Substrate-specific adapter of a DCX (DDB1-CUL4-X-box) E3 ubiquitin-protein ligase complex required for cell cycle control, DNA damage response and translesion DNA synthesis. The DCX(DTL) complex, also named CRL4(CDT2) complex, mediates the polyubiquitination and subsequent degradation of CDT1 and CDKN1A/p21(CIP1). CDT1 degradation in response to DNA damage is necessary to ensure proper cell cycle regulation of DNA replication. CDKN1A/p21(CIP1) degradation during S phase or following UV irradiation is essential to control replication licensing. Most substrates require their interaction with PCNA for their polyubiquitination: substrates interact with PCNA via their PIP-box, and those containing the 'K+4' motif in the PIP box, recruit the DCX(DTL) complex, leading to their degradation. In undamaged proliferating cells, the DCX(DTL) complex also promotes the 'Lys-164' monoubiquitination of PCNA, thereby being involved in PCNA-dependent translesion DNA synthesis.
Background References
1. Kiran S et al. HPVE6-USP46 Mediated Cdt2 Stabilization Reduces Set8 Mediated H4K20-Methylation to Induce Gene Expression Changes. Cancers (Basel). 2021 Dec
2. Wu X et al. DDB2 regulates DNA replication through PCNA-independent degradation of CDT2. Cell Biosci. 2021 Feb
Subcellular Location
Nucleus, Nucleus membrane, Cytoplasm, cytoskeleton, microtubule organizing center, centrosome, Chromosome.
Western blot analysis of CDT2 on different lysates with Rabbit anti-CDT2 antibody (HA721871) at 1/2,000 dilution.
Lane 1: HeLa cell lysate (20 µg/Lane) Lane 2: 293T cell lysate (20 µg/Lane) Lane 3: K-562 cell lysate (20 µg/Lane) Lane 4: Jurkat cell lysate (20 µg/Lane) Lane 5: Mouse spleen tissue lysate (40 µg/Lane)
Predicted band size: 79 kDa Observed band size: 79 kDa
Exposure time: 2 minutes;
4-20% SDS-PAGE gel.
Proteins were transferred to a PVDF membrane and blocked with 5% NFDM/TBST for 1 hour at room temperature. The primary antibody (HA721871) at 1/2,000 dilution was used in 5% NFDM/TBST at 4℃ overnight. Goat Anti-Rabbit IgG - HRP Secondary Antibody (HA1001) at 1/50,000 dilution was used for 1 hour at room temperature.
Flow cytometric analysis of HeLa cells labeling CDT2.
Cells were fixed and permeabilized. Then stained with the primary antibody (HA721871, 1μg/mL) (red) compared with Rabbit IgG Isotype Control (green). After incubation of the primary antibody at +4℃ for an hour, the cells were stained with a iFluor™ 488 conjugate-Goat anti-Rabbit IgG Secondary antibody (HA1121) at 1/1,000 dilution for 30 minutes at +4℃. Unlabelled sample was used as a control (cells without incubation with primary antibody; black).
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