α p chk1 ser 345  (Cell Signaling Technology Inc)


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    Cell Signaling Technology Inc α p chk1 ser 345
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    α P Chk1 Ser 345, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    1) Product Images from "TZAP overexpression induces telomere dysfunction and ALT-like activity in ATRX/DAXX-deficient cells"

    Article Title: TZAP overexpression induces telomere dysfunction and ALT-like activity in ATRX/DAXX-deficient cells

    Journal: iScience

    doi: 10.1016/j.isci.2023.106405

    Key resources table
    Figure Legend Snippet: Key resources table

    Techniques Used: Recombinant, Modification, Transfection, Electrophoresis, Blocking Assay, Imaging, Purification, Software

    α p chk1 ser 345  (Cell Signaling Technology Inc)


    Bioz Manufacturer Symbol Cell Signaling Technology Inc manufactures this product  
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    Cell Signaling Technology Inc α p chk1 ser 345
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    α P Chk1 Ser 345, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    1) Product Images from "TZAP overexpression induces telomere dysfunction and ALT-like activity in ATRX/DAXX-deficient cells"

    Article Title: TZAP overexpression induces telomere dysfunction and ALT-like activity in ATRX/DAXX-deficient cells

    Journal: iScience

    doi: 10.1016/j.isci.2023.106405

    Key resources table
    Figure Legend Snippet: Key resources table

    Techniques Used: Recombinant, Modification, Transfection, Electrophoresis, Blocking Assay, Imaging, Purification, Software

    α p chk1 ser 345  (Cell Signaling Technology Inc)


    Bioz Manufacturer Symbol Cell Signaling Technology Inc manufactures this product  
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    Cell Signaling Technology Inc α p chk1 ser 345
    Key resources table
    α P Chk1 Ser 345, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/α p chk1 ser 345/product/Cell Signaling Technology Inc
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    1) Product Images from "TZAP overexpression induces telomere dysfunction and ALT-like activity in ATRX/DAXX-deficient cells"

    Article Title: TZAP overexpression induces telomere dysfunction and ALT-like activity in ATRX/DAXX-deficient cells

    Journal: iScience

    doi: 10.1016/j.isci.2023.106405

    Key resources table
    Figure Legend Snippet: Key resources table

    Techniques Used: Recombinant, Modification, Transfection, Electrophoresis, Blocking Assay, Imaging, Purification, Software

    a p chk1 ser 345  (Cell Signaling Technology Inc)


    Bioz Manufacturer Symbol Cell Signaling Technology Inc manufactures this product  
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    Cell Signaling Technology Inc a p chk1 ser 345
    A P Chk1 Ser 345, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    p chk1 ser 345 antibodies  (Cell Signaling Technology Inc)


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    Cell Signaling Technology Inc p chk1 ser 345 antibodies
    P Chk1 Ser 345 Antibodies, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    p chk1 ser 345  (Cell Signaling Technology Inc)


    Bioz Manufacturer Symbol Cell Signaling Technology Inc manufactures this product  
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    Cell Signaling Technology Inc p chk1 ser 345
    P Chk1 Ser 345, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 96/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    rabbit ser 345 p chk1  (Cell Signaling Technology Inc)


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    Cell Signaling Technology Inc rabbit ser 345 p chk1
    Real-time PCR analysis of RRM2 ( A , D ) gene expression and number of surviving cells ( B , E ) under RRM2 siRNA knockdown for NCI-H295R and MUC-1, respectively. RRM2 gene expression upon etoposide and doxorubicin treatment for NCI-H295R and MUC-1 ( C , F ). Quantification of Western blots for <t>p-Chk1</t> ( G ), p-Chk2 ( H ), and p-H2AX ( I ) for no treatment, gemcitabine, cisplatin, and combination of both (gemcitabine and cisplatin) 24 h after treatment. Schematic illustration of the related DNA damage–repair pathway, including relevant therapeutic inhibitors ( J ). Stars represent significance vs. nontreated (*, p < 0.05; **, p < 0.01; ***, p < 0.001; ****, p < 0.0001); ns, non-significant.
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    Images

