The DHR1 domain of DOCK180 binds to SNX5 and regulates cation-independent mannose 6-phosphate receptor transport.
DOCK180 is the archetype of the DOCK180-family guanine nucleotide exchange factor for small GTPases Rac1 and Cdc42. DOCK180-family proteins share two conserved domains, called DOCK homology region (DHR)-1 and -2. Although the function of DHR2 is to activate Rac1, DHR1 is required for binding to phosphoinositides. To better understand the function of DHR1, we searched for its binding partners by direct nanoflow liquid chromatography/tandem mass spectrometry, and we identified sorting nexins (SNX) 1, 2, 5, and 6, which make up a multimeric protein complex mediating endosome-to-trans-Golgi-network (TGN) retrograde transport of the cation-independent mannose 6-phosphate receptor (CI-MPR). Among these SNX proteins, SNX5 was coimmunoprecipitated with DOCK180 most efficiently. In agreement with this observation, DOCK180 colocalized with SNX5 at endosomes. The RNA interference-mediated knockdowns of SNX5 and DOCK180, but not Rac1, resulted in the redistribution of CI-MPR from TGN to endosomes. Furthermore, expression of the DOCK180 DHR1 domain was sufficient to restore the perturbed CI-MPR distribution in DOCK180 knockdown cells. These data suggest that DOCK180 regulates CI-MPR trafficking via SNX5 and that this function is independent of its guanine nucleotide exchange factor activity toward Rac1.
Hara
Shigeo
S
Department of Pathology and Biology of Diseases, Graduate School of Medicine, Kyoto University, Kyoto 606-8501, Japan.
Kiyokawa
Etsuko
E
Iemura
Shun-ichiro
S
Natsume
Tohru
T
Wassmer
Thomas
T
Cullen
Peter J
PJ
Hiai
Hiroshi
H
Matsuda
Michiyuki
M
D002352:Carrier Proteins / Q000378:metabolism*; D002412:Cations; D011992:Endosomes / Q000378:metabolism; D005786:Gene Expression Regulation; D006056:Golgi Apparatus; D006367:HeLa Cells; D006801:Humans; D008954:Models, Biological; D017434:Protein Structure, Tertiary; D034622:RNA Interference; D017527:Receptor, IGF Type 2 / Q000378:metabolism*; D058305:Sorting Nexins; D033921:Vesicular Transport Proteins / Q000378:metabolism*; D020744:rac GTP-Binding Proteins / Q000378:metabolism*; D020830:rac1 GTP-Binding Protein / Q000378:metabolism
D002352:Carrier Proteins; D002412:Cations; C099818:DOCK1 protein, human; C107953:RAC1 protein, human; D017527:Receptor, IGF Type 2; C402667:SNX5 protein, human; D058305:Sorting Nexins; D033921:Vesicular Transport Proteins; D020744:rac GTP-Binding Proteins; D020830:rac1 GTP-Binding Protein