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slug promoter reporter construct  (Addgene inc)


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    Addgene inc slug promoter reporter construct
    Slug Promoter Reporter Construct, supplied by Addgene inc, used in various techniques. Bioz Stars score: 91/100, based on 5 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/product/plasmid+constructs/Slug_pGL2+(Plasmid+%2331695)/pm42894010-562-5-16
    Average 91 stars, based on 5 article reviews
    slug promoter reporter construct - by Bioz Stars, 2026-10
    91/100 stars

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    Related Articles

    Plasmid Preparation:

    Article Title: Avian photoreceptor homologies and the origin of double cones.
    Article Snippet: .. For reporter assays, two plasmid constructs were used: pX458-Ef1a-Cas9-H2B-mCherry (Addgene #159655),89 which contains a non-targeting control gRNA under the control of theU6 promoter and aCas9-T2A-mCherry cassette under the control of theEf1alpha promoter; and Ac-OPN1LW-GFP, which contains GFP under the control of a 1.8 Kb genomic fragment from the promoter region of Anolis carolinensis OPN1LW as previously described.24 To make an electroporation solution, Plasmid DNAs were precipitated by addition of 0.1 volumes of 3 M sodium acetate and 3 volumes of 100% ethanol, washed in 70% ethanol, resuspended to a final concentration of 1 mg/ml (pX458-Ef1a-Cas9-H2B-mCherry) or 0.5 mg/ml Ac-OPN1LW-GFP in 1X PBS, and combined in a single solution. ..

    Article Title: TLE1 drives EGFR-TKI resistance in EGFR-mutant lung adenocarcinoma through epithelial to mesenchymal transition.
    Article Snippet: .. To transiently express E-cadherin or ZEB1, cells were transfected with E-cadherin-containing plasmid constructs (28009, Addgene) or ZEB1-containing plasmid (42100, Addgene) using Lipofectamine 3000 (L3000015, Invitrogen), adhering to the provider’s protocol. ..

    Article Title: Nuclear translocation of β-catenin in Wg/Wnt signaling via the IFT-A microtubule–associated complex requires Pasovec/Gid8 proteins
    Article Snippet: .. Cells were then transfected by Lipofectin (Invitrogen, A12621) with plasmid constructs [LacZ, full-length β-cat (Addgene, 14717), or Gid8 mut : 1 μg each)] together with 2 μg of pGL-TOP or its negative control vector pGL-FOP. ..

    Article Title: Differential AXL expression and Arf1 regulation control stiffness-dependent Golgi organization in breast cancer cells
    Article Snippet: .. The peGFP-N1-AXL plasmid construct was obtained from Dr. Stéphane Bodin (CRMB CNRS). pVSV-G ( Addgene #138479 ), gag/pol ( Addgene #14887 ), AXL-pBABE ( Addgene #105936 ), pBABE-puro ( Addgene #1764 ), and pMIG-w ( Addgene #12282 ) plasmid constructs were purchased from Addgene. siRNA against human AXL sequence (siAXL) was designed and purchased from Merck (Sigma). .. The siRNA sequence used was described previously ( ; ; forward, 5′-GAAAGAAGGAGACCCGTTA-3′ and reverse, 5′-TAACGGGTCTCCTTCTTTC-3′). siRNA against the human Arf1 sequence (siArf1) was designed and purchased from Merck (Sigma).

    Article Title: RhoA functionally collaborates with HSPA1A to promote the migratory phenotype of cancer cells.
    Article Snippet: RhoA, a member of the GTPase family, plays a pivotal role in attaining a migratory phenotype, mainly by regulating cytoskeleton dynamics, cell adhesion and membrane protrusions.. Although many upstream regulators and downstream effectors of RhoA have been identified, the discovery of new interacting partners continues to expand its interactome, providing fresh insights into its regulation and function.. Co-immunoprecipitation and fluorescence microscopy were used to study the interaction, localization and morphological effects of HSPA1A and RhoA.

    Article Title: The NSP5, ORF6 and NSP13 of SARS-CoV-2 Cooperate to Modulate Inflammatory Cell Death Activation.
    Article Snippet: .. Plasmid Constructs: Expression plasmid for SARS-CoV-2 spike (Addgene, 158761) was a gift fromGerald Pao,[94] and expression plasmids for SARS-CoV-2 membrane protein (Addgene, 141386) and envelope protein (Addgene, 141385) were a gift from Nevan Krogan. ..