    1) Product Images from "Novel Insights into the Molecular Regulation of Ribonucleotide Reductase in Adrenocortical Carcinoma Treatment"

    Article Title: Novel Insights into the Molecular Regulation of Ribonucleotide Reductase in Adrenocortical Carcinoma Treatment

    Journal: Cancers

    doi: 10.3390/cancers13164200

    Real-time PCR analysis of RRM2 ( A , D ) gene expression and number of surviving cells ( B , E ) under RRM2 siRNA knockdown for NCI-H295R and MUC-1, respectively. RRM2 gene expression upon etoposide and doxorubicin treatment for NCI-H295R and MUC-1 ( C , F ). Quantification of Western blots for p-Chk1 ( G ), p-Chk2 ( H ), and p-H2AX ( I ) for no treatment, gemcitabine, cisplatin, and combination of both (gemcitabine and cisplatin) 24 h after treatment. Schematic illustration of the related DNA damage–repair pathway, including relevant therapeutic inhibitors ( J ). Stars represent significance vs. nontreated (*, p < 0.05; **, p < 0.01; ***, p < 0.001; ****, p < 0.0001); ns, non-significant.
    Figure Legend Snippet: Real-time PCR analysis of RRM2 ( A , D ) gene expression and number of surviving cells ( B , E ) under RRM2 siRNA knockdown for NCI-H295R and MUC-1, respectively. RRM2 gene expression upon etoposide and doxorubicin treatment for NCI-H295R and MUC-1 ( C , F ). Quantification of Western blots for p-Chk1 ( G ), p-Chk2 ( H ), and p-H2AX ( I ) for no treatment, gemcitabine, cisplatin, and combination of both (gemcitabine and cisplatin) 24 h after treatment. Schematic illustration of the related DNA damage–repair pathway, including relevant therapeutic inhibitors ( J ). Stars represent significance vs. nontreated (*, p < 0.05; **, p < 0.01; ***, p < 0.001; ****, p < 0.0001); ns, non-significant.

    Techniques Used: Real-time Polymerase Chain Reaction, Expressing, Western Blot

    phosphorylated p chk1 ser 345  (Cell Signaling Technology Inc)


    Bioz Manufacturer Symbol Cell Signaling Technology Inc manufactures this product  
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    Cell Signaling Technology Inc phosphorylated p chk1 ser 345
    Phosphorylated P Chk1 Ser 345, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 96/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    anti ser 345 checkpoint kinase 1 p chk1  (Cell Signaling Technology Inc)


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    Cell Signaling Technology Inc anti ser 345 checkpoint kinase 1 p chk1
    Anti Ser 345 Checkpoint Kinase 1 P Chk1, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 96/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    antibodies against ser 345 p chk1  (Cell Signaling Technology Inc)


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    Cell Signaling Technology Inc antibodies against ser 345 p chk1
    AICAr activates <t>Chk1,</t> and pharmacological inhibition of ATR/Chk1 prevents differentiation and cell cycle arrest. A–C , U937 cells were grown in the presence of AICAr (0.5 m m ), nucleosides, and uridine (30 and 300 μ m ) ( A ); increasing concentrations of AICAr (0.1, 0.2, or 0.5 m m ), brequinar (0.1 or 0.5 μ m ), and their combinations ( B ); or AICAr (0.2 and 0.5 m m ) and brequinar (0.5 μ m ) with caffeine (4 m m ) added 5–15 min prior to the addition of agents ( C ). Total cell lysates were isolated after 48 h and analyzed by Western blotting for the level of Ser-345–phosphorylated Chk1. Representative immunoblots from three independent experiments are shown. D–F , U937 cells were incubated with AICAr (0.2 and 0.5 m m ) and brequinar (0.5 μ m ) for 72 h. Caffeine (4 m m ) ( D ), Torin2 (100 n m ) ( E ), and VE-821 (2 μ m ) ( F ) were added 30 min before the addition of agents. Representative histograms of propidium-labeled cells are shown, and the number of viable cells and the expression of differentiation markers were determined as described under “Experimental procedures.” Results are mean ± S.E. ( error bars ) of at least three independent experiments. *, p < 0.05 compared with control ( ctrl ).
    Antibodies Against Ser 345 P Chk1, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 96/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    1) Product Images from "The ribonucleoside AICAr induces differentiation of myeloid leukemia by activating the ATR/Chk1 via pyrimidine depletion"

