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Addgene inc p50 expression plasmid
$PDLIM7 promotes p65 degradation in a LIM3 domain-dependent manner. (A) PDLIM7 interacts with p65 but not with <t>p50</t> subunit of NF- κ B. 293T cells were transfected with a FLAG-tagged p65 or p50 expression plasmid along with or without PDLIM7. Whole cell extracts were immunoprecipitated with anti-c-Myc and immunoblotted with anti-FLAG. Western blots are representative of four independent experiments. (B) Effect of PDLIM7 on cytoplasmic, soluble and insoluble nuclear p65 in NIH3T3 cells transfected with plasmids encoding FLAG-tagged p65 and increasing amounts of c-Myc-tagged PDLIM7. Cells were subjected to fractionation and analyzed with the indicated antibody. Western blots are representative of three independent experiments. (C) Western blot analysis for soluble and insoluble nuclear p65 in NIH3T3 cells transfected with plasmids encoding p65 without or with PDLIM7, then left untreated or treated for 4 h with MG132 (10 μM) and analyzed as in (B) . (D) NIH3T3 cells were transfected with FLAG-p65 along with or without c-Myc-PDLIM7, left untreated or treated for 4 h with MG132 (10 μM) or bafilomycin (100 μM). Cells were subjected to fractionation and analyzed with anti-FLAG antibody. Western blots are representative of three independent experiments. (E) Ubiquitination assay for p65 in 293T cells cotransfected with plasmids encoding His-tagged ubiquitin (His-Ub) and p65, together without or with wild-type or ΔLIM3 PDLIM7 and analyzed as in . Western blots are representative of three independent experiments. (F) Effect of ΔLIM3 PDLIM7 on cytoplasmic, soluble and insoluble nuclear p65 in NIH3T3 cells transfected with plasmids encoding p65, together without or with wild-type or ΔLIM3 PDLIM7 and analyzed as in (B) . Western blots are representative of five independent experiments. (G) Luciferase activity in MEFs transfected with an ELAM-1-luc and pGL4.32-Null renilla construct with or without plasmids encoding p65 in the absence or presence of expression plasmids encoding wild-type or ΔLIM3 PDLIM7. Data are representative of three independent experiments. Shown are the mean values ± SD. ** P < 0.01.$
P50 Expression Plasmid, supplied by Addgene inc, used in various techniques. Bioz Stars score: 93/100, based on 29 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/product/p50+expression+plasmid/p50+cFlag+pcDNA3+(Plasmid+%2320018)/pmc07417631-43-1-10
Average 93 stars, based on 29 article reviews

p50 expression plasmid - by Bioz Stars, 2026-08

93/100 stars

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1) Product Images from "PDLIM7 Synergizes With PDLIM2 and p62/Sqstm1 to Inhibit Inflammatory Signaling by Promoting Degradation of the p65 Subunit of NF-κB"

Article Title: PDLIM7 Synergizes With PDLIM2 and p62/Sqstm1 to Inhibit Inflammatory Signaling by Promoting Degradation of the p65 Subunit of NF-κB

