TRIAD1 and HHARI bind to and are activated by distinct neddylated Cullin-RING ligase complexes.
RING (Really Interesting New Gene)-in-between-RING (RBR) enzymes are a distinct class of E3 ubiquitin ligases possessing a cluster of three zinc-binding domains that cooperate to catalyse ubiquitin transfer. The regulation and biological function for most members of the RBR ligases is not known, and all RBR E3s characterized to date are auto-inhibited for in vitro ubiquitylation. Here, we show that TRIAD1 and HHARI, two members of the Ariadne subfamily ligases, associate with distinct neddylated Cullin-RING ligase (CRL) complexes. In comparison to the modest E3 ligase activity displayed by isolated TRIAD1 or HHARI, binding of the cognate neddylated CRL to TRIAD1 or HHARI greatly stimulates RBR ligase activity in vitro, as determined by auto-ubiquitylation, their ability to stimulate dissociation of a thioester-linked UBCH7∼ubiquitin intermediate, and reactivity with ubiquitin-vinyl methyl ester. Moreover, genetic evidence shows that RBR ligase activity impacts both the levels and activities of neddylated CRLs in vivo. Cumulatively, our work proposes a conserved mechanism of CRL-induced Ariadne RBR ligase activation and further suggests a reciprocal role of this special class of RBRs as regulators of distinct CRLs.
1] Scottish Institute for Cell Signalling, College of Life Sciences, University of Dundee, Dundee, UK [2] Medical Research Council Protein Phosphorylation and Ubiquitylation Unit, College of Life Sciences, University of Dundee, Dundee, UK.
Ian R
IR
Kelsall
David M
DM
Duda
Jennifer L
JL
Olszewski
Kay
0000000222899083
K
Hofmann
Axel
A
Knebel
Frédéric
F
Langevin
Nicola
N
Wood
Melanie
M
Wightman
Brenda A
BA
Schulman
Arno F
AF
Alpi
D002352:Carrier Proteins / Q000378:metabolism*; D044842:Cullin Proteins / Q000378:metabolism; D003517:Cyclopentanes / Q000494:pharmacology; D057809:HEK293 Cells; D006801:Humans; D000075687:NEDD8 Protein; D011743:Pyrimidines / Q000494:pharmacology; D025801:Ubiquitin / Q000378:metabolism; D044763:Ubiquitin-Conjugating Enzymes / Q000378:metabolism; D044767:Ubiquitin-Protein Ligases / Q000378:metabolism*; D054875:Ubiquitination; D014452:Ubiquitins / Q000037:antagonists & inhibitors / Q000378:metabolism
C496871:CUL5 protein, human; D002352:Carrier Proteins; D044842:Cullin Proteins; D003517:Cyclopentanes; D000075687:NEDD8 Protein; C082860:NEDD8 protein, human; D011743:Pyrimidines; D025801:Ubiquitin; D014452:Ubiquitins; C079437:UBE2L3 protein, human; D044763:Ubiquitin-Conjugating Enzymes; C419193:ARIH1 protein, human; C506696:ARIH2 protein, human; D044767:Ubiquitin-Protein Ligases; C539933:pevonedistat