    Article Title: Nuclear translocation of β-catenin in Wg/Wnt signaling via the IFT-A microtubule-associated complex requires Pasovec/Gid8 proteins.
    Article Snippet: .. Cells were then transfected by Lipofectin (Invitrogen, A12621) with plasmid constructs [LacZ, full- length β- cat (Addgene, 14717), or Gid8mut: 1 μ g each)] together with 2 μ g of pGL- TOP or its negative control vector pGL- FOP. ..

    Article Title: The N-terminal Segment of the Human ANKZF1 negatively regulates its internal mitochondrial targeting signal to prevent its mitochondrial localization
    Article Snippet: .. Plasmid constructs used in this study include the following: pCMV0-OCT-HA-eGFP (Addgene, #67479), pCMV0-SMAC-HA-eGFP (Addgene #67486), both the plasmids were a gift from Richard Kahn ( ). mCherry-Parkin (Addgene #23956) was a gift from Richard Youle , and Mito BFP (Addgene, #55248) ( ) was a gift from Michael Davidson. .. Δ73-ANKZF1-GFP, Δ210-ANKZF1-GFP, Δ220-ANKZF1-GFP, Δ230-ANKZF1-GFP, Δ240-ANKZF1-GFP, Δ250-ANKZF1-GFP, Δ330-ANKZF1-GFP, Δ370-ANKZF1-GFP, and Δ410-ANKZF1-GFP constructs were generated in the lab by using standard restriction digestion-based molecular cloning technique, and desired genes were digested with XhoI and KpnI restriction enzymes (NEB) and were ligated to pEGFP-N1 (Clontech) empty vector backbone.

    Construct:

    Article Title: Avian photoreceptor homologies and the origin of double cones.
    Article Snippet: .. For reporter assays, two plasmid constructs were used: pX458-Ef1a-Cas9-H2B-mCherry (Addgene #159655),89 which contains a non-targeting control gRNA under the control of theU6 promoter and aCas9-T2A-mCherry cassette under the control of theEf1alpha promoter; and Ac-OPN1LW-GFP, which contains GFP under the control of a 1.8 Kb genomic fragment from the promoter region of Anolis carolinensis OPN1LW as previously described.24 To make an electroporation solution, Plasmid DNAs were precipitated by addition of 0.1 volumes of 3 M sodium acetate and 3 volumes of 100% ethanol, washed in 70% ethanol, resuspended to a final concentration of 1 mg/ml (pX458-Ef1a-Cas9-H2B-mCherry) or 0.5 mg/ml Ac-OPN1LW-GFP in 1X PBS, and combined in a single solution. ..

    Article Title: TLE1 drives EGFR-TKI resistance in EGFR-mutant lung adenocarcinoma through epithelial to mesenchymal transition.
    Article Snippet: .. To transiently express E-cadherin or ZEB1, cells were transfected with E-cadherin-containing plasmid constructs (28009, Addgene) or ZEB1-containing plasmid (42100, Addgene) using Lipofectamine 3000 (L3000015, Invitrogen), adhering to the provider’s protocol. ..

    Article Title: Nuclear translocation of β-catenin in Wg/Wnt signaling via the IFT-A microtubule–associated complex requires Pasovec/Gid8 proteins
    Article Snippet: .. Cells were then transfected by Lipofectin (Invitrogen, A12621) with plasmid constructs [LacZ, full-length β-cat (Addgene, 14717), or Gid8 mut : 1 μg each)] together with 2 μg of pGL-TOP or its negative control vector pGL-FOP. ..

    Article Title: Differential AXL expression and Arf1 regulation control stiffness-dependent Golgi organization in breast cancer cells
    Article Snippet: .. The peGFP-N1-AXL plasmid construct was obtained from Dr. Stéphane Bodin (CRMB CNRS). pVSV-G ( Addgene #138479 ), gag/pol ( Addgene #14887 ), AXL-pBABE ( Addgene #105936 ), pBABE-puro ( Addgene #1764 ), and pMIG-w ( Addgene #12282 ) plasmid constructs were purchased from Addgene. siRNA against human AXL sequence (siAXL) was designed and purchased from Merck (Sigma). .. The siRNA sequence used was described previously ( ; ; forward, 5′-GAAAGAAGGAGACCCGTTA-3′ and reverse, 5′-TAACGGGTCTCCTTCTTTC-3′). siRNA against the human Arf1 sequence (siArf1) was designed and purchased from Merck (Sigma).

