SCCRO (DCUN1D1) is an essential component of the E3 complex for neddylation.
Covalent modification of cullins by the ubiquitin-like protein NEDD8 (neddylation) regulates protein ubiquitination by promoting the assembly of cullin-RING ligase E3 complexes. Like ubiquitination, neddylation results from an enzymatic cascade involving the sequential activity of a dedicated E1 (APPBP1/Uba3), E2 (Ubc12), and an ill-defined E3. We show that SCCRO (also known as DCUN1D1) binds to the components of the neddylation pathway (Cullin-ROC1, Ubc12, and CAND1) and augments but is not required for cullin neddylation in reactions using purified recombinant proteins. We also show that SCCRO recruits Ubc12 approximately NEDD8 to the CAND1-Cul1-ROC1 complex but that this is not sufficient to dissociate or overcome the inhibitory effects of CAND1 on cullin neddylation in purified protein assays. In contrast to findings in cellular systems where no binding is seen, we show that SCCRO and CAND1 can bind to the neddylated Cul1-ROC1 complex in assays using purified recombinant proteins. Although neddylated (not unneddylated) Cul1-ROC1 is released from CAND1 upon incubation with testis lysate from SCCRO+/+ mice, the addition of recombinant SCCRO is required to achieve the same results in lysate from SCCRO(-/-) mice. Combined, these results suggest that SCCRO is an important component of the neddylation E3 complex that functions to recruit charged E2 and is involved in the release of inhibitory effects of CAND1 on cullin-RING ligase E3 complex assembly and activity.
Kim
Alexander Y
AY
Department of Surgery, Laboratory of Epithelial Cancer Biology, Memorial Sloan-Kettering Cancer Center, New York, New York 10065, USA. ramanaty@mskcc.org
Bommeljé
Claire C
CC
Lee
Benjamin E
BE
Yonekawa
Yoshihiro
Y
Choi
Lydia
L
Morris
Luc G
LG
Huang
Guochang
G
Kaufman
Andrew
A
Ryan
Russel J H
RJ
Hao
Bing
B
Ramanathan
Y
Y
Singh
Bhuvanesh
B
D000818:Animals; D002352:Carrier Proteins / Q000737:chemistry / Q000378:metabolism; D002474:Cell-Free System; D044842:Cullin Proteins / Q000737:chemistry / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins; D051379:Mice; D018345:Mice, Knockout; D046912:Multiprotein Complexes / Q000737:chemistry / Q000378:metabolism*; D000075687:NEDD8 Protein; D015513:Oncogene Proteins / Q000737:chemistry / Q000378:metabolism*; D011499:Protein Processing, Post-Translational / Q000502:physiology*; D011506:Proteins; D011518:Proto-Oncogene Proteins; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism; D044767:Ubiquitin-Protein Ligases / Q000737:chemistry / Q000378:metabolism*; D014452:Ubiquitins / Q000737:chemistry / Q000378:metabolism
C486317:CAND1 protein, human; D002352:Carrier Proteins; C100318:Cullin 1; D044842:Cullin Proteins; C408631:DCUN1D1 protein, human; D047908:Intracellular Signaling Peptides and Proteins; D046912:Multiprotein Complexes; D000075687:NEDD8 Protein; C082860:NEDD8 protein, human; C513172:Nedd8 protein, mouse; D015513:Oncogene Proteins; D011506:Proteins; D011518:Proto-Oncogene Proteins; C479529:RBX1 protein, human; D011994:Recombinant Proteins; D014157:Transcription Factors; D014452:Ubiquitins; D044767:Ubiquitin-Protein Ligases