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PMID: 15601820 Published · ppublish English Comparative Study Journal Article Research Support, Non-U.S. Gov't

VHL-box and SOCS-box domains determine binding specificity for Cul2-Rbx1 and Cul5-Rbx2 modules of ubiquitin ligases.

Genes & development ·Vol. 18 ·No. 24 ·2004-12-15 ·Pages 3055-65

Kamura T, Maenaka K, Kotoshiba S, Matsumoto M, Kohda D, Conaway RC, Conaway JW, Nakayama KI

Abstract

The ECS (Elongin B/C-Cul2/Cul5-SOCS-box protein) complex is a member of a family of ubiquitin ligases that share a Cullin-Rbx module. SOCS-box proteins recruit substrates to the ECS complex and are linked to Cullin-Rbx via Elongin B/C. VHL has been implicated as a SOCS-box protein, but lacks a C-terminal sequence (downstream of the BC box) of the SOCS box. We now show that VHL specifically interacts with endogenous Cul2-Rbx1 in mammalian cells, whereas SOCS-box proteins associate with Cul5-Rbx2. We also identify LRR-1 and FEM1B as proteins that share a region of homology with VHL (the VHL box, including the BC box and downstream residues) and associate with Cul2-Rbx1. ECS complexes can thus be classified into two distinct protein assemblies, that is, those that contain a subunit with a VHL box (composed of the BC box and a downstream Cul2 box) that interacts with Cul2-Rbx1, and those that contain a subunit with a SOCS box (BC box and downstream Cul5 box) that interacts with Cul5-Rbx2. Domain-swapping analyses showed that the specificity of interaction of VHL-box and SOCS-box proteins with Cullin-Rbx modules is determined by the Cul2 and Cul5 boxes, respectively. Finally, RNAi-mediated knockdown of the Cul2-Rbx1 inhibited the VHL-mediated degradation of HIF-2alpha, whereas knockdown of Cul5-Rbx2 did not affect it. These data suggest that the functions of the Cul2-Rbx1 and Cul5-Rbx2 modules are distinct.

MeSH Terms
Amino Acid Sequence Carrier Proteins/metabolism Cell Cycle Proteins/metabolism Cells, Cultured Cullin Proteins/genetics,metabolism Gene Components Humans Immunoblotting Immunoprecipitation Intracellular Signaling Peptides and Proteins/genetics,metabolism Mass Spectrometry Models, Chemical Molecular Sequence Data Protein Binding Protein Structure, Tertiary Proteins/metabolism RNA Interference Repressor Proteins/genetics,metabolism Sequence Alignment Suppressor of Cytokine Signaling 1 Protein Suppressor of Cytokine Signaling Proteins Tumor Suppressor Proteins/genetics,metabolism Ubiquitin-Protein Ligases/genetics,metabolism Von Hippel-Lindau Tumor Suppressor Protein
Chemicals
CUL2 protein, human CUL5 protein, human Carrier Proteins Cell Cycle Proteins Cullin Proteins FEM1B protein, human Intracellular Signaling Peptides and Proteins LRR1 protein, human Proteins RBX1 protein, human Repressor Proteins SOCS1 protein, human Suppressor of Cytokine Signaling 1 Protein Suppressor of Cytokine Signaling Proteins Tumor Suppressor Proteins Ubiquitin-Protein Ligases Von Hippel-Lindau Tumor Suppressor Protein VHL protein, human
Authors & Affiliations
8 authors, click to expand affiliations / ORCID
Kamura Takumi
Department of Molecular and Cellular Biology, Medical Institute of Bioregulation, Kyushu University, Higashi-ku, Fukuoka, Fukuoka 812-8582, Japan.
Maenaka Katsumi
Kotoshiba Shuhei
Matsumoto Masaki
Kohda Daisuke
Conaway Ronald C
Conaway Joan Weliky
Nakayama Keiichi I
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Article Info
Journal
Genes & development
Abbr.
Genes Dev
ISSN
0890-9369
DOI
10.1101/gad.1252404
Published
2004-12-15
Pages
3055-65
Language
English
Region
United States
NLM ID
8711660
PMCID
PMC535916
Subset
IM
External Links
PubMed source DOI: 10.1101/gad.1252404 PMC Full Text View journal details
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