Journal: Frontiers in Immunology

doi: 10.3389/fimmu.2020.01559

$PDLIM7 promotes p65 degradation in a LIM3 domain-dependent manner. (A) PDLIM7 interacts with p65 but not with p50 subunit of NF- κ B. 293T cells were transfected with a FLAG-tagged p65 or p50 expression plasmid along with or without PDLIM7. Whole cell extracts were immunoprecipitated with anti-c-Myc and immunoblotted with anti-FLAG. Western blots are representative of four independent experiments. (B) Effect of PDLIM7 on cytoplasmic, soluble and insoluble nuclear p65 in NIH3T3 cells transfected with plasmids encoding FLAG-tagged p65 and increasing amounts of c-Myc-tagged PDLIM7. Cells were subjected to fractionation and analyzed with the indicated antibody. Western blots are representative of three independent experiments. (C) Western blot analysis for soluble and insoluble nuclear p65 in NIH3T3 cells transfected with plasmids encoding p65 without or with PDLIM7, then left untreated or treated for 4 h with MG132 (10 μM) and analyzed as in (B) . (D) NIH3T3 cells were transfected with FLAG-p65 along with or without c-Myc-PDLIM7, left untreated or treated for 4 h with MG132 (10 μM) or bafilomycin (100 μM). Cells were subjected to fractionation and analyzed with anti-FLAG antibody. Western blots are representative of three independent experiments. (E) Ubiquitination assay for p65 in 293T cells cotransfected with plasmids encoding His-tagged ubiquitin (His-Ub) and p65, together without or with wild-type or ΔLIM3 PDLIM7 and analyzed as in . Western blots are representative of three independent experiments. (F) Effect of ΔLIM3 PDLIM7 on cytoplasmic, soluble and insoluble nuclear p65 in NIH3T3 cells transfected with plasmids encoding p65, together without or with wild-type or ΔLIM3 PDLIM7 and analyzed as in (B) . Western blots are representative of five independent experiments. (G) Luciferase activity in MEFs transfected with an ELAM-1-luc and pGL4.32-Null renilla construct with or without plasmids encoding p65 in the absence or presence of expression plasmids encoding wild-type or ΔLIM3 PDLIM7. Data are representative of three independent experiments. Shown are the mean values ± SD. ** P < 0.01.$
Figure Legend Snippet: PDLIM7 promotes p65 degradation in a LIM3 domain-dependent manner. (A) PDLIM7 interacts with p65 but not with p50 subunit of NF- κ B. 293T cells were transfected with a FLAG-tagged p65 or p50 expression plasmid along with or without PDLIM7. Whole cell extracts were immunoprecipitated with anti-c-Myc and immunoblotted with anti-FLAG. Western blots are representative of four independent experiments. (B) Effect of PDLIM7 on cytoplasmic, soluble and insoluble nuclear p65 in NIH3T3 cells transfected with plasmids encoding FLAG-tagged p65 and increasing amounts of c-Myc-tagged PDLIM7. Cells were subjected to fractionation and analyzed with the indicated antibody. Western blots are representative of three independent experiments. (C) Western blot analysis for soluble and insoluble nuclear p65 in NIH3T3 cells transfected with plasmids encoding p65 without or with PDLIM7, then left untreated or treated for 4 h with MG132 (10 μM) and analyzed as in (B) . (D) NIH3T3 cells were transfected with FLAG-p65 along with or without c-Myc-PDLIM7, left untreated or treated for 4 h with MG132 (10 μM) or bafilomycin (100 μM). Cells were subjected to fractionation and analyzed with anti-FLAG antibody. Western blots are representative of three independent experiments. (E) Ubiquitination assay for p65 in 293T cells cotransfected with plasmids encoding His-tagged ubiquitin (His-Ub) and p65, together without or with wild-type or ΔLIM3 PDLIM7 and analyzed as in . Western blots are representative of three independent experiments. (F) Effect of ΔLIM3 PDLIM7 on cytoplasmic, soluble and insoluble nuclear p65 in NIH3T3 cells transfected with plasmids encoding p65, together without or with wild-type or ΔLIM3 PDLIM7 and analyzed as in (B) . Western blots are representative of five independent experiments. (G) Luciferase activity in MEFs transfected with an ELAM-1-luc and pGL4.32-Null renilla construct with or without plasmids encoding p65 in the absence or presence of expression plasmids encoding wild-type or ΔLIM3 PDLIM7. Data are representative of three independent experiments. Shown are the mean values ± SD. ** P < 0.01.

Techniques Used: Transfection, Expressing, Plasmid Preparation, Immunoprecipitation, Western Blot, Fractionation, Ubiquitin Assay, Luciferase, Activity Assay, Construct

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Dynein binds to the Kv7.4 C terminus. (A) Amino acid alignment of the end of the c-helix of Xenopus Kv7.1 (used for the modeling in B) and human Kv7.1–Kv7.5 channels. (B) Close view of the Kv7 channel c-helix using the structure of Xenopus Kv7.1 suggests that several residues (highlighted in green) in the dynein-binding motifs indeed are accessible to the intracellular environment. Those in red are not exposed to the intracellular environment and are therefore unlikely to be required for dynein recognition. (C, i–iii) Representative whole-cell voltage clamp recordings and respective I-V relations compared between Kv7.4 and Kv7.4-Q580A (i); when cotransfected with p50/dynamitin (ii); or incubation with 3 µM ciliobrevin D (iii). Statistical comparisons were made with a two-way ANOVA, followed by a Bonferroni multiple comparisons test, where P < 0.05, P < 0.01, and P < 0.001 are depicted by *, **, and ***, respectively. (C iv) Mean V 1/2 for steady-state activation was compared for each condition with a one-way ANVOA. (D) Docking simulations performed with SwissDock on Kv7.1, Kv7.4, Kv7.1-Q560A, and Kv7.4-Q580A showing ciliobrevin D binding to both mutant channels but neither WT channel. Each of the four spirals (pink, green, orange, and gold) represent an intercellular C terminus of each of the four Kv7 protein α subunits that multimerize to form a functional channel. Mean values are shown with error bars depicting the SEM.

