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resource source identifier antibodies rabbit polyclonal anti cul4a  (Cell Signaling Technology Inc)


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    Cell Signaling Technology Inc resource source identifier antibodies rabbit polyclonal anti cul4a
    Figure 1. A Toolbox to Monitor Endogenous CRL4 Complexes Schematic representation of the PIKES approach: (1) 293T/17 cells were genetically engineered to carry a 3xFLAG or 3xHA on <t>Cul4A</t> or <t>Cul4B</t> or both to enable rapid and efficient (2) immunoprecipitation of endogenous CRL4 assemblages. (3) Endogenous CRL4 ligase complexes were then evaluated systematically using global and targeted proteomic approaches. First, the protein interaction network of interest was analyzed at endogenous protein levels, followed by a kinetic characterization of interactions, and the development of chemical and biochemical methods to control and preserve the composition of protein complexes. Subsequently, the cellular concentrations and complex stoichiometries were analyzed using QconCAT standards, and induced changes in complex composition were monitored quantitatively. See also Figures S1, S7, and S8.
    Resource Source Identifier Antibodies Rabbit Polyclonal Anti Cul4a, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 95/100, based on 145 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/resource source identifier antibodies rabbit polyclonal anti cul4a/product/Cell Signaling Technology Inc
    Average 95 stars, based on 145 article reviews
    resource source identifier antibodies rabbit polyclonal anti cul4a - by Bioz Stars, 2026-03
    95/100 stars

    Images

    1) Product Images from "PIKES Analysis Reveals Response to Degraders and Key Regulatory Mechanisms of the CRL4 Network."

    Article Title: PIKES Analysis Reveals Response to Degraders and Key Regulatory Mechanisms of the CRL4 Network.

    Journal: Molecular cell

    doi: 10.1016/j.molcel.2019.12.013

    Figure 1. A Toolbox to Monitor Endogenous CRL4 Complexes Schematic representation of the PIKES approach: (1) 293T/17 cells were genetically engineered to carry a 3xFLAG or 3xHA on Cul4A or Cul4B or both to enable rapid and efficient (2) immunoprecipitation of endogenous CRL4 assemblages. (3) Endogenous CRL4 ligase complexes were then evaluated systematically using global and targeted proteomic approaches. First, the protein interaction network of interest was analyzed at endogenous protein levels, followed by a kinetic characterization of interactions, and the development of chemical and biochemical methods to control and preserve the composition of protein complexes. Subsequently, the cellular concentrations and complex stoichiometries were analyzed using QconCAT standards, and induced changes in complex composition were monitored quantitatively. See also Figures S1, S7, and S8.
    Figure Legend Snippet: Figure 1. A Toolbox to Monitor Endogenous CRL4 Complexes Schematic representation of the PIKES approach: (1) 293T/17 cells were genetically engineered to carry a 3xFLAG or 3xHA on Cul4A or Cul4B or both to enable rapid and efficient (2) immunoprecipitation of endogenous CRL4 assemblages. (3) Endogenous CRL4 ligase complexes were then evaluated systematically using global and targeted proteomic approaches. First, the protein interaction network of interest was analyzed at endogenous protein levels, followed by a kinetic characterization of interactions, and the development of chemical and biochemical methods to control and preserve the composition of protein complexes. Subsequently, the cellular concentrations and complex stoichiometries were analyzed using QconCAT standards, and induced changes in complex composition were monitored quantitatively. See also Figures S1, S7, and S8.

