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  • Review Article
  • Published:

Function and regulation of cullin–RING ubiquitin ligases

Nature Reviews Molecular Cell Biology volume 6, pages 9–20 (2005)Cite this article

Key Points

  • The cullin–RING ligases (CRLs) comprise a superfamily of ubiquitin ligases that are implicated in the regulation of a diverse array of eukaryotic functions.

  • The various cullin proteins function as a rigid scaffold for the assembly of this modular class of ligase. All cullins associate with a RING protein through their C-terminal domain, whereas the N-terminal region recruits a wide variety of receptor proteins that confer substrate specificity.

  • The cullin–RING module is often referred to as the catalytic core, because it is common to all CRLs. It recruits ubiquitin-conjugating enzymes (E2s) and activates the transfer of ubiquitin from the E2 to the substrate through an as-yet-unclear mechanism that does not involve a CRL–ubiquitin-thioester intermediate.

  • The substrate receptors for CRLs are generally linked to the catalytic core through adaptor proteins that are specific for each cullin-family member. Numerous substrate receptors can be recruited to each CRL core, which increases the diversity of proteins that can be targeted for ubiquitylation.

  • In most cases, the recognition of a substrate by a CRL requires post-translational modification of the substrate. This further increases the repertoire of substrates that can be targeted to a given CRL and also links protein ubiquitylation and turnover to numerous signalling pathways.

  • CRL activity can be regulated by numerous mechanisms, which include the turnover of substrate receptors, the reversible attachment of the ubiquitin-like protein NEDD8 to cullins, and the sequestration of cullins by CAND1.

Abstract

Cullin–RING complexes comprise the largest known class of ubiquitin ligases. Owing to the great diversity of their substrate-receptor subunits, it is possible that there are hundreds of distinct cullin–RING ubiquitin ligases in eukaryotic cells, which establishes these enzymes as key mediators of post-translational protein regulation. In this review, we focus on the composition, regulation and function of cullin–RING ligases, and describe how these enzymes can be characterized by a set of general principles.

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Figure 1: The modularity of cullin–RING ligases.
Figure 2: The structure of SCFSKP2.
Figure 3: Role of cullin–RING ligases in diverse cellular processes.
Figure 4: Substrate targeting by cullin–RING ligases.
Figure 5: The regulation of cullin–RING-ligase activity through the CAND1/NEDD8 cycle.

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Acknowledgements

We thank C.-T. Chien, M. Estelle, E. Kipreos, M. Pagano and D. Wolf for their help with the online supplementary information S2 (table). We also thank G. Kleiger for preparing figure 2. M.D.P. is a postdoctoral fellow and R.J.D. is an assistant investigator of the Howard Hughes Medical Institute, which supports their work.

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Authors and Affiliations

  1. Division of Biology and Howard Hughes Medical Institute, California Institute of Technology, 1200 East California Boulevard, Pasadena, 91125, California, USA

    Matthew D. Petroski & Raymond J. Deshaies

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  1. Matthew D. Petroski
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Supplementary information

Related links

Related links

DATABASES

Flybase

Ci

Slmb

Interpro

BTB domain

F-box domain

SOCS/BC-box domain

Protein Data Bank

1LDK

Saccharomyces genome database

Cdc4

Cdc34

Grr1

Met30

S. pombe gene database

Pop1

Pop2

Swiss-Prot

APC2

CAND1

CDC34

CUL1

CUL2

CUL3

CUL4A

CUL4B

CUL5

CUL7

DDB1

elongin B

elongin C

FBH1

FBX2

FSN-1

hnRNP U

Jun

MEL-26

NEDD8

PARC

ROC1

RPM-1

SKP1

SKP2

β-TrCP1

β-TrCP2

UBC3

UBC4/5

UBC12

VHL

FURTHER INFORMATION

HGNC-Gene-Family Nomenclature — F-Box gene family

Protein families (Pfam) database

Simple Modular Architecture Research Tool (SMART)

Glossary

UBIQUITIN LIGASE (E3)

A protein or protein complex that facilitates the transfer of ubiquitin from the active-site cysteine of a ubiquitin-conjugating enzyme (E2) to a Lys residue of a substrate. Two main classes have been identified on the basis of the presence of either a HECT domain or a RING-like motif.

CULLIN

A family of proteins that is characterized by the presence of a distinct globular C-terminal domain (cullin-homology domain) and a series of N-terminal repeats of a five-helix bundle (cullin repeats).

SCF

A multisubunit ubiquitin ligase (E3). It consists of SKP1, CUL1 and an F-box protein that confers substrate specificity, as well as a RING protein that is also known as HRT1, RBX1 or ROC1.

CULLIN–RING LIGASES

(CRLs). A superfamily of ubiquitin ligases that is characterized by an enzymatic core that contains a cullin-family member and a RING protein. The core is linked to specific substrates by adaptor proteins (or domains) and various receptor subunits.

ANAPHASE-PROMOTING COMPLEX/CYCLOSOME

(APC/C). A multisubunit ubiquitin ligase that contains a RING subunit (APC11) and a distant cullin homologue (APC2). It has a key role in regulating the eukaryotic cell cycle.

