cdc25 phosphatase inhibitor ii  (Santa Cruz Biotechnology)

Journal: International Journal of Medical Sciences

Article Title: Cell Recognition Molecule L1 Regulates Cell Surface Glycosylation to Modulate Cell Survival and Migration

doi: 10.7150/ijms.20479

Figure Lengend Snippet: Effects of inhibitors of signal transduction pathways on cell migration, cell survival and the protein expressions of FUT9 and ST6Gal1. After incubation for 1 hour of L1-CHO cells treated with or without L1Ab, inhibitors of PLCγ (U73122, 10.5 μM), CDC25 phosphatase (CDC25 inhibitor II, 1.05 μM), PI3K (LY294002, 16.5 μM), Erk (Erk inhibitor, 50 μM), JNK (JNK inhibitor, 200nM), ATM/ATR (Caffeine, 1mM) or PKA inhibitor (KT5720, 280 nM) were added into the culture medium and the cells were incubated further for 24 hours. A. After treatment with inhibitors of the signal transduction pathways, cell migration assay was performed. Cell migration was significantly inhibited in L1Ab-treated L1-CHO cells by LY294002 and Erk inhibitor. B. After treatment with inhibitors of the signal transduction pathways, cell survival was quantified by MTT assay. U73122, LY294002, JNK inhibitor, Caffeine and Erk inhibitor significantly repressed cell survival in L1-CHO cells. C. Western blot was used to detect the expression of transferases. The protein expressions of ST6Gal1 and FUT9 were significantly downregulated in L1-CHO cells treated with LY294002 and Erk inhibitor. * : p <0.05; ** : p <0.01, by Student's test comparison with the no inhibitor control.

Article Snippet: Co. Ltd., Japan), Tunicamycin (Calbiochem, CA, USA), U73122 (Calbiochem, CA, USA), Cdc25 phosphatase inhibitor II (Calbiochem, CA, USA), LY294002 (Sigma, MO, USA), Erk inhibitor (3-(2-Aminoethyl)-5-((4-ethoxyphenyl) methylene) -2,4-thiazolidinedione, HCl; Calbiochem, CA, USA) and KT5720 (Sigma, MO, USA) are available commercially.

Techniques: Transduction, Migration, Incubation, Cell Migration Assay, MTT Assay, Western Blot, Expressing

Journal: Frontiers in Genetics

Article Title: WIP1 Contributes to the Adaptation of Fanconi Anemia Cells to DNA Damage as Determined by the Regulatory Network of the Fanconi Anemia and Checkpoint Recovery Pathways

doi: 10.3389/fgene.2019.00411

Figure Lengend Snippet: Boolean functions for the nodes in the FA-CHKREC network.

Article Snippet: We tested this model-derived hypothesis in the EUFA316+EV cell line, a lymphoblast cell line derived from a FA patient with inherited mutations in FANCG , using chemical inhibitors against the components of the CHKREC nodes, namely GSK830371 (Sigma-Aldrich) for WIP1 inhibition, TC-S 7010 (Selleckchem) for inhibiting Aurora A, BI2536 (Selleckchem) for inhibiting PLK1 and CDC25 Phosphatase Inhibitor II (Santa Cruz Biotechnologies) for inhibiting CDC25. shows survival curves from the different CHKREC inhibitors tested.

Techniques:

Journal: Frontiers in Genetics

Article Title: WIP1 Contributes to the Adaptation of Fanconi Anemia Cells to DNA Damage as Determined by the Regulatory Network of the Fanconi Anemia and Checkpoint Recovery Pathways

doi: 10.3389/fgene.2019.00411

Figure Lengend Snippet: Biological meaning of the attractors obtained in the FA-CHKREC network.

Article Snippet: We tested this model-derived hypothesis in the EUFA316+EV cell line, a lymphoblast cell line derived from a FA patient with inherited mutations in FANCG , using chemical inhibitors against the components of the CHKREC nodes, namely GSK830371 (Sigma-Aldrich) for WIP1 inhibition, TC-S 7010 (Selleckchem) for inhibiting Aurora A, BI2536 (Selleckchem) for inhibiting PLK1 and CDC25 Phosphatase Inhibitor II (Santa Cruz Biotechnologies) for inhibiting CDC25. shows survival curves from the different CHKREC inhibitors tested.

