All tags p21 Cdk-cylin complex and its effectors

Cdk-cylin complex and its effectors

Investigate Cdk-cyclin and its associating protein targets in cell cycle and cancer with highly cited reagents.

Cdk (cyclin-dependent kinase) associates with cyclins to play a crucial role in cell cycle progression. p21 directly binds to Cdk-cyclin complexes maintaining Cdk in an inactive state1.

Inactive 
Cdk cannot phosphorylate downstream targets thereby leading to cell cycle arrest2

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Contents



​​Cdk1 (Cdc2, p34Cdc2)

Cdk1 interacts with cyclin B to form an active heterodimer3. Cdk1-cyclin B1 complex progresses the cell cycle through G2/M mitotic phase. Uncontrolled Cdk1 activation can lead to apoptosis2.

TypeReagent nameDescriptionAbID
AntibodyAnti-CDK1 antibodyType: Rabbit monoclonal antibody
Reactivity: Hu
Applications: ICC/IF, Flow Cyt, WB, IP, IHC-P
ab133327
Anti-CDK1 (phospho Y15) antibodyY15: Phosphorylated inhibitory residue of Cdk1 causing G1 block4.

Type: Rabbit polyclonal antibody
Reactivity: Hu, Mo, Rt
Applications: WB, ELISA
ab47594
Anti-CDK1 (phospho T14) antibodyT14: Phosphorylated inhibitory residue of Cdk1 causing G1 block4.

Type: Rabbit polyclonal antibody
Reactivity: Hu, Mo, Rt
Applications: WB, ELISA
ab58509
Anti-CDK1 (phospho T161) antibodyT161: Phosphorylated residue that stabilizes Cdk-cyclin B1 complex5.

Type: Rabbit polyclonal antibody
Reactivity: Hu, Mo, Rt
Applications: WB, ELISA
ab194874
KitCDK1 SimpleStep® ELISA kit

Get fast, reproducible results in 90 minutes.

Reactivity: Hu
Range: 93.75 pg/ml-6000 pg/ml
Method: Colorimetric

ab212162
Small
molecule compound
Ro-3306
(Cdk1 inhibitor)
Selective, reversible small molecule Cdk1 inhibitor (Ki = 35 nM Cdk1/cyclin B).ab141491
Bleomycin
(G1/S, G2M blocker)
Induces cell cycle exit and down-regulation of mitotic cyclins. Causes increased accumulation of cyclin B1-Cdk1-p21 complexes2.ab142977

Click here for the full list of Cdk inhibitors


Cyclin B1

Cyclin B1 levels rise during G2 phase facilitating Cdk1-cyclin B1 complex formation6. Cdk1-cyclin B1 is activated by Cdc25 phosphatase7.

TypeReagent nameDescriptionAbID
Antibody

Anti-Cyclin B1 antibodyValidated using a knockout cell line

Type: Rabbit monoclonal antibody
Reactivity: Hu
Applications: WB, IHC-P, Flow Cyt, ICC, IP
ab32053
Anti-Cyclin B1 (p S126) antibodyS126-phosphorylated cyclin B1 binds to T57-phosphorylated p21. Promotes kinase activation and G2M progression8.

Type: Rabbit monoclonal antibody
Reactivity: Hu
​Applications: IP, WB
ab133439
Small molecule compoundPaclitaxel (Taxol)Cyclin B1 suppression increases taxol sensitivity and inhibits tumor growth9.ab120143
Leptomycin BInduces endogenous nuclear cyclin B1 accumulation and prevents nuclear export10.ab120501

Click here for conjugated Cdk antibodies


Cdk 2 (CDKN2, p33)

Binding of p21 to Cdk2-cyclin E complex arrests cells at the G1/S checkpoint. Cdk2 inhibition by p21 is therefore crucial for DNA damage repair11.

TypeReagent nameDescriptionAbID
AntibodyAnti-Cdk2 antibodyValidated using a knockout cell lineType: Rabbit monoclonal antibody
Reactivity: Hu, Mo, Rt
Applications: ICC/IF, IP, WB, Flow Cyt, IHC-P
ab32147
Anti-CDK2 (phospho Y15) antibodyY15: Inhibitory phosphorylation site by Wee1 kinase12.

Type: Rabbit monoclonal antibody
Reactivity: Hu, Mo, Rt
Applications: IHC-P, WB, IP
ab76146
Anti-CDK2 (phospho T14) antibodyT14: Inhibitory phosphorylation site by Wee1 kinase12.