    Article Title: The ribonucleoside AICAr induces differentiation of myeloid leukemia by activating the ATR/Chk1 via pyrimidine depletion

    Journal: The Journal of Biological Chemistry

    doi: 10.1074/jbc.RA119.009396

    AICAr activates Chk1, and pharmacological inhibition of ATR/Chk1 prevents differentiation and cell cycle arrest. A–C , U937 cells were grown in the presence of AICAr (0.5 m m ), nucleosides, and uridine (30 and 300 μ m ) ( A ); increasing concentrations of AICAr (0.1, 0.2, or 0.5 m m ), brequinar (0.1 or 0.5 μ m ), and their combinations ( B ); or AICAr (0.2 and 0.5 m m ) and brequinar (0.5 μ m ) with caffeine (4 m m ) added 5–15 min prior to the addition of agents ( C ). Total cell lysates were isolated after 48 h and analyzed by Western blotting for the level of Ser-345–phosphorylated Chk1. Representative immunoblots from three independent experiments are shown. D–F , U937 cells were incubated with AICAr (0.2 and 0.5 m m ) and brequinar (0.5 μ m ) for 72 h. Caffeine (4 m m ) ( D ), Torin2 (100 n m ) ( E ), and VE-821 (2 μ m ) ( F ) were added 30 min before the addition of agents. Representative histograms of propidium-labeled cells are shown, and the number of viable cells and the expression of differentiation markers were determined as described under “Experimental procedures.” Results are mean ± S.E. ( error bars ) of at least three independent experiments. *, p < 0.05 compared with control ( ctrl ).
    Figure Legend Snippet: AICAr activates Chk1, and pharmacological inhibition of ATR/Chk1 prevents differentiation and cell cycle arrest. A–C , U937 cells were grown in the presence of AICAr (0.5 m m ), nucleosides, and uridine (30 and 300 μ m ) ( A ); increasing concentrations of AICAr (0.1, 0.2, or 0.5 m m ), brequinar (0.1 or 0.5 μ m ), and their combinations ( B ); or AICAr (0.2 and 0.5 m m ) and brequinar (0.5 μ m ) with caffeine (4 m m ) added 5–15 min prior to the addition of agents ( C ). Total cell lysates were isolated after 48 h and analyzed by Western blotting for the level of Ser-345–phosphorylated Chk1. Representative immunoblots from three independent experiments are shown. D–F , U937 cells were incubated with AICAr (0.2 and 0.5 m m ) and brequinar (0.5 μ m ) for 72 h. Caffeine (4 m m ) ( D ), Torin2 (100 n m ) ( E ), and VE-821 (2 μ m ) ( F ) were added 30 min before the addition of agents. Representative histograms of propidium-labeled cells are shown, and the number of viable cells and the expression of differentiation markers were determined as described under “Experimental procedures.” Results are mean ± S.E. ( error bars ) of at least three independent experiments. *, p < 0.05 compared with control ( ctrl ).