Journal: The Journal of General Physiology

Article Title: Dynein regulates Kv7.4 channel trafficking from the cell membrane

doi: 10.1085/jgp.202012760

Figure Lengend Snippet: Dynein binds to the Kv7.4 C terminus. (A) Amino acid alignment of the end of the c-helix of Xenopus Kv7.1 (used for the modeling in B) and human Kv7.1–Kv7.5 channels. (B) Close view of the Kv7 channel c-helix using the structure of Xenopus Kv7.1 suggests that several residues (highlighted in green) in the dynein-binding motifs indeed are accessible to the intracellular environment. Those in red are not exposed to the intracellular environment and are therefore unlikely to be required for dynein recognition. (C, i–iii) Representative whole-cell voltage clamp recordings and respective I-V relations compared between Kv7.4 and Kv7.4-Q580A (i); when cotransfected with p50/dynamitin (ii); or incubation with 3 µM ciliobrevin D (iii). Statistical comparisons were made with a two-way ANOVA, followed by a Bonferroni multiple comparisons test, where P < 0.05, P < 0.01, and P < 0.001 are depicted by *, **, and ***, respectively. (C iv) Mean V 1/2 for steady-state activation was compared for each condition with a one-way ANVOA. (D) Docking simulations performed with SwissDock on Kv7.1, Kv7.4, Kv7.1-Q560A, and Kv7.4-Q580A showing ciliobrevin D binding to both mutant channels but neither WT channel. Each of the four spirals (pink, green, orange, and gold) represent an intercellular C terminus of each of the four Kv7 protein α subunits that multimerize to form a functional channel. Mean values are shown with error bars depicting the SEM.

Article Snippet: Kv7.4 and Kv7.4-Q580A were coexpressed with 1 µg p50/dynamitin (catalog: HG14449-UT; SinoBiological).

Techniques: Binding Assay, Incubation, Activation Assay, Mutagenesis, Functional Assay

Journal: Frontiers in Immunology

Article Title: PDLIM7 Synergizes With PDLIM2 and p62/Sqstm1 to Inhibit Inflammatory Signaling by Promoting Degradation of the p65 Subunit of NF-κB

doi: 10.3389/fimmu.2020.01559

Figure Lengend Snippet: PDLIM7 promotes p65 degradation in a LIM3 domain-dependent manner. (A) PDLIM7 interacts with p65 but not with p50 subunit of NF- κ B. 293T cells were transfected with a FLAG-tagged p65 or p50 expression plasmid along with or without PDLIM7. Whole cell extracts were immunoprecipitated with anti-c-Myc and immunoblotted with anti-FLAG. Western blots are representative of four independent experiments. (B) Effect of PDLIM7 on cytoplasmic, soluble and insoluble nuclear p65 in NIH3T3 cells transfected with plasmids encoding FLAG-tagged p65 and increasing amounts of c-Myc-tagged PDLIM7. Cells were subjected to fractionation and analyzed with the indicated antibody. Western blots are representative of three independent experiments. (C) Western blot analysis for soluble and insoluble nuclear p65 in NIH3T3 cells transfected with plasmids encoding p65 without or with PDLIM7, then left untreated or treated for 4 h with MG132 (10 μM) and analyzed as in (B) . (D) NIH3T3 cells were transfected with FLAG-p65 along with or without c-Myc-PDLIM7, left untreated or treated for 4 h with MG132 (10 μM) or bafilomycin (100 μM). Cells were subjected to fractionation and analyzed with anti-FLAG antibody. Western blots are representative of three independent experiments. (E) Ubiquitination assay for p65 in 293T cells cotransfected with plasmids encoding His-tagged ubiquitin (His-Ub) and p65, together without or with wild-type or ΔLIM3 PDLIM7 and analyzed as in . Western blots are representative of three independent experiments. (F) Effect of ΔLIM3 PDLIM7 on cytoplasmic, soluble and insoluble nuclear p65 in NIH3T3 cells transfected with plasmids encoding p65, together without or with wild-type or ΔLIM3 PDLIM7 and analyzed as in (B) . Western blots are representative of five independent experiments. (G) Luciferase activity in MEFs transfected with an ELAM-1-luc and pGL4.32-Null renilla construct with or without plasmids encoding p65 in the absence or presence of expression plasmids encoding wild-type or ΔLIM3 PDLIM7. Data are representative of three independent experiments. Shown are the mean values ± SD. ** P < 0.01.

Article Snippet: FLAG-tagged p50 expression plasmid (p50 cFLAG pcDNA3) was purchased from Addgene (#20018).

Techniques: Transfection, Expressing, Plasmid Preparation, Immunoprecipitation, Western Blot, Fractionation, Ubiquitin Assay, Luciferase, Activity Assay, Construct