Distinct Rab-binding domains mediate the interaction of Rabaptin-5 with GTP-bound Rab4 and Rab5.
Rabaptin-5 functions as an effector for the small GTPase Rab5, a regulator of endocytosis and early endosome fusion. We have searched for structural determinants that confer functional specificity on Rabaptin-5. Here we report that native cytosolic Rabaptin-5 is present in a homodimeric state and dimerization depends upon the presence of its coiled-coil predicted sequences. A 73 residue C-terminal region of Rabaptin-5 is necessary and sufficient both for the interaction with Rab5 and for Rab5-dependent recruitment of the protein on early endosomes. Surprisingly, we uncovered the presence of an additional Rab-binding domain at the N-terminus of Rabaptin-5. This domain mediates the direct interaction with the GTP-bound form of Rab4, a small GTPase that has been implicated in recycling from early endosomes to the cell surface. Based on these results, we propose that Rabaptin-5 functions as a molecular linker between two sequentially acting GTPases to coordinate endocytic and recycling traffic.
Vitale
G
G
European Molecular Biology Laboratory, Postfach 10.2209, D-69012 Heidelberg, Germany.
Rybin
V
V
Christoforidis
S
S
Thornqvist
P
P
McCaffrey
M
M
Stenmark
H
H
Zerial
M
M
D000595:Amino Acid Sequence; D000818:Animals; D002417:Cattle; D002460:Cell Line; D006224:Cricetinae; D003600:Cytosol / Q000737:chemistry; D019281:Dimerization; D011992:Endosomes / Q000378:metabolism; D019204:GTP-Binding Proteins / Q000032:analysis / Q000378:metabolism*; D006160:Guanosine Triphosphate / Q000378:metabolism*; D006367:HeLa Cells; D006801:Humans; D008565:Membrane Proteins / Q000032:analysis / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D011485:Protein Binding; D017433:Protein Structure, Secondary; D011993:Recombinant Fusion Proteins; D016415:Sequence Alignment; D033921:Vesicular Transport Proteins; D020695:rab4 GTP-Binding Proteins; D020696:rab5 GTP-Binding Proteins
D008565:Membrane Proteins; C097227:RABEP1 protein, human; D011993:Recombinant Fusion Proteins; D033921:Vesicular Transport Proteins; D006160:Guanosine Triphosphate; D019204:GTP-Binding Proteins; D020695:rab4 GTP-Binding Proteins; D020696:rab5 GTP-Binding Proteins