Role of Rab9 GTPase in facilitating receptor recruitment by TIP47

Science

Volumn 292, Issue 5520, 2001, Pages 1373-1376

  Carroll, Kate S ^a   Hanna, John ^a   Simon, Iris ^a,b   Krise, Jeff ^a   Barbero, Pierre ^a   Pfeffer, Suzanne R ^a  

^a STANFORD UNIVERSITY SCHOOL OF MEDICINE   (United States)

^b Eos Biotechnology   (United States)

Author keywords

[No Author keywords available]

Indexed keywords

CONFORMATIONS; CYTOLOGY; PHOSPHATES;

VESICLE BUDDING;

ENZYMES;

ADAPTOR PROTEIN; GUANOSINE TRIPHOSPHATASE; LYSOSOME ENZYME; MEMBRANE PROTEIN; PROTEIN TIP47; RAB PROTEIN; RAB9 GUANOSINE TRIPHOSPHATASE; SOMATOMEDIN B RECEPTOR; UNCLASSIFIED DRUG;

ARTICLE; BINDING AFFINITY; BINDING SITE; ENDOSOME; GOLGI COMPLEX; NONHUMAN; PRIORITY JOURNAL; PROTEIN BINDING; PROTEIN CONFORMATION; PROTEIN TRANSPORT;

AMINO ACID SUBSTITUTION; ANIMALS; BINDING SITES; CATTLE; CYTOPLASM; DNA-BINDING PROTEINS; ENDOSOMES; GOLGI APPARATUS; GUANOSINE 5'-O-(3-THIOTRIPHOSPHATE); INTRACELLULAR SIGNALING PEPTIDES AND PROTEINS; PREGNANCY PROTEINS; PROTEIN BINDING; PROTEIN STRUCTURE, TERTIARY; PROTEIN TRANSPORT; RAB GTP-BINDING PROTEINS; RECEPTOR, IGF TYPE 2; RECOMBINANT FUSION PROTEINS; SUBSTRATE SPECIFICITY;

EID: 0035906951     PISSN: 00368075     EISSN: None     Source Type: Journal    
DOI: 10.1126/science.1056791     Document Type: Article

References (19)

1. S. Kornfeld, Annu. Rev. Biochem. 61, 307 (1992).
2. _, 1. Mellman, Annu. Rev. Cell Biol. 5, 483 (1989).
3. E. Diaz, S. R. Pfeffer, Cell 93, 433 (1998).
4. D. Lombardi, et al. EMBO J. 12, 677 (1993).
5. M. A. Riederer, T. Soldati, A. D. Shapiro, J. Lin, S. R. Pfeffer, J. Cell Biol. 125, 573 (1994).
6. His-tagged TIP47 was purified after expression in Escherichia coli (3); 10% glycerol was added. GST-CI-MPR cytoplasmic domain and TIP47 constructs, and antibodies against TIP47 were as described (3). Texas Red sulfonyl chloride (Molecular Probes, Eugene, Oregon), was covalently attached to TIP47 and used for binding as described (7). Cytosolic TIP47 was purified from bovine kidney cytosol (3.6 g) by chromatography on Fast Flow Q-Sepharose (18 ml), eluted with 0 to 400 mM NaCl in 25 mM Tris pH 7.5. Fractions were dialyzed into 10 mM Na phosphate pH 7.5 and applied onto a hydroxyapatite column (7 ml), eluted with a Na phosphate gradient at pH 7.5. Purification for Fig. 1A included an additional anion exchange step at pH 6.8. This material is 150-fold enriched for TIP47.
7. J. P. Krise, P. M. Sincock, J. G. Orsel, S. R. Pfeffer. J. Biol. Chem. 275, 25188 (2000).
8. For Rab9-GTP we utilized Rab9 CLLL, which is a wild-type protein with an altered COOH-terminus, to enhance stability upon expression in E. coli. The protein shows entirely wild-type nucleotide binding properties, becomes mono-geranylgeranylated in vitro, and can support in vitro transport of the CI-MPR. Rab9 Q66L was used in Fig. 1 only; this protein was generated by polymerase chain reaction mutagenesis and is defective in nucleotide hydrolysis, favoring the GTP-bound state.
9. P. Chavrier, M. Vingron, C. Sander, K. Simons, M. Zerial, Mol. Cell Biol. 10, 6578 (1990).
10. Y. Feng, B. Press, A. Wandinger-Ness, J. Cell Biol. 131, 1435 (1995).
11. F. Schimmöller, I. Simon, S. R. Pfeffer. J. Biol. Chem. 273, 22161 (1998).
12. 167AlaAla were transiently transfected into cells stably expressing a CD-MPR that contains a tyrosine sulfation site to enable monitoring of endosome-to-TGN transport in vivo (13). Transport was measured 43 hours after Fugene (Roche Diagnostics, Indianapolis) transfection as described (13). Data were corrected for the efficiency of transfection (36%), which was determined in parallel by immunoftuorescence microscopy using cells transfected with a plasmid encoding green fluorescent protein.
13. C. Itin, C. Rancaño, Y. Nakajima, S. R. Pfeffer, S. R. J. Biol. Chem. 272, 27737 (1997).
14. S. R. Pfeffer, Nature Cell Biol. 1, E17 (1999).
15. P. Novick, M. Zerial, Curr. Opin. Cell Biol. 9, 496 (1997).
16. G. Jedd, J. Mulholland, N. Segev, J. Cell Biol. 137, 563 (1997).
17. H. McLauchlan et al., Curr. Biol. 8, 34 (1998).
18. T. Soldati, M. A. Riederer, S. R. Pfeffer, Mol. Biol. Cell 4, 425 (1993).
19. Research was supported by a research grant (DK37332) from the National Institutes of Health; I.S. and J.K. were postdoctoral fellows of the Deutsche Forschungsgemeinschaft and the Pharmaceutical Research and Manufacturers of America Foundation, respectively; K.S.C. was a predoctoral trainee of NIH. We thank L. Ding and B. O'Connor for contributions to the identification of the Rab9 binding site.