    Article Title: RhoA functionally collaborates with HSPA1A to promote the migratory phenotype of cancer cells.
    Article Snippet: RhoA, a member of the GTPase family, plays a pivotal role in attaining a migratory phenotype, mainly by regulating cytoskeleton dynamics, cell adhesion and membrane protrusions.. Although many upstream regulators and downstream effectors of RhoA have been identified, the discovery of new interacting partners continues to expand its interactome, providing fresh insights into its regulation and function.. Co-immunoprecipitation and fluorescence microscopy were used to study the interaction, localization and morphological effects of HSPA1A and RhoA.

    Article Title: The NSP5, ORF6 and NSP13 of SARS-CoV-2 Cooperate to Modulate Inflammatory Cell Death Activation.
    Article Snippet: .. Plasmid Constructs: Expression plasmid for SARS-CoV-2 spike (Addgene, 158761) was a gift fromGerald Pao,[94] and expression plasmids for SARS-CoV-2 membrane protein (Addgene, 141386) and envelope protein (Addgene, 141385) were a gift from Nevan Krogan. ..

    Article Title: Nuclear translocation of β-catenin in Wg/Wnt signaling via the IFT-A microtubule-associated complex requires Pasovec/Gid8 proteins.
    Article Snippet: .. Cells were then transfected by Lipofectin (Invitrogen, A12621) with plasmid constructs [LacZ, full- length β- cat (Addgene, 14717), or Gid8mut: 1 μ g each)] together with 2 μ g of pGL- TOP or its negative control vector pGL- FOP. ..

    Article Title: The N-terminal Segment of the Human ANKZF1 negatively regulates its internal mitochondrial targeting signal to prevent its mitochondrial localization
    Article Snippet: .. Plasmid constructs used in this study include the following: pCMV0-OCT-HA-eGFP (Addgene, #67479), pCMV0-SMAC-HA-eGFP (Addgene #67486), both the plasmids were a gift from Richard Kahn ( ). mCherry-Parkin (Addgene #23956) was a gift from Richard Youle , and Mito BFP (Addgene, #55248) ( ) was a gift from Michael Davidson. .. Δ73-ANKZF1-GFP, Δ210-ANKZF1-GFP, Δ220-ANKZF1-GFP, Δ230-ANKZF1-GFP, Δ240-ANKZF1-GFP, Δ250-ANKZF1-GFP, Δ330-ANKZF1-GFP, Δ370-ANKZF1-GFP, and Δ410-ANKZF1-GFP constructs were generated in the lab by using standard restriction digestion-based molecular cloning technique, and desired genes were digested with XhoI and KpnI restriction enzymes (NEB) and were ligated to pEGFP-N1 (Clontech) empty vector backbone.

    Control:

    Article Title: Avian photoreceptor homologies and the origin of double cones.
    Article Snippet: .. For reporter assays, two plasmid constructs were used: pX458-Ef1a-Cas9-H2B-mCherry (Addgene #159655),89 which contains a non-targeting control gRNA under the control of theU6 promoter and aCas9-T2A-mCherry cassette under the control of theEf1alpha promoter; and Ac-OPN1LW-GFP, which contains GFP under the control of a 1.8 Kb genomic fragment from the promoter region of Anolis carolinensis OPN1LW as previously described.24 To make an electroporation solution, Plasmid DNAs were precipitated by addition of 0.1 volumes of 3 M sodium acetate and 3 volumes of 100% ethanol, washed in 70% ethanol, resuspended to a final concentration of 1 mg/ml (pX458-Ef1a-Cas9-H2B-mCherry) or 0.5 mg/ml Ac-OPN1LW-GFP in 1X PBS, and combined in a single solution. ..

    Electroporation:

    Article Title: Avian photoreceptor homologies and the origin of double cones.
    Article Snippet: .. For reporter assays, two plasmid constructs were used: pX458-Ef1a-Cas9-H2B-mCherry (Addgene #159655),89 which contains a non-targeting control gRNA under the control of theU6 promoter and aCas9-T2A-mCherry cassette under the control of theEf1alpha promoter; and Ac-OPN1LW-GFP, which contains GFP under the control of a 1.8 Kb genomic fragment from the promoter region of Anolis carolinensis OPN1LW as previously described.24 To make an electroporation solution, Plasmid DNAs were precipitated by addition of 0.1 volumes of 3 M sodium acetate and 3 volumes of 100% ethanol, washed in 70% ethanol, resuspended to a final concentration of 1 mg/ml (pX458-Ef1a-Cas9-H2B-mCherry) or 0.5 mg/ml Ac-OPN1LW-GFP in 1X PBS, and combined in a single solution. ..