    Techniques Used: Inhibition, Isolation, Western Blot, Incubation, Labeling, Expressing

    Down-regulation of Chk1 reduced the effects of AICAr and brequinar on the expression of differentiation markers and S-phase arrest. U937 cells were transfected with siRNA against Chk1 (mixture of four sequences), and respective nontargeting siRNA was used as a negative control. AICAr (0.2 or 0.5 m m ) and brequinar ( Breq ) (0.5 μ m ) were added 24 h after transfection. A , total cell lysates were isolated 3 or 48 h after the addition of agents and analyzed by Western blotting for the level of Chk1. Representative immunoblots from three independent experiments are shown. B , representative histograms of propidium-labeled cells. C , the number of viable cells and the expression of differentiation markers were determined as described under “Experimental procedures.” Results are mean ± S.E. ( error bars ) of three independent experiments. *, p < 0.05 compared with control ( ctrl ).
    Figure Legend Snippet: Down-regulation of Chk1 reduced the effects of AICAr and brequinar on the expression of differentiation markers and S-phase arrest. U937 cells were transfected with siRNA against Chk1 (mixture of four sequences), and respective nontargeting siRNA was used as a negative control. AICAr (0.2 or 0.5 m m ) and brequinar ( Breq ) (0.5 μ m ) were added 24 h after transfection. A , total cell lysates were isolated 3 or 48 h after the addition of agents and analyzed by Western blotting for the level of Chk1. Representative immunoblots from three independent experiments are shown. B , representative histograms of propidium-labeled cells. C , the number of viable cells and the expression of differentiation markers were determined as described under “Experimental procedures.” Results are mean ± S.E. ( error bars ) of three independent experiments. *, p < 0.05 compared with control ( ctrl ).

    Techniques Used: Expressing, Transfection, Negative Control, Isolation, Western Blot, Labeling

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    Cell Signaling Technology Inc α p chk1 ser 345
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    α P Chk1 Ser 345, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Real-time PCR analysis of RRM2 ( A , D ) gene expression and number of surviving cells ( B , E ) under RRM2 siRNA knockdown for NCI-H295R and MUC-1, respectively. RRM2 gene expression upon etoposide and doxorubicin treatment for NCI-H295R and MUC-1 ( C , F ). Quantification of Western blots for <t>p-Chk1</t> ( G ), p-Chk2 ( H ), and p-H2AX ( I ) for no treatment, gemcitabine, cisplatin, and combination of both (gemcitabine and cisplatin) 24 h after treatment. Schematic illustration of the related DNA damage–repair pathway, including relevant therapeutic inhibitors ( J ). Stars represent significance vs. nontreated (*, p < 0.05; **, p < 0.01; ***, p < 0.001; ****, p < 0.0001); ns, non-significant.
    Rabbit Ser 345 P Chk1, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 96/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Real-time PCR analysis of RRM2 ( A , D ) gene expression and number of surviving cells ( B , E ) under RRM2 siRNA knockdown for NCI-H295R and MUC-1, respectively. RRM2 gene expression upon etoposide and doxorubicin treatment for NCI-H295R and MUC-1 ( C , F ). Quantification of Western blots for <t>p-Chk1</t> ( G ), p-Chk2 ( H ), and p-H2AX ( I ) for no treatment, gemcitabine, cisplatin, and combination of both (gemcitabine and cisplatin) 24 h after treatment. Schematic illustration of the related DNA damage–repair pathway, including relevant therapeutic inhibitors ( J ). Stars represent significance vs. nontreated (*, p < 0.05; **, p < 0.01; ***, p < 0.001; ****, p < 0.0001); ns, non-significant.
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    Real-time PCR analysis of RRM2 ( A , D ) gene expression and number of surviving cells ( B , E ) under RRM2 siRNA knockdown for NCI-H295R and MUC-1, respectively. RRM2 gene expression upon etoposide and doxorubicin treatment for NCI-H295R and MUC-1 ( C , F ). Quantification of Western blots for <t>p-Chk1</t> ( G ), p-Chk2 ( H ), and p-H2AX ( I ) for no treatment, gemcitabine, cisplatin, and combination of both (gemcitabine and cisplatin) 24 h after treatment. Schematic illustration of the related DNA damage–repair pathway, including relevant therapeutic inhibitors ( J ). Stars represent significance vs. nontreated (*, p < 0.05; **, p < 0.01; ***, p < 0.001; ****, p < 0.0001); ns, non-significant.
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    Cell Signaling Technology Inc antibodies against ser 345 p chk1
    AICAr activates <t>Chk1,</t> and pharmacological inhibition of ATR/Chk1 prevents differentiation and cell cycle arrest. A–C , U937 cells were grown in the presence of AICAr (0.5 m m ), nucleosides, and uridine (30 and 300 μ m ) ( A ); increasing concentrations of AICAr (0.1, 0.2, or 0.5 m m ), brequinar (0.1 or 0.5 μ m ), and their combinations ( B ); or AICAr (0.2 and 0.5 m m ) and brequinar (0.5 μ m ) with caffeine (4 m m ) added 5–15 min prior to the addition of agents ( C ). Total cell lysates were isolated after 48 h and analyzed by Western blotting for the level of Ser-345–phosphorylated Chk1. Representative immunoblots from three independent experiments are shown. D–F , U937 cells were incubated with AICAr (0.2 and 0.5 m m ) and brequinar (0.5 μ m ) for 72 h. Caffeine (4 m m ) ( D ), Torin2 (100 n m ) ( E ), and VE-821 (2 μ m ) ( F ) were added 30 min before the addition of agents. Representative histograms of propidium-labeled cells are shown, and the number of viable cells and the expression of differentiation markers were determined as described under “Experimental procedures.” Results are mean ± S.E. ( error bars ) of at least three independent experiments. *, p < 0.05 compared with control ( ctrl ).
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    Key resources table