Techniques: Activation Assay

Journal: Frontiers in Genetics

Article Title: WIP1 Contributes to the Adaptation of Fanconi Anemia Cells to DNA Damage as Determined by the Regulatory Network of the Fanconi Anemia and Checkpoint Recovery Pathways

doi: 10.3389/fgene.2019.00411

Figure Lengend Snippet: Inactivation of CHKREC nodes in FA mutants promotes CCA and reduces FA cell survival. (A) Double KO simulations of the FAcore and components of the CHKREC (WIP1, CDK1-AurA, PLK1, CDC25, and CycB-CDK1) showing that FA cell division will be blocked since the CycB-CDK1 node cannot be activated, driving the system to a cyclic CCA attractors. Only attractors are shown. Nodes in the simulations are grouped by color according to functional categories: DNA damage in black, DNA repair pathways in blue, Checkpoint in red and CHKREC in green. Inactive nodes are colorless, whereas active nodes are colored according to their functional category. (B) Schematics showing that upon CHKREC inhibition, the division of FA mutant cells with unrepaired DNA damage will be blocked and the cell will remain in a CCA attractor. In biological terms, cell division blockade may drive the cell to senescence or cell dead. (C) Screening of multiple CHKREC chemical inhibitors showing that the FAcore mutant cell line EUFA316+EV ( FANCG deficient) is more sensitive to CHKREC inhibition than its corrected counterpart EUFA316+G. Refer to , , to see the trajectories followed by these and other FAcore and FANCD2I double null mutants, respectively, before arriving to an attractor.

Article Snippet: We tested this model-derived hypothesis in the EUFA316+EV cell line, a lymphoblast cell line derived from a FA patient with inherited mutations in FANCG , using chemical inhibitors against the components of the CHKREC nodes, namely GSK830371 (Sigma-Aldrich) for WIP1 inhibition, TC-S 7010 (Selleckchem) for inhibiting Aurora A, BI2536 (Selleckchem) for inhibiting PLK1 and CDC25 Phosphatase Inhibitor II (Santa Cruz Biotechnologies) for inhibiting CDC25. shows survival curves from the different CHKREC inhibitors tested.

Techniques: Functional Assay, Inhibition, Mutagenesis

Journal: Scientific Reports

Article Title: PPM1D controls nucleolar formation by up-regulating phosphorylation of nucleophosmin

doi: 10.1038/srep33272

Figure Lengend Snippet: ( a ) In vitro sequential phosphorylation of NPM by CDK1-PLK1. His-NPM were expressed in E. coli and purified. 2 μg of His-NPM were incubated with 10 units of CDK1-cyclinB for 60 min at 30 °C, prior to incubation with 100 ng of PLK1 for 60 min at 30 °C. 2x sample buffer was added to stop the reaction. Samples were separated by SDS-PAGE and were subjected to Western blotting with same antibodies as in ( b ) In vivo sequential phosphorylation of NPM. MCF-7 cells were transfected with either HA-NPM(WT) or HA-NPM(T199A) for 40 h then treated with 4 μg/ml nocodazole for 16 h. Lysates were then purified with mouse monoclonal anti-HA and analysed by Western blotting with polyclonal anti-HA and the same antibodies as . ( c ) Decrease of phosphorylated-CDC25C at Ser216 by PPM1D overexpression in H1299 clones. All arrows indicate CDC25C with different modification states. Cells were lysed 2 h after exposure to 30 J/m 2 of UV radiation and analysed by Western blotting with rabbit monoclonal anti-CDC25C(pS216) and the same antibodies as . ( d ) Decrease of phosphorylated-NPM at Thr199 and Ser4 by CDC25 inhibitor in MCF-7. Cells were treated with 3 μM CDC25 inhibitor for 6 h and subsequently treated with 4 μg/ml nocodazole for 16 h. Cells were then harvested by mitotic shake off and analysed by Western blotting with mouse monoclonal anti-NPM, rabbit polyclonal anti NPM(pT199), rabbit monoclonal anti-NPM(pS4), and mouse monoclonal anti-actin.

Article Snippet: For the experiments with CDC25 inhibitor, cells were treated with 3 μM CDC25 Phosphatase Inhibitor II (NSC663284, Santa Cruz Biotechnology) for 6 h and subsequently treated with 4 μg/ml nocodazole for 16 h. Cells were then harvested by mitotic shake off and analysed by Western blotting.

Techniques: In Vitro, Phospho-proteomics, Purification, Incubation, SDS Page, Western Blot, In Vivo, Transfection, Over Expression, Clone Assay, Modification