Type: Rabbit monoclonal antibody
Reactivity: Hu, Rt, ChHa
Applications: WB, IP, IHC-P, ICC
ab68265
Anti-CDK2 (phospho T160) antibodyT160: Phosphorylation that stabilizes the Cdk2-cyclin complex12.

Type: Rabbit monoclonal antibody
Reactivity: Hu, Mo, Rt
​Applications: IHC-P, WB
ab194868
Small molecule compoundFlavopiridol
(Cdk2 inhibitor)
Potent inhibitor Cdk2 kinase activity (IC50 = 100 nM at 400 μM ATP)13.ab141300
Roscovitine
(Cdk2 inhibitor)
Potent and selective inhibitor of Cdk2, Cdk5, Cdc2. Exhibits IC50 = 0.7 μM for Cdk2/cyclin A, Cdk2/cyclin E14.ab141847


Cyclin E1 

Cdk2-cyclin E1 complex facilitates phosphorylation of retinoblastoma protein (Rb), E2F transcription factor release and DNA S phase entry15

TypeReagent nameDescriptionAbID
AntibodyAnti-Cyclin E1 antibodyType: Rabbit monoclonal antibody
Reactivity: Hu, Mo, Rt
Applications: IHC-P, WB
ab33911
Small molecule compoundSorafenib
(kinase inhibitor)
Cyclin E1 expression is not suppressed in sorafenib-resistant cancer16.ab141966


Cyclin A1

p53 activates cyclin A1 promoter. Cdk2-cyclin A1 complex functions in DNA double strand break repair17.

TypeReagent nameDescriptionAbID
AntibodyAnti-Cyclin A1/A2 antibodyType: Rabbit monoclonal antibody
Reactivity: Hu
Applications: WB, IHC-P, IP
ab185619
Anti-Ku70 antibodyKu70 is a Cdk2-cyclin A1 interactor that plays a role in DNA repair17.

Type: Rabbit monoclonal antibody
Reactivity: Hu
Applications: WB, IHC-P, ICC/IF, Flow Cyt, IP
ab92450


Cdc25c 

Cdc25 dephosphorylates Y15 and T14 on Cdk2, the rate-limiting step in Cdk activation/cell-cycle progression18.

TypeReagent nameDescriptionAbID
AntibodyAnti-Cdc25C antibodyValidated using a knockout cell lineType: Rabbit monoclonal antibody
Reactivity: Hu
Applications: WB, IHC-P, ICC/IF, Flow Cyt, IP
ab32444
Small molecule compoundNSC 663284
(Cdc25 inhibitor)
Potent and selective inhibitor of Cdc25 (Ki = 89 nM for Cdc25C)19.ab144408
Gossypin
(Cdc25 activator)
Induces G2/M arrest and inhibits cell proliferation via Chk1 phosphorylation of Cdc25C20.ab145623
LysateCdc25C lysateCdc25C overexpression 293T lysate suitable for WB as a control.

Concentration: 100 µl at 1.7 mg/ml (1x sample buffer)
ab94219


Cdc6

Protein expression of Cdc6 increases during G1 phase. Cdc6 regulates initiation of DNA replication and its levels are stabilized by Cdk2-cyclin E activity21.

TypeReagent nameDescriptionAbID
AntibodyAnti-Cdc6 antibodyType: Rabbit monoclonal antibody
Reactivity: Hu
Applications: WB, IHC-P, ICC
ab109315
Anti-Cdc6 (phospho S54) antibodyPhosphorylation on S54 prevents Cdc6 from APC/C-dependent proteolysis22.

Type: Rabbit monoclonal antibody
Reactivity: Hu
Applications: WB, ICC/IF
ab75809
Anti-Cdc6 (phospho S106) antibodyCdk2-Cyclin E phosphorylation site required to induce G2 arrest23.

Type: Rabbit monoclonal antibody
Reactivity: Hu
Applications: WB
ab76422
Anti-CRM1 antibody (Nuclear export factor)CRM1 regulates Cyclin B1 nuclear export and localization10.

Type: Rabbit monoclonal antibody
Reactivity: Hu, Mo, Rt
​Applications: IHC-P, Flow Cyt, ICC/IF, WB
ab191081
Small molecule compoundMG-132
(proteasome inhibitor)
MG-132 leads to rapid accumulation of Cdc6. Effects of MG-132 are reversible with cyclohexamide (ab120093). Useful for Cdc6 degradation and localization studies24.ab141003


Cdk4/6 and cyclin D1/2

Required for efficient cell proliferation and plays a role in tumorigenesis25. Kinase activity of Cdk4-cyclin D complex is inhibited by increase in p21 levels26. Cyclin D2 acts as a Cdk2 binding partner in absence of Cdk4 and Cdk627.