    Techniques Used: Immunoprecipitation, Control

    Figure 3. Individual CRL4 Ligases Show Cullin-Scaffold Preferences and Differ up to 100-Fold in Absolute Abundance (A) Schematic of CRL4 QconCAT experiments to determine cellular concentrations and complex stoichiometries. (B–D) Absolute quantification of CRL4 components in whole cell lysates as well as immunoprecipitated Cul4 complexes. *Cul4 scaffolds were measured by targeting peptides common (cP) or unique (uP) to Cul4A and Cul4B. (E) Summary of cellular organization of CRL4 complexes. (F) and (G) Cellular concentrations of assembled CRL4s as well as DCAFs. (H) Correlation plot of percentage assembled (percentage of DCAF Cul4-bound versus free) and CRL4 ligase abundance (percentage of individual Cul4DCAF
    Figure Legend Snippet: Figure 3. Individual CRL4 Ligases Show Cullin-Scaffold Preferences and Differ up to 100-Fold in Absolute Abundance (A) Schematic of CRL4 QconCAT experiments to determine cellular concentrations and complex stoichiometries. (B–D) Absolute quantification of CRL4 components in whole cell lysates as well as immunoprecipitated Cul4 complexes. *Cul4 scaffolds were measured by targeting peptides common (cP) or unique (uP) to Cul4A and Cul4B. (E) Summary of cellular organization of CRL4 complexes. (F) and (G) Cellular concentrations of assembled CRL4s as well as DCAFs. (H) Correlation plot of percentage assembled (percentage of DCAF Cul4-bound versus free) and CRL4 ligase abundance (percentage of individual Cul4DCAF

    Techniques Used: Immunoprecipitation



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    resource source identifier antibodies rabbit polyclonal anti cul4a  (Cell Signaling Technology Inc)
    95
    Cell Signaling Technology Inc resource source identifier antibodies rabbit polyclonal anti cul4a
    Figure 1. A Toolbox to Monitor Endogenous CRL4 Complexes Schematic representation of the PIKES approach: (1) 293T/17 cells were genetically engineered to carry a 3xFLAG or 3xHA on <t>Cul4A</t> or <t>Cul4B</t> or both to enable rapid and efficient (2) immunoprecipitation of endogenous CRL4 assemblages. (3) Endogenous CRL4 ligase complexes were then evaluated systematically using global and targeted proteomic approaches. First, the protein interaction network of interest was analyzed at endogenous protein levels, followed by a kinetic characterization of interactions, and the development of chemical and biochemical methods to control and preserve the composition of protein complexes. Subsequently, the cellular concentrations and complex stoichiometries were analyzed using QconCAT standards, and induced changes in complex composition were monitored quantitatively. See also Figures S1, S7, and S8.
    Resource Source Identifier Antibodies Rabbit Polyclonal Anti Cul4a, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 95/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/resource source identifier antibodies rabbit polyclonal anti cul4a/product/Cell Signaling Technology Inc
    Average 95 stars, based on 1 article reviews
    resource source identifier antibodies rabbit polyclonal anti cul4a - by Bioz Stars, 2026-03
    95/100 stars
    		  Buy from Supplier
    resource source identifier antibodies rabbit polyclonal anti cul4a  (Bethyl)
    93
    Bethyl resource source identifier antibodies rabbit polyclonal anti cul4a
    Figure 1. A Toolbox to Monitor Endogenous CRL4 Complexes Schematic representation of the PIKES approach: (1) 293T/17 cells were genetically engineered to carry a 3xFLAG or 3xHA on <t>Cul4A</t> or <t>Cul4B</t> or both to enable rapid and efficient (2) immunoprecipitation of endogenous CRL4 assemblages. (3) Endogenous CRL4 ligase complexes were then evaluated systematically using global and targeted proteomic approaches. First, the protein interaction network of interest was analyzed at endogenous protein levels, followed by a kinetic characterization of interactions, and the development of chemical and biochemical methods to control and preserve the composition of protein complexes. Subsequently, the cellular concentrations and complex stoichiometries were analyzed using QconCAT standards, and induced changes in complex composition were monitored quantitatively. See also Figures S1, S7, and S8.
    Resource Source Identifier Antibodies Rabbit Polyclonal Anti Cul4a, supplied by Bethyl, used in various techniques. Bioz Stars score: 93/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/resource source identifier antibodies rabbit polyclonal anti cul4a/product/Bethyl
    Average 93 stars, based on 1 article reviews
    resource source identifier antibodies rabbit polyclonal anti cul4a - by Bioz Stars, 2026-03
    93/100 stars
    		  Buy from Supplier