SKP1

(S-phase-kinase-associated protein-1). A 23-kDa protein that functions as an adaptor between CUL1 and F-box proteins. SKP1 was identified as a protein that, together with SKP2, associates with cyclin-A–CDK complexes. It might have other functions, such as binding to centromeres and regulating the assembly of vacuolar ATPases.

RING-H2 DOMAIN

A protein motif that consists of a defined pattern of cysteine and histidine residues (Cys-X(2)-Cys-X(9–39)-Cys-X(1–3)-His-X(2–3)-His-X(2)-Cys-X(4–48)-Cys-X(2)-Cys; where X is any amino acid) and that coordinates two zinc molecules. This motif interacts directly with E2 enzymes. Proteins such as MDM2, which have variations of the basic RING motif, can nonetheless sustain ubiquitylation. In CRL RING proteins, the final Cys is an Asp.

BTB DOMAIN

A domain that was identified in 'Broad-complex, Tramtrack and Bric-a-brac' proteins in D. melanogaster and that is usually located in the N-terminal region of proteins. Originally known as the POZ domain, this motif is found in several virus proteins. Numerous BTB-domain proteins contain a second protein–protein interaction motif, such as zinc-finger and Kelch motifs.

F-BOX MOTIF

Originally identified in cyclin F, this structural motif adopts a fold that is similar to the BTB domain, binds to SKP1 and is found in receptors that assemble into CUL1-containing cullin–RING ligases. Proteins that contain the F-box motif are found in all eukaryotes.

AUTOUBIQUITYLATION

This term is loosely used to refer to the covalent transfer of ubiquitins to a Lys of an E2 or E3 component of the ubiquitin-conjugating machinery. This mechanism might regulate the assembly of some cullin–RING ligases by causing the proteasome-mediated degradation of some substrate receptors when substrate levels are depleted.

DEGRON

A portion of a protein that is necessary and sufficient to confer its degradation by the ubiquitin–proteasome system.

LYS-48-LINKED POLYUBIQUITIN CHAIN

Ubiquitin polymers in which the ε-NH2 group of Lys48 of ubiquitin is linked by an isopeptide bond to the C terminus of the next ubiquitin molecule in the chain. Such a chain can either be unanchored or attached at its proximal (C-terminal) end to the ε-NH2 group of a substrate Lys.

PROTEASOME

A 2-MDa protein complex that degrades ubiquitylated proteins in an ATP-dependent manner.

ALLOVALENCY

A kinetic model that indicates that the affinity observed for some ligand–receptor interactions might increase nonlinearly depending on the polyvalency and flexibility of the ligand. This hypothesis has been proposed for Sic1, as increasing the number of phosphate groups on Sic1 from five to six significantly increases its binding affinity for Cdc4, even though Cdc4 seems to contain only one key phosphopeptide-binding site.

HECT DOMAIN

('homologous to E6-AP C terminus' domain). HECT- and RING-domain-containing proteins represent the two main classes of E3 ubiquitin ligases. In contrast to RING ligases, HECT-domain ligases form an essential thioester intermediate with ubiquitin as it is being transferred from the E2 enzyme to the substrate.

UBC4/5 FAMILY

Two related E2-enzyme families that are structurally distinct from UBC3, the E2 that has been shown, using genetics, to interact with SCF. Although UBC4 and UBC5 ubiquitylate SCF substrates in vitro, it is unclear if they do so in vivo.

NEDD8

A small protein that is greater than 50% identical to ubiquitin and is conjugated as a single molecule to a specific Lys residue in all cullin-family members. ATP, a heterodimeric E1 (ULA1–UBA3) and the E2 UBC12 are required for the covalent attachment of NEDD8.

'HIT AND RUN' HYPOTHESIS

A mechanistic model for ubiquitin transfer by SCF ubiquitin ligases, which proposes that the dissociation of the E2 enzyme Cdc34 from SCF is required for ubiquitin transfer to the substrate.

NEDDYLATION/DENEDDYLATION

The attachment/removal of NEDD8, in this case, to/from cullin-family members. This cyclical process might regulate the assembly and activity of cullin–RING ligases. It has recently been reported that the RING ligase MDM2 is also regulated by neddylation.

COP9 SIGNALOSOME

(CSN). An eight-subunit complex that was originally identified in plants. It cleaves NEDD8 from cullins and also associates with the deubiquitylating enzyme UBP12.

'JAMM' MOTIF

A metalloprotease motif (His-X-His-X(10)-Asp) that was originally identified in the CSN5 subunit of the COP9 signalosome and the RPN11 subunit of 26S proteasome. The JAMM motif is thought to be directly involved in the cleavage of NEDD8 from cullins.

CAND1/TIP120A

(cullin-associated and neddylation-dissociated protein-1/TATA-binding-protein interacting protein-120A). A protein that specifically associates with deneddylated cullins to sequester them in an unassembled and inactive state. Putative CAND1 homologues have been identified in most eukaryotic model organisms, except for Saccharomyces cerevisiae.

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Petroski, M., Deshaies, R. Function and regulation of cullin–RING ubiquitin ligases. Nat Rev Mol Cell Biol 6, 9–20 (2005). https://doi.org/10.1038/nrm1547

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