    Journal: iScience

    Article Title: TZAP overexpression induces telomere dysfunction and ALT-like activity in ATRX/DAXX-deficient cells

    doi: 10.1016/j.isci.2023.106405

    Figure Lengend Snippet: Key resources table

    Article Snippet: 8 hours later, the media of the cells was exchanged for antibiotic-containing media and cells were either transfected again or harvested for experiments 72 hours later. . Antibody sources α-GAPDH (# PAB17013 , Abnova), α-γH2A.x Ser 139 (JBW301, #05-636, Millipore), α-Myc-tag (9B11, #2276S, Cell Signaling), α-p-Chk1 Ser 345 (133D3, #2348S, Cell Signaling), α-PML (#sc-5621, Santa Cruz) α-p-Chk2 Thr 68 (#2661S, Cell Signaling), α-p-ATR Thr 1989 (#ab223258, Abcam), α-p-ATM Ser 1981 (D25E5, #13050S, Cell Signaling), α-ATRX (#sc-15408, Santa Cruz), α-DAXX (#sc-7152, Santa Cruz), α-H3.3 (#ab176840, Abcam), α-RMI1 (#NB100-1720, Novus), α-FLAG-tag (M2, #F1804-1MG, Sigma), α-RPA32 (9H8, #ab2175, Abcam), α-RPA70 (#sc-28304, Santa Cruz). α-BLM (R7019), α-TRF1 (R6839) and α-TRF2 (R6841) were raised in our lab. All antibodies were used for Western Blotting apart from α-BLM, α-Myc and α-FLAG that were also used for IF and α-RPA32, α-RPA70, α-PML, α-TRF1 and α-TRF2 that were used for IF only.