TypeReagent nameDescriptionAbID
AntibodyAnti-Cdk4 antibodyValidated using a knockout cell lineType: Rabbit monoclonal antibody
Reactivity: Hu
Applications: WB, IHC-P, Flow Cyt, ICC/IF
ab108357

Anti-Cdk6 antibodyValidated using a knockout cell line

Type: Rabbit monoclonal antibody
Reactivity: Hu
Applications: Flow Cyt, ICC/IF, IHC-P, WB
ab124821
Anti-Cyclin D1 antibodyType: Rabbit monoclonal antibody
Reactivity: Hu, Mo, Rt
Applications: WB, IP, ICC/IF, IHC-P
ab134175
Anti-Cyclin D2 antibodyType: Rabbit monoclonal antibody
Reactivity: Hu
Applications: ICC/IF, IP, WB
ab207604
KitCyclin D1 SimpleStep ELISA® kit 

Quantitatively measure cyclin D1 protein in human cell and tissue extract samples.

Reactivity: Hu
Range: 187 pg/ml - 12000 pg/ml;
Method: Colorimetric

ab214571
Small molecule compoundPalbociclib
(Cdk4/6 inhibitor)
Highly potent Cdk4/6 inhibitor (IC50 = 11 nM and 16 nM for Cdk4/6)28.ab218118
Fascaplysin
(Cdk4 inhibitor)
Potent, selective ATP-competitive CDK4 inhibitor (IC50 = 350 nM)29.ab146177


Retinoblastoma protein (Rb)

Rb is inactivated following phosphorylated by Cdk2 and Cdk4/6. p21 inhibition of Cdk leads to dephosphorylation and activation of Rb. Rb forms a complex with E2F transcription factor and stalls cell cycle progression30. p21 can negatively regulate Rb stalling of cell cycle via a feedback loop31.

TypeReagent nameDescriptionAbID
AntibodyAnti-Rb binding protein 6 antibodyType: Mouse polyclonal antibody
Reactivity: Hu
Applications: WB, ICC/IF
ab168511
Anti-Rb binding protein 1 antibodyType: Rabbit polyclonal antibody
Reactivity: Hu
Applications: ELISA, WB
ab25913
Small molecule compoundSilibinin
(Rb activator)
Increases levels of unphosphorylated Rb (active) 4-fold with 70-90% decrease in phosphorylated Rb (inactive)32.ab142948
PD0332991
(Rb activator)
Clinically shown to activate Rb and promote tumor suppression33.ab218118
PAI-1
(Rb activator)
Plasminogen Activator Inhibitor (PAI-1) colocalizes with Rb and increases Rb activity34.ab142349


MDM2 

A negative regulator of p21. MDM2 inhibition elevates p21 while its overexpression has the opposite effect. MDM2 binds p21 and directs it to proteasome-mediated degradation35.

TypeReagent nameDescriptionAbID
AntibodyAnti-MDM2 antibodyType: Mouse monoclonal
Reactivity: Hu, Mo
Applications: Flow Cyt, ICC/IF, IHC-P, IP, WB
ab16895
ProteinRecombinant Human MDM2Protein for use as a control in WB.ab167941
Small molecule compoundNutlin-3a
(MDM2 inhibitor)
Potent p53-MDM2 binding inhibitor (IC50 = 90 nM)36. Causes activation of p21 at levels greater than etoposide (ab120227).ab144428
NSC 66811
(MDM2 inhibitor)
Potent, cell-permeable MDM2 inhibitor (Ki = 120 nM). Induces p21, p53 and MDM2 accumulation in human colon cancer cells37.ab142190


p21 cell cycle analysis

Modulating p21 and its associated Cdk target proteins can cause DNA damage, replication stress and DNA accumulation. Studying DNA changes during cell cycle arrest will help better characterize cell cycle progression and the effect of target modulation.

