The Eukaryote Linear Motif resource for Functional Sites in Proteins
Functional site class:
CRL4-Cdt2 binding PIP degron
Functional site description:
This PCNA interaction protein (PIP) degron motif occurs in chromatin-associated proteins and is recognised by the WD40 repeat domain of Cdt2, a DDB1/CUL4-associated factor (DCAF) of the CRL4DCAF E3 ubiquitin ligase. The PIP degron overlaps a PIP box, which is required to bind PCNA. The chromatin bound complex consisting of PCNA and the PIP degron-carrying protein is targeted for ubiquitylation by CRL4Cdt2 and thereby marked for subsequent degradation after DNA damage or during S phase. PIP degron-containing proteins are involved in a broad range of cell regulation. For example, Cdt1 plays a crucial role in replication licensing, whereas human p21Cip1 and Xenopus Xic1 are Cdk inhibitors.
ELMs with this model: DEG_CRL4_CDT2_1  DEG_CRL4_CDT2_2 
Description:
This PIP degron is recognised by Cdt2, a subunit of the CRL4Cdt2 ubiquitin ligase, in a PCNA-dependent manner. This degron overlaps either a canonical (LIG_PCNA) or a non-canonical PIP box that interacts with PCNA. The canonical PIP box contains an [NQ] residue at the first position that enhances the affinity of the PIP degron to PCNA via hydrogen bonds. The hydrophobic and aromatic residues represent the main part of the PIP box and perform hydrophobic interactions with a deep cavity and proline residues of PCNA. Both aromatic residues are highly conserved in Vertebrates' PIP degrons whereas they can be substituted by methionines in non-Vertebrates. Based on the crystal structure of the PCNA-p21Cip1 complex (1AXC), it is known that the PIP degron residues [ILMV]-T-[DEN] form an α helix. It has been shown that the threonine within this α helix is crucial for the recognition by CRL4Cdt2, however, it is suggested that this residue rather binds and enhances the affinity to PCNA than to the ubiquitin ligase. Furthermore, it is proposed that the [DEN] residue at the third helix position is required for PCNA binding as well. Additionally, 2 to 3 basic residues, directly adjacent to each other, are part of this PIP degron. It is likely that the first basic residue interacts with parts of the Cdt2 WD40 repeats whereas the downstream basic residues interact with and mask a negatively charged surface area of PCNA. Furthermore, negatively charged residues directly downstream of the PIP degron are sufficient to abolish the recognition by the ubiquitin ligase. It has been shown that the preformed complex consisting of chromatin-bound PCNA and the PIP-degron is required to induce ubiquitylation of the PIP degron-carrying protein. Therefore, it is likely that Cdt2 not only recognises parts of the PIP degron but also interacts with PCNA (Havens,2009). However, more biochemical research and crystal structure analysis are required to confirm these interactions.
Pattern: [NQ]{0,1}..[ILMV][ST][DEN][FY][FY].{2,3}[KR]{2,3}[^DE]
Pattern Probability: 1.624e-07
Present in taxon: Vertebrata
Interaction Domains:
  • PCNA_C (PF02747) Proliferating cell nuclear antigen, C-terminal domain (Stochiometry: 1 : 1)
  • WD40 (PF00400) WD domain, G-beta repeat (Stochiometry: 1 : 1)
o See 6 Instances for DEG_CRL4_CDT2_1
o Abstract
Ubiquitin-mediated proteolysis has diverse regulatory functions in eukaryotic cells (Hershko,1998). The role of ubiquitylation in regulating cell state by targeting proteins for proteasomal destruction is a major field of research. The ubiquitylation of a specific protein is performed by a ubiquitin-protein ligase (also known as E3) in an ATP-dependent manner. Ubiquitin is covalently bound to a ubiquitin-activating enzyme (E1), which transfers the ubiquitin to a ubiquitin-conjugating enzyme (E2). The E2 binds to the E3 ligase, which specifically recognises the target protein (Kamura,2003). There are two major types of E3 enzymes that ubiquitylate the substrate in different ways: HECT-type E3s first form an E3-ubiquitin thioester conjugate and then transfer the ubiquitin to the substrate. In contrast, RING-type E3s do not form this thioester bond but they coordinate zinc ions in a domain that binds the E2 (Pickart,2002; Havens,2011). The cullin ring ligase 4 (CRL4) is a RING ubiquitin ligase containing the CUL4 as a scaffold protein, which is linked to the damage-specific DNA-binding protein 1 (DDB1), an adaptor, as well as to the RING domain protein Rbx1 that binds the E2. DDB1 recruits the DDB1/CUL4-associated factor (DCAF) representing the specific substrate-recognition subunit of CRL4DCAF ubiquitin ligases.
The Proliferating Cell Nuclear Antigen (PCNA) is a homotrimer and forms a ring encircling double-stranded DNA. It slides along DNA as a clamp, which is required to ensure processivity of the DNA polymerase complex and of a broad range of other proteins involved in DNA metabolism. PCNA is loaded onto DNA during S phase or after DNA damage. This sliding clamp is highly conserved in Eukaroytes with homologues present in Bacteria, Archaea and some viruses (Warbrick,2000). Proteins bind to PCNA via the PCNA interaction protein (PIP) box (LIG_PCNA). A subset of PIP box-carrying proteins contain a PIP box overlapping degron, termed PIP degron (LIG_CRL4_Cdt2_1; LIG_CRL4_Cdt2_2). This degron is recognised by the WD40 repeat domain of the Cdc10-dependent transcript 2 (Cdt2), the target recognition subunit of the CRL4Cdt2 ubiquitin ligase. In order to ubiquitylate PIP degron-carrying proteins, the PIP box must first bind to chromatin-associated PCNA and subsequently, CRL4Cdt2 binds to the preformed complex. It has been observed that ubiquitylation of these PIP degron-carrying proteins by CRL4Cdt2 occurs during S phase or after DNA damage.
o 7 selected references:

o 9 GO-Terms:

o 6 Instances for DEG_CRL4_CDT2_1
(click table headers for sorting; Notes column: =Number of Switches, =Number of Interactions)
Protein NameGene NameStartEndSubsequenceLogic#Ev.OrganismNotes
CDT1_XENLA cdt1 4 16 MSQMRVTDFFSQSKRGTAAQ TP 6 Xenopus laevis (African clawed frog)
2 
CDN1C_HUMAN CDKN1C 270 282 LSGPLISDFFAKRKRSAPEK TP 2 Homo sapiens (Human)
CDT1_HUMAN CDT1 4 15 MEQRRVTDFFARRRPGPPRI TP 3 Homo sapiens (Human)
2 
Q91603_XENLA Xicl 172 184 EITTPITDYFPKRKKILSAK TP 6 Xenopus laevis (African clawed frog)
2 
CDN1A_HUMAN CDKN1A 145 157 RRQTSMTDFYHSKRRLIFSK TP 11 Homo sapiens (Human)
2 
SETD8_HUMAN SETD8 220 231 TQQNRKLTDFYPVRRSSRKS TP 9 Homo sapiens (Human)
2 
Please cite: The Eukaryotic Linear Motif Resource ELM: 10 Years and Counting (PMID:24214962)

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