    Techniques: Recombinant, Modification, Transfection, Electrophoresis, Blocking Assay, Imaging, Purification, Software

    Real-time PCR analysis of RRM2 ( A , D ) gene expression and number of surviving cells ( B , E ) under RRM2 siRNA knockdown for NCI-H295R and MUC-1, respectively. RRM2 gene expression upon etoposide and doxorubicin treatment for NCI-H295R and MUC-1 ( C , F ). Quantification of Western blots for p-Chk1 ( G ), p-Chk2 ( H ), and p-H2AX ( I ) for no treatment, gemcitabine, cisplatin, and combination of both (gemcitabine and cisplatin) 24 h after treatment. Schematic illustration of the related DNA damage–repair pathway, including relevant therapeutic inhibitors ( J ). Stars represent significance vs. nontreated (*, p < 0.05; **, p < 0.01; ***, p < 0.001; ****, p < 0.0001); ns, non-significant.

    Journal: Cancers

    Article Title: Novel Insights into the Molecular Regulation of Ribonucleotide Reductase in Adrenocortical Carcinoma Treatment

    doi: 10.3390/cancers13164200

    Figure Lengend Snippet: Real-time PCR analysis of RRM2 ( A , D ) gene expression and number of surviving cells ( B , E ) under RRM2 siRNA knockdown for NCI-H295R and MUC-1, respectively. RRM2 gene expression upon etoposide and doxorubicin treatment for NCI-H295R and MUC-1 ( C , F ). Quantification of Western blots for p-Chk1 ( G ), p-Chk2 ( H ), and p-H2AX ( I ) for no treatment, gemcitabine, cisplatin, and combination of both (gemcitabine and cisplatin) 24 h after treatment. Schematic illustration of the related DNA damage–repair pathway, including relevant therapeutic inhibitors ( J ). Stars represent significance vs. nontreated (*, p < 0.05; **, p < 0.01; ***, p < 0.001; ****, p < 0.0001); ns, non-significant.

    Article Snippet: Antibodies used: RRM2 (#ab57653, Abcam), rabbit Ser 345 p-Chk1 (#2348, Cell Signaling, Danvers, MA, USA), rabbit Thr68 p-Chk2 (#2661, Cell Signaling), p-H2AX (#9718, Cell Signaling), and beta- actin (#A5316, Sigma-Aldrich).

    Techniques: Real-time Polymerase Chain Reaction, Expressing, Western Blot

    AICAr activates Chk1, and pharmacological inhibition of ATR/Chk1 prevents differentiation and cell cycle arrest. A–C , U937 cells were grown in the presence of AICAr (0.5 m m ), nucleosides, and uridine (30 and 300 μ m ) ( A ); increasing concentrations of AICAr (0.1, 0.2, or 0.5 m m ), brequinar (0.1 or 0.5 μ m ), and their combinations ( B ); or AICAr (0.2 and 0.5 m m ) and brequinar (0.5 μ m ) with caffeine (4 m m ) added 5–15 min prior to the addition of agents ( C ). Total cell lysates were isolated after 48 h and analyzed by Western blotting for the level of Ser-345–phosphorylated Chk1. Representative immunoblots from three independent experiments are shown. D–F , U937 cells were incubated with AICAr (0.2 and 0.5 m m ) and brequinar (0.5 μ m ) for 72 h. Caffeine (4 m m ) ( D ), Torin2 (100 n m ) ( E ), and VE-821 (2 μ m ) ( F ) were added 30 min before the addition of agents. Representative histograms of propidium-labeled cells are shown, and the number of viable cells and the expression of differentiation markers were determined as described under “Experimental procedures.” Results are mean ± S.E. ( error bars ) of at least three independent experiments. *, p < 0.05 compared with control ( ctrl ).