TypeReagent nameDescriptionAbID
KitCell Cycle Assay Kit (Fluorometric)Monitor cell cycle progression in G0/G1, S and G2/M phases using flow cytometry38.
Ex/Em = 490 nm/520 nm
ab112116
ProteinCell Cycle and Apoptosis WB CocktailControls for WB of molecular markers for the G1/S and M phases of the cell cycle (pCdk/pHH3/Actin/PARP)ab139417
Small molecule compoundBrdUCell proliferation reagents can be used for labelling DNA and studying DNA accumulation following modulation of p21 and other cell cycle proteins39. Detection antibodies for BrdU (ab6326), ldU (ab181664).ab142567
CldUab213715
IdUab142581
EdUThymidine analog and BrdU alternative that does not require antibodies or DNA denaturation40. EdU can be used to study DNA damage following p21 induction39.ab146186

Click here for the cell proliferation guide

References

  1. Harper, J. W. et al. Inhibition of cyclin-dependent kinases by p21. Mol. Biol. Cell 6, 387–400 (1995). Read more
  2. Charrier-Savournin, F. B. et al. p21-Mediated nuclear retention of cyclin B1-Cdk1 in response to genotoxic stress. Mol. Biol. Cell 15, 3965–76 (2004). Read more
  3. Castedo, M., Perfettini, J.-L., Roumier, T. & Kroemer, G. Cyclin-dependent kinase-1: linking apoptosis to cell cycle and mitotic catastrophe. Cell Death Differ. 9, 1287–1293 (2002). Read more
  4. Potapova, T. A., Daum, J. R., Byrd, K. S. & Gorbsky, G. J. Fine tuning the cell cycle: activation of the Cdk1 inhibitory phosphorylation pathway during mitotic exit. Mol. Biol. Cell 20, 1737–48 (2009). Read more
  5. Larochelle, S. et al. Requirements for Cdk7 in the Assembly of Cdk1/Cyclin B and Activation of Cdk2 Revealed by Chemical Genetics in Human Cells. Mol. Cell 25, 839–850 (2007). Read more
  6. Pines, J. & Hunter, T. Isolation of a human cyclin cDNA: evidence for cyclin mRNA and protein regulation in the cell cycle and for interaction with p34cdc2. Cell 58, 833–46 (1989). Read more
  7. Boutros, R., Dozier, C. & Ducommun, B. The when and wheres of CDC25 phosphatases. Curr. Opin. Cell Biol. 18, 185–91 (2006). Read more
  8. Dash, B. C. & El-Deiry, W. S. Phosphorylation of p21 in G2/M promotes cyclin B-Cdc2 kinase activity. Mol. Cell. Biol. 25, 3364–87 (2005). Read more
  9. Androic, I. et al. Targeting cyclin B1 inhibits proliferation and sensitizes breast cancer cells to taxol. BMC Cancer 8, 391 (2008). Read more
  10. Yang, J. et al. Control of Cyclin B1 localization through regulated binding of the nuclear export factor CRM1. Genes Dev. 12, 2131–2143 (1998). Read more
  11. O’Connor, P. M. Mammalian G1 and G2 phase checkpoints. Cancer Surv. 29, 151–82 (1997). Read more
  12. Bártová, I., Otyepka, M., Kríz, Z. & Koca, J. Activation and inhibition of cyclin-dependent kinase-2 by phosphorylation; a molecular dynamics study reveals the functional importance of the glycine-rich loop. Protein Sci. 13, 1449–57 (2004). Read more
  13. Carlson, B. A., Dubay, M. M., Sausville, E. A., Brizuela, L. & Worland, P. J. Flavopiridol induces G1 arrest with inhibition of cyclin-dependent kinase (CDK) 2 and CDK4 in human breast carcinoma cells. Cancer Res. 56, 2973–8 (1996). Read more
  14. Meijer, L. et al. Biochemical and Cellular Effects of Roscovitine, a Potent and Selective Inhibitor of the Cyclin-Dependent Kinases cdc2, cdk2 and cdk5. Eur. J. Biochem. 243, 527–536 (1997). Read more
  15. Dimova, D. K. & Dyson, N. J. The E2F transcriptional network: old acquaintances with new faces. Oncogene 24, 2810–2826 (2005). Read more
  16. Hsu, C. et al. Cyclin E1 Inhibition can Overcome Sorafenib Resistance in Hepatocellular Carcinoma Cells Through Mcl-1 Suppression. Clin. Cancer Res. 22, 2555–2564 (2016). Read more
  17. Müller-Tidow, C. et al. The cyclin A1-CDK2 complex regulates DNA double-strand break repair. Mol. Cell. Biol. 24, 8917–28 (2004). Read more
  18. Donzelli, M. & Draetta, G. F. Regulating mammalian checkpoints through Cdc25 inactivation. EMBO Rep. 4, 671–7 (2003). Read more
