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Molecular and Cellular Biology, January 2007, p. 605-621, Vol. 27, No. 2
0270-7306/07/$08.00+0     doi:10.1128/MCB.00699-06
Copyright © 2007, American Society for Microbiology. All Rights Reserved.

Btn2, a Hook1 Ortholog and Potential Batten Disease-Related Protein, Mediates Late Endosome-Golgi Protein Sorting in Yeast

Department of Molecular Genetics, Weizmann Institute of Science, Rehovot 76100, Israel

Received 23 April 2006/ Returned for modification 30 June 2006/ Accepted 1 November 2006

BTN2 gene expression in the yeast Saccharomyces cerevisiae is up-regulated in response to the deletion of BTN1, which encodes the ortholog of a human Batten disease protein. We isolated Btn2 as a Snc1 v-SNARE binding protein using the two-hybrid assay and examined its role in intracellular protein trafficking. We show that Btn2 is an ortholog of theDrosophila and mammalian Hook1 proteins that interact with SNAREs, cargo proteins, and coat components involved in endosome-Golgi protein sorting. By immunoprecipitation, it was found that Btn2 bound the yeast endocytic SNARE complex (e.g., Snc1 and Snc2 [Snc1/2], Tlg1, Tlg2, and Vti1), the Snx4 sorting nexin, and retromer (e.g., Vps26 and Vps35). In in vitro binding assays, recombinant His₆-tagged Btn2 bound glutathione S-transferase (GST)-Snc1 and GST-Vps26. Btn2-green fluorescent protein and Btn2-red fluorescent protein colocalize with Tlg2, Snx4, and Vps27 to a compartment adjacent to the vacuole that corresponds to a late endosome. The deletion of BTN2 blocks Yif1 retrieval back to the Golgi apparatus, while the localization of Ste2, Fur4, Snc1, Vps10, carboxypeptidases Y (CPY) and S (CPS), Sed5, and Sec7 is unaltered in btn2 cells. Yif1 delivery to the vacuole was observed in other late endosome-Golgi trafficking mutants, including ypt6, snx4, and vps26 cells. Thus, Btn2 facilitates specific protein retrieval from a late endosome to the Golgi apparatus, a process which may be adversely affected in patients with Batten disease.

Published ahead of print on 13 November 2006.