    Image Search Results


    Figure 1. A Toolbox to Monitor Endogenous CRL4 Complexes Schematic representation of the PIKES approach: (1) 293T/17 cells were genetically engineered to carry a 3xFLAG or 3xHA on Cul4A or Cul4B or both to enable rapid and efficient (2) immunoprecipitation of endogenous CRL4 assemblages. (3) Endogenous CRL4 ligase complexes were then evaluated systematically using global and targeted proteomic approaches. First, the protein interaction network of interest was analyzed at endogenous protein levels, followed by a kinetic characterization of interactions, and the development of chemical and biochemical methods to control and preserve the composition of protein complexes. Subsequently, the cellular concentrations and complex stoichiometries were analyzed using QconCAT standards, and induced changes in complex composition were monitored quantitatively. See also Figures S1, S7, and S8.

    Journal: Molecular cell

    Article Title: PIKES Analysis Reveals Response to Degraders and Key Regulatory Mechanisms of the CRL4 Network.

    doi: 10.1016/j.molcel.2019.12.013

    Figure Lengend Snippet: Figure 1. A Toolbox to Monitor Endogenous CRL4 Complexes Schematic representation of the PIKES approach: (1) 293T/17 cells were genetically engineered to carry a 3xFLAG or 3xHA on Cul4A or Cul4B or both to enable rapid and efficient (2) immunoprecipitation of endogenous CRL4 assemblages. (3) Endogenous CRL4 ligase complexes were then evaluated systematically using global and targeted proteomic approaches. First, the protein interaction network of interest was analyzed at endogenous protein levels, followed by a kinetic characterization of interactions, and the development of chemical and biochemical methods to control and preserve the composition of protein complexes. Subsequently, the cellular concentrations and complex stoichiometries were analyzed using QconCAT standards, and induced changes in complex composition were monitored quantitatively. See also Figures S1, S7, and S8.