  19. Pu, L., Amoscato, A. A., Bier, M. E. & Lazo, J. S. Dual G1 and G2 phase inhibition by a novel, selective Cdc25 inhibitor 6-chloro-7-[corrected](2-morpholin-4-ylethylamino)-quinoline-5,8-dione. J. Biol. Chem. 277, 46877–85 (2002). Read more
  20. Shi, L. et al. Gossypin Induces G2/M Arrest in Human Malignant Glioma U251 Cells by the Activation of Chk1/Cdc25C Pathway. Cell. Mol. Neurobiol. 32, 289–296 (2012). Read more
  21. Duursma, A. M. & Agami, R. CDK-Dependent Stabilization of Cdc6: Linking Growth and Stress Signals to Activation of DNA Replication. Cell Cycle 4, 1725–1728 (2005). Read more
  22. Mailand, N. & Diffley, J. F. X. CDKs Promote DNA Replication Origin Licensing in Human Cells by Protecting Cdc6 from APC/C-Dependent Proteolysis. Cell 122, 915–926 (2005). Read more
  23. Yoshida, K. et al. CDC6 interaction with ATR regulates activation of a replication checkpoint in higher eukaryotic cells. J. Cell Sci. 123, (2010). Read more
  24. Clijsters, L. & Wolthuis, R. PIP-box-mediated degradation prohibits re-accumulation of Cdc6 during S phase. J. Cell Sci. 127, (2014). Read more
  25. Cole, A. M. et al. Cyclin D2-cyclin-dependent kinase 4/6 is required for efficient proliferation and tumorigenesis following Apc loss. Cancer Res. 70, 8149–58 (2010). Read more
  26. Kehn, K. et al. The role of cyclin D2 and p21/waf1 in human T-cell leukemia virus type 1 infected cells. Retrovirology 1, 6 (2004). Read more
  27. Baker, S. J. & Reddy, E. P. CDK4: A Key Player in the Cell Cycle, Development, and Cancer. Genes Cancer 3, 658–69 (2012). Read more
  28. Fry, D. W. et al. Specific inhibition of cyclin-dependent kinase 4/6 by PD 0332991 and associated antitumor activity in human tumor xenografts. Mol. Cancer Ther. 3, 1427–38 (2004). Read more
  29. Hamilton, G. Cytotoxic effects of fascaplysin against small cell lung cancer cell lines. Mar. Drugs 12, 1377–89 (2014). Read more
  30. Münger, K. & Howley, P. M. Human papillomavirus immortalization and transformation functions. Virus Res. 89, 213–28 (2002). Read more
  31. Broude, E. V et al. p21Waf1/Cip1/Sdi1 mediates retinoblastoma protein degradation. Oncogene 26, 6954–6958 (2007). Read more
  32. Tyagi, A., Agarwal, C. & Agarwal, R. Inhibition of retinoblastoma protein (Rb) phosphorylation at serine sites and an increase in Rb-E2F complex formation by silibinin in androgen-dependent human prostate carcinoma LNCaP cells: role in prostate cancer prevention. Mol. Cancer Ther. 1, 525–32 (2002). Read more
  33. Knudsen, E. S. & Wang, J. Y. J. Targeting the RB-pathway in cancer therapy. Clin. Cancer Res. 16, 1094–9 (2010). Read more
  34. Darnell, G. A. et al. Inhibition of retinoblastoma protein degradation by interaction with the serpin plasminogen activator inhibitor 2 via a novel consensus motif. Mol. Cell. Biol. 23, 6520–32 (2003). Read more
  35. Zhang, Z. et al. MDM2 Is a Negative Regulator of p21WAF1/CIP1, Independent of p53. J. Biol. Chem. 279, 16000–16006 (2004). Read more
  36. Vassilev, L. T. et al. In Vivo Activation of the p53 Pathway by Small-Molecule Antagonists of MDM2. Science (80-. ). 303, 844–848 (2004). Read more
  37. Lu, Y. et al. Discovery of a nanomolar inhibitor of the human murine double minute 2 (MDM2)-p53 interaction through an integrated, virtual database screening strategy. J. Med. Chem. 49, 3759–3762 (2006).Read more
  38. García-Castro, B. et al. Restoration of WNT4 inhibits cell growth in leukemia-derived cell lines. BMC Cancer 13, 557 (2013). Read more
  39. Galanos, P. et al. Chronic p53-independent p21 expression causes genomic instability by deregulating replication licensing. Nat. Cell Biol. 18, 777 (2016). Read more
  40. Zeng, C. B. et al. Evaluation of 5-ethynyl-2 ’-deoxyuridine staining as a sensitive and reliable method for studying cell proliferation in the adult nervous system. Brain Res. 1319, 21–32 (2010).Read more


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