    Journal: The Journal of Biological Chemistry

    Article Title: The ribonucleoside AICAr induces differentiation of myeloid leukemia by activating the ATR/Chk1 via pyrimidine depletion

    doi: 10.1074/jbc.RA119.009396

    Figure Lengend Snippet: AICAr activates Chk1, and pharmacological inhibition of ATR/Chk1 prevents differentiation and cell cycle arrest. A–C , U937 cells were grown in the presence of AICAr (0.5 m m ), nucleosides, and uridine (30 and 300 μ m ) ( A ); increasing concentrations of AICAr (0.1, 0.2, or 0.5 m m ), brequinar (0.1 or 0.5 μ m ), and their combinations ( B ); or AICAr (0.2 and 0.5 m m ) and brequinar (0.5 μ m ) with caffeine (4 m m ) added 5–15 min prior to the addition of agents ( C ). Total cell lysates were isolated after 48 h and analyzed by Western blotting for the level of Ser-345–phosphorylated Chk1. Representative immunoblots from three independent experiments are shown. D–F , U937 cells were incubated with AICAr (0.2 and 0.5 m m ) and brequinar (0.5 μ m ) for 72 h. Caffeine (4 m m ) ( D ), Torin2 (100 n m ) ( E ), and VE-821 (2 μ m ) ( F ) were added 30 min before the addition of agents. Representative histograms of propidium-labeled cells are shown, and the number of viable cells and the expression of differentiation markers were determined as described under “Experimental procedures.” Results are mean ± S.E. ( error bars ) of at least three independent experiments. *, p < 0.05 compared with control ( ctrl ).

    Article Snippet: Cell lysis buffer (catalog number 9803), antibodies against Ser-345 p-Chk1 (catalog number 2348), Chk1 (catalog number 2360), anti-rabbit IgG (catalog number 7074), and anti-mouse IgG (catalog number 7076) conjugated to horseradish peroxidase were purchased from Cell Signaling Technology (Beverly, MA).

    Techniques: Inhibition, Isolation, Western Blot, Incubation, Labeling, Expressing

    Down-regulation of Chk1 reduced the effects of AICAr and brequinar on the expression of differentiation markers and S-phase arrest. U937 cells were transfected with siRNA against Chk1 (mixture of four sequences), and respective nontargeting siRNA was used as a negative control. AICAr (0.2 or 0.5 m m ) and brequinar ( Breq ) (0.5 μ m ) were added 24 h after transfection. A , total cell lysates were isolated 3 or 48 h after the addition of agents and analyzed by Western blotting for the level of Chk1. Representative immunoblots from three independent experiments are shown. B , representative histograms of propidium-labeled cells. C , the number of viable cells and the expression of differentiation markers were determined as described under “Experimental procedures.” Results are mean ± S.E. ( error bars ) of three independent experiments. *, p < 0.05 compared with control ( ctrl ).

    Journal: The Journal of Biological Chemistry

    Article Title: The ribonucleoside AICAr induces differentiation of myeloid leukemia by activating the ATR/Chk1 via pyrimidine depletion

    doi: 10.1074/jbc.RA119.009396

    Figure Lengend Snippet: Down-regulation of Chk1 reduced the effects of AICAr and brequinar on the expression of differentiation markers and S-phase arrest. U937 cells were transfected with siRNA against Chk1 (mixture of four sequences), and respective nontargeting siRNA was used as a negative control. AICAr (0.2 or 0.5 m m ) and brequinar ( Breq ) (0.5 μ m ) were added 24 h after transfection. A , total cell lysates were isolated 3 or 48 h after the addition of agents and analyzed by Western blotting for the level of Chk1. Representative immunoblots from three independent experiments are shown. B , representative histograms of propidium-labeled cells. C , the number of viable cells and the expression of differentiation markers were determined as described under “Experimental procedures.” Results are mean ± S.E. ( error bars ) of three independent experiments. *, p < 0.05 compared with control ( ctrl ).

    Article Snippet: Cell lysis buffer (catalog number 9803), antibodies against Ser-345 p-Chk1 (catalog number 2348), Chk1 (catalog number 2360), anti-rabbit IgG (catalog number 7074), and anti-mouse IgG (catalog number 7076) conjugated to horseradish peroxidase were purchased from Cell Signaling Technology (Beverly, MA).

    Techniques: Expressing, Transfection, Negative Control, Isolation, Western Blot, Labeling