    Article Snippet: REAGENT or RESOURCE SOURCE IDENTIFIER Antibodies Rabbit polyclonal anti-Cul4A (epitope Cell Signaling Technology 2699S, RRID:AB_2086563 Rabbit polyclonal anti-Cul4B (epitope AA Bethyl A303-863A, RRID:AB_2620214 Rabbit polyclonal anti-Cul4B (epitope AA Sigma-Aldrich C9995, RRID:AB_1840781 Mouse monoclonal anti-Cul1 (D-5) Santa Cruz Biotechnology Sc-17775, RRID:AB_627325 Rabbit polyclonal anti-Cand1 Bethyl A302-901A, RRID:AB_10663486 Rabbit monoclonal anti-DDB2 Abcam ab181136 Rabbit monoclonal anti-DDB1 Abcam ab109027, RRID:AB_10859111 Rabbit polyclonal anti-pCHK-1 Cell Signaling Technology 2341S, RRID:AB_330023 Rabbit polyclonal anti-pH2A.x Cell Signaling Technology 2577S, RRID:AB_2118010 Rabbit polyclonal anti-CRBN Sigma-Aldrich HPA045910-100UL, RRID:AB_10960409 Rabbit polyclonal anti-CSN5 Santa Cruz Biotechnology Sc-393725 Rat monoclonal anti-HA-HRP (clone 3F10) Roche 34071100 Mouse monoclonal anti-FLAG Sigma-Aldrich A8592-2MG, RRID:AB_439702 Mouse monoclonal anti-GAPDH Santa Cruz Biotechnology sc-365062, RRID:AB_10847862 Rabbit monoclonal anti-BRD4 Abcam ab128874, RRID:AB_11145462 Rabbit monoclonal anti-CK1alpha Abcam ab206662 Rabbit polyclonal anti-ZFP91 Bethyl A303-245A, RRID:AB_10953803 Rabbit polyclonal anti-RBM39 Bethyl A300-291A, RRID:AB_263411 Mouse monoclonlal anti-b-actin Sigma-Aldrich A5316, RRID:AB_476743 Rabbit monoclonal anti-GFP Abcam ab32146, RRID:AB_732717 Rabbit TrueBlot Anti-Rabbit IgG HRP Rockland 18-8816-33, RRID:AB_2610848 Donkey-anti-Mouse-HRP secondary antibody Jackson Laboratories 715-035-151, RRID:AB_2340771 Donkey anti-Rabbit-HRP secondary antibody Jackson Laboratories 711-035-152, RRID:AB_10015282 IRDye 800CW Donkey-anti-Mouse secondary antibody Li-COR 926-32212, RRID:AB_621847 IRDye 680RD Donkey-anti-Rabbit secondary antibody Li-COR 926-68073, RRID:AB_10954442 EZview Red Anti-HA Affinity Gel Sigma-Aldrich E6779-1ML, RRID:AB_10109562 EZview Red Anti-FLAG Affinity Gel Sigma-Aldrich F2426-1ML, RRID:AB_2616449 EZview Red GSH Affinity Gel Sigma-Aldrich E6406-1ML Chemicals, Peptides, and Recombinant Proteins CRL4 SpikeMix This study, synthesized by JPT Peptide Technologies, Berlin, Germany N/A CRL4 QconCAT This study, synthesized by PolyQuant GmbH, Bad Abbach, Germany N/A LysC protease Wako 125-02543 Trypsin protease Promega V511C MLN4924 Calbiochem 5.05477.0001 CSN5i-3 Custom synthesis N/A MLN7243 MedKoo Biosciences 206163 Bortezomib Selleck Chemicals S1013 CB5083 Custom synthesis N/A Indisulam MedKoo Biosciences 201540 Lenalidomide Selleck Chemicals S1029 (Continued on next page) e1 Molecular Cell 77, 1–15.e1–e9, March 5, 2020

    Techniques: Immunoprecipitation, Control

    Figure 3. Individual CRL4 Ligases Show Cullin-Scaffold Preferences and Differ up to 100-Fold in Absolute Abundance (A) Schematic of CRL4 QconCAT experiments to determine cellular concentrations and complex stoichiometries. (B–D) Absolute quantification of CRL4 components in whole cell lysates as well as immunoprecipitated Cul4 complexes. *Cul4 scaffolds were measured by targeting peptides common (cP) or unique (uP) to Cul4A and Cul4B. (E) Summary of cellular organization of CRL4 complexes. (F) and (G) Cellular concentrations of assembled CRL4s as well as DCAFs. (H) Correlation plot of percentage assembled (percentage of DCAF Cul4-bound versus free) and CRL4 ligase abundance (percentage of individual Cul4DCAF

    Journal: Molecular cell

    Article Title: PIKES Analysis Reveals Response to Degraders and Key Regulatory Mechanisms of the CRL4 Network.

    doi: 10.1016/j.molcel.2019.12.013

    Figure Lengend Snippet: Figure 3. Individual CRL4 Ligases Show Cullin-Scaffold Preferences and Differ up to 100-Fold in Absolute Abundance (A) Schematic of CRL4 QconCAT experiments to determine cellular concentrations and complex stoichiometries. (B–D) Absolute quantification of CRL4 components in whole cell lysates as well as immunoprecipitated Cul4 complexes. *Cul4 scaffolds were measured by targeting peptides common (cP) or unique (uP) to Cul4A and Cul4B. (E) Summary of cellular organization of CRL4 complexes. (F) and (G) Cellular concentrations of assembled CRL4s as well as DCAFs. (H) Correlation plot of percentage assembled (percentage of DCAF Cul4-bound versus free) and CRL4 ligase abundance (percentage of individual Cul4DCAF

