Characterization of novel Rab6-interacting proteins involved in endosome-to-TGN transport.
Rab6 GTPase regulates intracellular transport at the level of the Golgi complex. Using the yeast two-hybrid screen, we have isolated two clones that specifically interact with the three isoforms of Rab6 present in mammalian cells (Rab6A, A' and B). The cDNAs encode two proteins of 976 and 1120 amino acids (calculated molecular mass of 112 and 128 kDa, respectively) that we named Rab6IP2A and Rab6IP2B (for Rab6 Interacting Protein 2). The two proteins likely correspond to spliced variants of the same gene. Rab6IP2s have no significant homology with other known proteins, including Rab effectors or partners. They are ubiquitously expressed, mostly cytosolic and found in high molecular mass complexes in brain cytosol. We show that Rab6IP2s can be recruited on Golgi membranes in a Rab6:GTP-dependent manner. The overexpression of any form of Rab6IP2 has no detectable effect on the secretory pathway. In contrast, the retrograde transport of the Shiga toxin B subunit between the plasma membrane and the Golgi complex is partly inhibited in cells overexpressing the Rab6-binding domain of Rab6IP2. Our data suggest that Rab6IP2s is involved in the pathway regulated by Rab6A'.
Monier
Solange
S
Laboratories Molecular mechanisms of intracellular transport, Institut Curie, 26 rue d'Ulm, 75248 Paris Cedex 05, France. smonier@curie.fr
Jollivet
Florence
F
Janoueix-Lerosey
Isabelle
I
Johannes
Ludger
L
Goud
Bruno
B
D000595:Amino Acid Sequence; D000818:Animals; D001692:Biological Transport; D015152:Blotting, Northern; D001921:Brain / Q000378:metabolism; D002417:Cattle; D002850:Chromatography, Gel; D003001:Cloning, Molecular; D003600:Cytosol / Q000378:metabolism; D018076:DNA, Complementary / Q000378:metabolism; D011992:Endosomes / Q000378:metabolism*; D015723:Gene Library; D005982:Glutathione Transferase / Q000378:metabolism; D006056:Golgi Apparatus / Q000378:metabolism*; D006160:Guanosine Triphosphate / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D051379:Mice; D008856:Microscopy, Fluorescence; D008969:Molecular Sequence Data; D010957:Plasmids / Q000378:metabolism; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D021122:Protein Subunits; D017386:Sequence Homology, Amino Acid; D022641:Shiga Toxin 2 / Q000378:metabolism; D014018:Tissue Distribution; D014162:Transfection; D020798:Two-Hybrid System Techniques; D020691:rab GTP-Binding Proteins / Q000096:biosynthesis / Q000235:genetics* / Q000378:metabolism*; D021601:trans-Golgi Network / Q000378:metabolism*; D009419:Nerve Tissue Proteins
D018076:DNA, Complementary; D005982:Glutathione Transferase; D006160:Guanosine Triphosphate; D021122:Protein Subunits; D022641:Shiga Toxin 2; D020691:rab GTP-Binding Proteins; C094135:Rab6 protein; C466022:Rab6IP2A protein, mouse; C450841:shiga toxin 2 B subunit; C486769:Erc1 protein, mouse; D009419:Nerve Tissue Proteins