ArfGAP1 interacts with coat proteins through tryptophan-based motifs.
The Arf1 GTPase-activating protein ArfGAP1 regulates vesicular traffic through the COPI system. This protein consists of N-terminal catalytic domain, lipid packing sensors (the ALPS motifs) in the central region, and a carboxy part of unknown function. The carboxy part contains several diaromatic sequences that are reminiscent of motifs known to interact with clathrin adaptors. In pull-down experiments using GST-fused peptides from rat ArfGAP1, a peptide containing a (329)WETF sequence interacted strongly with clathrin adaptors AP1 and AP2, whereas a major coatomer-binding determinant was identified within the extreme carboxy terminal peptide ((405)AADEGWDNQNW). Mutagenesis and peptide competition experiments revealed that this determinant is required for coatomer binding to full-length ArfGAP1, and that interaction is mediated through the delta-subunit of the coatomer adaptor-like subcomplex. Evidence for a role of the carboxy motif in ArfGAP1-coatomer interaction in vivo is provided by means of a reporter fusion assay. Our findings point to mechanistic differences between ArfGAP1 and the other ArfGAPs known to function in the COPI system.
Rawet
Moran
M
Department of Biology, Technion-Israel Institute of Technology, Haifa 32000, Israel.
Levi-Tal
Sharon
S
Szafer-Glusman
Edith
E
Parnis
Anna
A
Cassel
Dan
D
D033961:Adaptor Protein Complex 1 / Q000378:metabolism; D033962:Adaptor Protein Complex 2 / Q000378:metabolism; D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D000818:Animals; D020755:Coat Protein Complex I / Q000378:metabolism*; D020690:GTPase-Activating Proteins / Q000235:genetics / Q000378:metabolism*; D051381:Rats; D014364:Tryptophan / Q000235:genetics / Q000378:metabolism
D033961:Adaptor Protein Complex 1; D033962:Adaptor Protein Complex 2; C503239:Arfgap1 protein, rat; D020755:Coat Protein Complex I; D020690:GTPase-Activating Proteins; D014364:Tryptophan