    Article Snippet: REAGENT or RESOURCE SOURCE IDENTIFIER Antibodies Rabbit polyclonal anti-Cul4A (epitope Cell Signaling Technology 2699S, RRID:AB_2086563 Rabbit polyclonal anti-Cul4B (epitope AA Bethyl A303-863A, RRID:AB_2620214 Rabbit polyclonal anti-Cul4B (epitope AA Sigma-Aldrich C9995, RRID:AB_1840781 Mouse monoclonal anti-Cul1 (D-5) Santa Cruz Biotechnology Sc-17775, RRID:AB_627325 Rabbit polyclonal anti-Cand1 Bethyl A302-901A, RRID:AB_10663486 Rabbit monoclonal anti-DDB2 Abcam ab181136 Rabbit monoclonal anti-DDB1 Abcam ab109027, RRID:AB_10859111 Rabbit polyclonal anti-pCHK-1 Cell Signaling Technology 2341S, RRID:AB_330023 Rabbit polyclonal anti-pH2A.x Cell Signaling Technology 2577S, RRID:AB_2118010 Rabbit polyclonal anti-CRBN Sigma-Aldrich HPA045910-100UL, RRID:AB_10960409 Rabbit polyclonal anti-CSN5 Santa Cruz Biotechnology Sc-393725 Rat monoclonal anti-HA-HRP (clone 3F10) Roche 34071100 Mouse monoclonal anti-FLAG Sigma-Aldrich A8592-2MG, RRID:AB_439702 Mouse monoclonal anti-GAPDH Santa Cruz Biotechnology sc-365062, RRID:AB_10847862 Rabbit monoclonal anti-BRD4 Abcam ab128874, RRID:AB_11145462 Rabbit monoclonal anti-CK1alpha Abcam ab206662 Rabbit polyclonal anti-ZFP91 Bethyl A303-245A, RRID:AB_10953803 Rabbit polyclonal anti-RBM39 Bethyl A300-291A, RRID:AB_263411 Mouse monoclonlal anti-b-actin Sigma-Aldrich A5316, RRID:AB_476743 Rabbit monoclonal anti-GFP Abcam ab32146, RRID:AB_732717 Rabbit TrueBlot Anti-Rabbit IgG HRP Rockland 18-8816-33, RRID:AB_2610848 Donkey-anti-Mouse-HRP secondary antibody Jackson Laboratories 715-035-151, RRID:AB_2340771 Donkey anti-Rabbit-HRP secondary antibody Jackson Laboratories 711-035-152, RRID:AB_10015282 IRDye 800CW Donkey-anti-Mouse secondary antibody Li-COR 926-32212, RRID:AB_621847 IRDye 680RD Donkey-anti-Rabbit secondary antibody Li-COR 926-68073, RRID:AB_10954442 EZview Red Anti-HA Affinity Gel Sigma-Aldrich E6779-1ML, RRID:AB_10109562 EZview Red Anti-FLAG Affinity Gel Sigma-Aldrich F2426-1ML, RRID:AB_2616449 EZview Red GSH Affinity Gel Sigma-Aldrich E6406-1ML Chemicals, Peptides, and Recombinant Proteins CRL4 SpikeMix This study, synthesized by JPT Peptide Technologies, Berlin, Germany N/A CRL4 QconCAT This study, synthesized by PolyQuant GmbH, Bad Abbach, Germany N/A LysC protease Wako 125-02543 Trypsin protease Promega V511C MLN4924 Calbiochem 5.05477.0001 CSN5i-3 Custom synthesis N/A MLN7243 MedKoo Biosciences 206163 Bortezomib Selleck Chemicals S1013 CB5083 Custom synthesis N/A Indisulam MedKoo Biosciences 201540 Lenalidomide Selleck Chemicals S1029 (Continued on next page) e1 Molecular Cell 77, 1–15.e1–e9, March 5, 2020

