Gamma-adaptin interacts directly with Rabaptin-5 through its ear domain.
In yeast two-hybrid screening using gamma1-adaptin, a subunit of the AP-1 adaptor complex of clathrin-coated vesicles derived from the trans-Golgi network (TGN), as bait, we found that it could interact with Rabaptin-5, an effector of Rab5 and Rab4 that regulates membrane docking with endosomes. Further two-hybrid analysis revealed that the interaction occurs between the ear domain of gamma1-adaptin and the COOH-terminal coiled-coil region of Rabaptin-5. Pull down assay with a fusion protein between glutathione S-transferase and the ear domain of gamma1-adaptin and coimmunoprecipitation analysis revealed that the interaction occurs in vitro and in vivo. Immunocytochemical analysis showed that gamma1-adaptin and Rabaptin-5 colocalize to a significant extent on perinuclear structures, probably on recycling endosomes, and are redistributed into the cytoplasm upon treatment with brefeldin A. These results suggest that the gamma1-adaptin-Rabaptin-5 interaction may play a role in membrane trafficking between the TGN and endosomes.
Shiba
Yoko
Y
Institute of Biological Sciences and Gene Research Center, University of Tsukuba, Tsukuba Science City, Ibaraki 305-8572, Japan.
Takatsu
Hiroyuki
H
Shin
Hye-Won
HW
Nakayama
Kazuhisa
K
D033982:Adaptor Protein Complex gamma Subunits; D000818:Animals; D020126:Brefeldin A / Q000494:pharmacology; D002478:Cells, Cultured; D003593:Cytoplasm / Q000378:metabolism; D011992:Endosomes / Q000378:metabolism* / Q000648:ultrastructure; D006367:HeLa Cells / Q000187:drug effects / Q000378:metabolism; D006801:Humans; D007150:Immunohistochemistry; D008565:Membrane Proteins / Q000378:metabolism*; D017434:Protein Structure, Tertiary; D021381:Protein Transport / Q000502:physiology; D051381:Rats; D020798:Two-Hybrid System Techniques; D033921:Vesicular Transport Proteins; D021601:trans-Golgi Network / Q000378:metabolism*
D033982:Adaptor Protein Complex gamma Subunits; D008565:Membrane Proteins; C097227:RABEP1 protein, human; D033921:Vesicular Transport Proteins; D020126:Brefeldin A