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Eukaryotic Cell, December 2006, p. 2104-2113, Vol. 5, No. 12
1535-9778/06/$08.00+0     doi:10.1128/EC.00347-05
Copyright © 2006, American Society for Microbiology. All Rights Reserved.

Skp1p Regulates Soi3p/Rav1p Association with Endosomal Membranes but Is Not Required for Vacuolar ATPase Assembly{triangledown}

E. J. Brace, Leah P. Parkinson, and Robert S. Fuller*

Department of Biological Chemistry, University of Michigan Medical Center, Ann Arbor, Michigan 48109-0606

Received 16 November 2005/ Accepted 29 September 2006

Skp1p is an essential component of SCF-type E3 ubiquitin ligase complexes and associates with these through binding to F-box proteins. Skp1p also binds F-box proteins in a number of non-SCF complexes. The Skp1p-associated yeast protein Soi3p/Rav1p (hereafter referred to as Rav1p) is a component of the RAVE complex required for regulated assembly of vacuolar ATPase (V-ATPase). Rav1p is also involved in transport of TGN proteins and endocytic cargo between early and late endosomes. To evaluate the role of Skp1p in the RAVE complex, we made use of the fact that overexpression of Rav1p is toxic because it sequesters Skp1p from essential interactions. We isolated a separation of function allele of SKP1, skp1(Asn108Tyr), that completely abrogated the Rav1p interaction but allowed Skp1p to perform other essential cellular functions. Cells containing the skp1(Asn108Tyr) allele as the sole source of Skp1p exhibited normal V-ATPase assembly and activity. However, in the skp1(Asn108Tyr) mutant strain, the membrane-associated pool of Rav1-green fluorescent protein was increased, suggesting that Skp1p is important for the release of Rav1p from endosomal membranes where it functions in V-ATPase assembly. Thus, although part of the RAVE complex, Skp1p does not appear to be involved in V-ATPase assembly but instead in the cycling of the complex off membranes. This work also provides a generalizable approach to defining the roles of interactions of Skp1p with individual F-box proteins through the isolation of special alleles of SKP1.

* Corresponding author. Mailing address: Department of Biological Chemistry, University of Michigan Medical Center, Ann Arbor, MI 48109-0606. Phone: (734) 936-9764. Fax: (734) 763-7799. E-mail: bfuller{at}umich.edu.

{triangledown} Published ahead of print on 13 October 2006.

Eukaryotic Cell, December 2006, p. 2104-2113, Vol. 5, No. 12
1535-9778/06/$08.00+0     doi:10.1128/EC.00347-05
Copyright © 2006, American Society for Microbiology. All Rights Reserved.



This article has been cited by other articles:

  • Smardon, A. M., Kane, P. M. (2007). RAVE Is Essential for the Efficient Assembly of the C Subunit with the Vacuolar H+-ATPase. J. Biol. Chem. 282: 26185-26194 [Abstract] [Full Text]  


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