    Techniques: Immunoprecipitation

    Figure 1. A Toolbox to Monitor Endogenous CRL4 Complexes Schematic representation of the PIKES approach: (1) 293T/17 cells were genetically engineered to carry a 3xFLAG or 3xHA on Cul4A or Cul4B or both to enable rapid and efficient (2) immunoprecipitation of endogenous CRL4 assemblages. (3) Endogenous CRL4 ligase complexes were then evaluated systematically using global and targeted proteomic approaches. First, the protein interaction network of interest was analyzed at endogenous protein levels, followed by a kinetic characterization of interactions, and the development of chemical and biochemical methods to control and preserve the composition of protein complexes. Subsequently, the cellular concentrations and complex stoichiometries were analyzed using QconCAT standards, and induced changes in complex composition were monitored quantitatively. See also Figures S1, S7, and S8.

    Journal: Molecular cell

    Article Title: PIKES Analysis Reveals Response to Degraders and Key Regulatory Mechanisms of the CRL4 Network.

    doi: 10.1016/j.molcel.2019.12.013

    Figure Lengend Snippet: Figure 1. A Toolbox to Monitor Endogenous CRL4 Complexes Schematic representation of the PIKES approach: (1) 293T/17 cells were genetically engineered to carry a 3xFLAG or 3xHA on Cul4A or Cul4B or both to enable rapid and efficient (2) immunoprecipitation of endogenous CRL4 assemblages. (3) Endogenous CRL4 ligase complexes were then evaluated systematically using global and targeted proteomic approaches. First, the protein interaction network of interest was analyzed at endogenous protein levels, followed by a kinetic characterization of interactions, and the development of chemical and biochemical methods to control and preserve the composition of protein complexes. Subsequently, the cellular concentrations and complex stoichiometries were analyzed using QconCAT standards, and induced changes in complex composition were monitored quantitatively. See also Figures S1, S7, and S8.

