implicated in mediating cross-talk between Rac (a member of the Rho family) and Arf small GTPases. Here we show that Arfaptin binds specifically to GTP-bound Arf1 and Arf6, but binds to Rac.GTP and Rac.GDP with similar affinities. The X-ray structure of Arfaptin reveals an elongated, crescent-shaped dimer of three-helix coiled-coils. Structures of Arfaptin with Rac bound to either GDP or the slowly hydrolysable analogue GMPPNP show that the switch regions adopt similar conformations in both complexes. Our data highlight fundamental differences between the molecular mechanisms of Rho and Ras family signalling, and suggest a model of Arfaptin-mediated synergy between the Arf and Rho family signalling pathways.
Tarricone
C
C
Division of Protein Structure, National Institute for Medical Research, Mill Hill, London NW7 IAA, UK.
Xiao
B
B
Justin
N
N
Walker
P A
PA
Rittinger
K
K
Gamblin
S J
SJ
Smerdon
S J
SJ
D020823:ADP-Ribosylation Factor 1 / Q000235:genetics / Q000378:metabolism; D000089982:ADP-Ribosylation Factor 6; D020727:ADP-Ribosylation Factors / Q000235:genetics / Q000378:metabolism*; D048868:Adaptor Proteins, Signal Transducing; D000595:Amino Acid Sequence; D001667:Binding, Competitive; D002151:Calorimetry; D002352:Carrier Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D018360:Crystallography, X-Ray; D019281:Dimerization; D005454:Fluorescence Polarization; D006153:Guanosine Diphosphate / Q000378:metabolism; D006160:Guanosine Triphosphate / Q000378:metabolism; D006165:Guanylyl Imidodiphosphate / Q000378:metabolism; D006801:Humans; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009154:Mutation / Q000235:genetics; D011485:Protein Binding; D011487:Protein Conformation; D016415:Sequence Alignment; D015398:Signal Transduction; D013696:Temperature; D017186:Titrimetry; D020744:rac GTP-Binding Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*
D000089982:ADP-Ribosylation Factor 6; C101158:ARFIP2 protein, human; D048868:Adaptor Proteins, Signal Transducing; D002352:Carrier Proteins; D006153:Guanosine Diphosphate; D006165:Guanylyl Imidodiphosphate; D006160:Guanosine Triphosphate; D020823:ADP-Ribosylation Factor 1; D020727:ADP-Ribosylation Factors; C000715348:ARF6 protein, human; D020744:rac GTP-Binding Proteins