Skp1 forms multiple protein complexes, including RAVE, a regulator of V-ATPase assembly

SCF ubiquitin ligases are composed of Skp1, Cdc53, Hrt1 and one member of a large family of substrate receptors known as F-box proteins (FBPs). Here we report the identification, using sequential rounds of epitope tagging, affinity purification and mass spectrometry, of 16 Skp1 and Cdc53-associated...

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Veröffentlicht in:Nature cell biology 2001-04, Vol.3 (4), p.384-391
Hauptverfasser: Shevchenko, Andrej, Deshaies, Raymond J, Seol, Jae Hong, Shevchenko, Anna
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Sprache:eng
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Zusammenfassung:SCF ubiquitin ligases are composed of Skp1, Cdc53, Hrt1 and one member of a large family of substrate receptors known as F-box proteins (FBPs). Here we report the identification, using sequential rounds of epitope tagging, affinity purification and mass spectrometry, of 16 Skp1 and Cdc53-associated proteins in budding yeast, including all components of SCF, 9 FBPs, Yjr033 (Rav1) and Ydr202 (Rav2). Rav1, Rav2 and Skp1 form a complex that we have named `regulator of the (H + )-ATPase of the vacuolar and endosomal membranes' (RAVE), which associates with the V 1 domain of the vacuolar membrane (H + )-ATPase (V-ATPase). V-ATPases are conserved throughout eukaryotes, and have been implicated in tumour metastasis and multidrug resistance, and here we show that RAVE promotes glucose-triggered assembly of the V-ATPase holoenzyme. Previous systematic genome-wide two-hybrid screens yielded 17 proteins that interact with Skp1 and Cdc53, only 3 of which overlap with those reported here. Thus, our results provide a distinct view of the interactions that link proteins into a comprehensive cellular network.
ISSN:1465-7392
1476-4679
DOI:10.1038/35070067