    Article Snippet: REAGENT or RESOURCE SOURCE IDENTIFIER Antibodies Rabbit polyclonal anti-Cul4A (epitope Cell Signaling Technology 2699S, RRID:AB_2086563 Rabbit polyclonal anti-Cul4B (epitope AA Bethyl A303-863A, RRID:AB_2620214 Rabbit polyclonal anti-Cul4B (epitope AA Sigma-Aldrich C9995, RRID:AB_1840781 Mouse monoclonal anti-Cul1 (D-5) Santa Cruz Biotechnology Sc-17775, RRID:AB_627325 Rabbit polyclonal anti-Cand1 Bethyl A302-901A, RRID:AB_10663486 Rabbit monoclonal anti-DDB2 Abcam ab181136 Rabbit monoclonal anti-DDB1 Abcam ab109027, RRID:AB_10859111 Rabbit polyclonal anti-pCHK-1 Cell Signaling Technology 2341S, RRID:AB_330023 Rabbit polyclonal anti-pH2A.x Cell Signaling Technology 2577S, RRID:AB_2118010 Rabbit polyclonal anti-CRBN Sigma-Aldrich HPA045910-100UL, RRID:AB_10960409 Rabbit polyclonal anti-CSN5 Santa Cruz Biotechnology Sc-393725 Rat monoclonal anti-HA-HRP (clone 3F10) Roche 34071100 Mouse monoclonal anti-FLAG Sigma-Aldrich A8592-2MG, RRID:AB_439702 Mouse monoclonal anti-GAPDH Santa Cruz Biotechnology sc-365062, RRID:AB_10847862 Rabbit monoclonal anti-BRD4 Abcam ab128874, RRID:AB_11145462 Rabbit monoclonal anti-CK1alpha Abcam ab206662 Rabbit polyclonal anti-ZFP91 Bethyl A303-245A, RRID:AB_10953803 Rabbit polyclonal anti-RBM39 Bethyl A300-291A, RRID:AB_263411 Mouse monoclonlal anti-b-actin Sigma-Aldrich A5316, RRID:AB_476743 Rabbit monoclonal anti-GFP Abcam ab32146, RRID:AB_732717 Rabbit TrueBlot Anti-Rabbit IgG HRP Rockland 18-8816-33, RRID:AB_2610848 Donkey-anti-Mouse-HRP secondary antibody Jackson Laboratories 715-035-151, RRID:AB_2340771 Donkey anti-Rabbit-HRP secondary antibody Jackson Laboratories 711-035-152, RRID:AB_10015282 IRDye 800CW Donkey-anti-Mouse secondary antibody Li-COR 926-32212, RRID:AB_621847 IRDye 680RD Donkey-anti-Rabbit secondary antibody Li-COR 926-68073, RRID:AB_10954442 EZview Red Anti-HA Affinity Gel Sigma-Aldrich E6779-1ML, RRID:AB_10109562 EZview Red Anti-FLAG Affinity Gel Sigma-Aldrich F2426-1ML, RRID:AB_2616449 EZview Red GSH Affinity Gel Sigma-Aldrich E6406-1ML Chemicals, Peptides, and Recombinant Proteins CRL4 SpikeMix This study, synthesized by JPT Peptide Technologies, Berlin, Germany N/A CRL4 QconCAT This study, synthesized by PolyQuant GmbH, Bad Abbach, Germany N/A LysC protease Wako 125-02543 Trypsin protease Promega V511C MLN4924 Calbiochem 5.05477.0001 CSN5i-3 Custom synthesis N/A MLN7243 MedKoo Biosciences 206163 Bortezomib Selleck Chemicals S1013 CB5083 Custom synthesis N/A Indisulam MedKoo Biosciences 201540 Lenalidomide Selleck Chemicals S1029 (Continued on next page) e1 Molecular Cell 77, 1–15.e1–e9, March 5, 2020

    Techniques: Immunoprecipitation, Control

    Figure 3. Individual CRL4 Ligases Show Cullin-Scaffold Preferences and Differ up to 100-Fold in Absolute Abundance (A) Schematic of CRL4 QconCAT experiments to determine cellular concentrations and complex stoichiometries. (B–D) Absolute quantification of CRL4 components in whole cell lysates as well as immunoprecipitated Cul4 complexes. *Cul4 scaffolds were measured by targeting peptides common (cP) or unique (uP) to Cul4A and Cul4B. (E) Summary of cellular organization of CRL4 complexes. (F) and (G) Cellular concentrations of assembled CRL4s as well as DCAFs. (H) Correlation plot of percentage assembled (percentage of DCAF Cul4-bound versus free) and CRL4 ligase abundance (percentage of individual Cul4DCAF

    Journal: Molecular cell

    Article Title: PIKES Analysis Reveals Response to Degraders and Key Regulatory Mechanisms of the CRL4 Network.

    doi: 10.1016/j.molcel.2019.12.013

    Figure Lengend Snippet: Figure 3. Individual CRL4 Ligases Show Cullin-Scaffold Preferences and Differ up to 100-Fold in Absolute Abundance (A) Schematic of CRL4 QconCAT experiments to determine cellular concentrations and complex stoichiometries. (B–D) Absolute quantification of CRL4 components in whole cell lysates as well as immunoprecipitated Cul4 complexes. *Cul4 scaffolds were measured by targeting peptides common (cP) or unique (uP) to Cul4A and Cul4B. (E) Summary of cellular organization of CRL4 complexes. (F) and (G) Cellular concentrations of assembled CRL4s as well as DCAFs. (H) Correlation plot of percentage assembled (percentage of DCAF Cul4-bound versus free) and CRL4 ligase abundance (percentage of individual Cul4DCAF

    Article Snippet: REAGENT or RESOURCE SOURCE IDENTIFIER Antibodies Rabbit polyclonal anti-Cul4A (epitope Cell Signaling Technology 2699S, RRID:AB_2086563 Rabbit polyclonal anti-Cul4B (epitope AA Bethyl A303-863A, RRID:AB_2620214 Rabbit polyclonal anti-Cul4B (epitope AA Sigma-Aldrich C9995, RRID:AB_1840781 Mouse monoclonal anti-Cul1 (D-5) Santa Cruz Biotechnology Sc-17775, RRID:AB_627325 Rabbit polyclonal anti-Cand1 Bethyl A302-901A, RRID:AB_10663486 Rabbit monoclonal anti-DDB2 Abcam ab181136 Rabbit monoclonal anti-DDB1 Abcam ab109027, RRID:AB_10859111 Rabbit polyclonal anti-pCHK-1 Cell Signaling Technology 2341S, RRID:AB_330023 Rabbit polyclonal anti-pH2A.x Cell Signaling Technology 2577S, RRID:AB_2118010 Rabbit polyclonal anti-CRBN Sigma-Aldrich HPA045910-100UL, RRID:AB_10960409 Rabbit polyclonal anti-CSN5 Santa Cruz Biotechnology Sc-393725 Rat monoclonal anti-HA-HRP (clone 3F10) Roche 34071100 Mouse monoclonal anti-FLAG Sigma-Aldrich A8592-2MG, RRID:AB_439702 Mouse monoclonal anti-GAPDH Santa Cruz Biotechnology sc-365062, RRID:AB_10847862 Rabbit monoclonal anti-BRD4 Abcam ab128874, RRID:AB_11145462 Rabbit monoclonal anti-CK1alpha Abcam ab206662 Rabbit polyclonal anti-ZFP91 Bethyl A303-245A, RRID:AB_10953803 Rabbit polyclonal anti-RBM39 Bethyl A300-291A, RRID:AB_263411 Mouse monoclonlal anti-b-actin Sigma-Aldrich A5316, RRID:AB_476743 Rabbit monoclonal anti-GFP Abcam ab32146, RRID:AB_732717 Rabbit TrueBlot Anti-Rabbit IgG HRP Rockland 18-8816-33, RRID:AB_2610848 Donkey-anti-Mouse-HRP secondary antibody Jackson Laboratories 715-035-151, RRID:AB_2340771 Donkey anti-Rabbit-HRP secondary antibody Jackson Laboratories 711-035-152, RRID:AB_10015282 IRDye 800CW Donkey-anti-Mouse secondary antibody Li-COR 926-32212, RRID:AB_621847 IRDye 680RD Donkey-anti-Rabbit secondary antibody Li-COR 926-68073, RRID:AB_10954442 EZview Red Anti-HA Affinity Gel Sigma-Aldrich E6779-1ML, RRID:AB_10109562 EZview Red Anti-FLAG Affinity Gel Sigma-Aldrich F2426-1ML, RRID:AB_2616449 EZview Red GSH Affinity Gel Sigma-Aldrich E6406-1ML Chemicals, Peptides, and Recombinant Proteins CRL4 SpikeMix This study, synthesized by JPT Peptide Technologies, Berlin, Germany N/A CRL4 QconCAT This study, synthesized by PolyQuant GmbH, Bad Abbach, Germany N/A LysC protease Wako 125-02543 Trypsin protease Promega V511C MLN4924 Calbiochem 5.05477.0001 CSN5i-3 Custom synthesis N/A MLN7243 MedKoo Biosciences 206163 Bortezomib Selleck Chemicals S1013 CB5083 Custom synthesis N/A Indisulam MedKoo Biosciences 201540 Lenalidomide Selleck Chemicals S1029 (Continued on next page) e1 Molecular Cell 77, 1–15.e1–e9, March 5, 2020

    Techniques: Immunoprecipitation