    Concentration Assay:

    Article Title: Avian photoreceptor homologies and the origin of double cones.
    Article Snippet: .. For reporter assays, two plasmid constructs were used: pX458-Ef1a-Cas9-H2B-mCherry (Addgene #159655),89 which contains a non-targeting control gRNA under the control of theU6 promoter and aCas9-T2A-mCherry cassette under the control of theEf1alpha promoter; and Ac-OPN1LW-GFP, which contains GFP under the control of a 1.8 Kb genomic fragment from the promoter region of Anolis carolinensis OPN1LW as previously described.24 To make an electroporation solution, Plasmid DNAs were precipitated by addition of 0.1 volumes of 3 M sodium acetate and 3 volumes of 100% ethanol, washed in 70% ethanol, resuspended to a final concentration of 1 mg/ml (pX458-Ef1a-Cas9-H2B-mCherry) or 0.5 mg/ml Ac-OPN1LW-GFP in 1X PBS, and combined in a single solution. ..

    Transfection:

    Article Title: TLE1 drives EGFR-TKI resistance in EGFR-mutant lung adenocarcinoma through epithelial to mesenchymal transition.
    Article Snippet: .. To transiently express E-cadherin or ZEB1, cells were transfected with E-cadherin-containing plasmid constructs (28009, Addgene) or ZEB1-containing plasmid (42100, Addgene) using Lipofectamine 3000 (L3000015, Invitrogen), adhering to the provider’s protocol. ..

    Article Title: Nuclear translocation of β-catenin in Wg/Wnt signaling via the IFT-A microtubule–associated complex requires Pasovec/Gid8 proteins
    Article Snippet: .. Cells were then transfected by Lipofectin (Invitrogen, A12621) with plasmid constructs [LacZ, full-length β-cat (Addgene, 14717), or Gid8 mut : 1 μg each)] together with 2 μg of pGL-TOP or its negative control vector pGL-FOP. ..

    Article Title: Nuclear translocation of β-catenin in Wg/Wnt signaling via the IFT-A microtubule-associated complex requires Pasovec/Gid8 proteins.
    Article Snippet: .. Cells were then transfected by Lipofectin (Invitrogen, A12621) with plasmid constructs [LacZ, full- length β- cat (Addgene, 14717), or Gid8mut: 1 μ g each)] together with 2 μ g of pGL- TOP or its negative control vector pGL- FOP. ..

    Negative Control:

    Article Title: Nuclear translocation of β-catenin in Wg/Wnt signaling via the IFT-A microtubule–associated complex requires Pasovec/Gid8 proteins
    Article Snippet: .. Cells were then transfected by Lipofectin (Invitrogen, A12621) with plasmid constructs [LacZ, full-length β-cat (Addgene, 14717), or Gid8 mut : 1 μg each)] together with 2 μg of pGL-TOP or its negative control vector pGL-FOP. ..

    Article Title: Nuclear translocation of β-catenin in Wg/Wnt signaling via the IFT-A microtubule-associated complex requires Pasovec/Gid8 proteins.
    Article Snippet: .. Cells were then transfected by Lipofectin (Invitrogen, A12621) with plasmid constructs [LacZ, full- length β- cat (Addgene, 14717), or Gid8mut: 1 μ g each)] together with 2 μ g of pGL- TOP or its negative control vector pGL- FOP. ..

    Sequencing:

    Article Title: Differential AXL expression and Arf1 regulation control stiffness-dependent Golgi organization in breast cancer cells
    Article Snippet: .. The peGFP-N1-AXL plasmid construct was obtained from Dr. Stéphane Bodin (CRMB CNRS). pVSV-G ( Addgene #138479 ), gag/pol ( Addgene #14887 ), AXL-pBABE ( Addgene #105936 ), pBABE-puro ( Addgene #1764 ), and pMIG-w ( Addgene #12282 ) plasmid constructs were purchased from Addgene. siRNA against human AXL sequence (siAXL) was designed and purchased from Merck (Sigma). .. The siRNA sequence used was described previously ( ; ; forward, 5′-GAAAGAAGGAGACCCGTTA-3′ and reverse, 5′-TAACGGGTCTCCTTCTTTC-3′). siRNA against the human Arf1 sequence (siArf1) was designed and purchased from Merck (Sigma).

    Expressing:

    Article Title: The NSP5, ORF6 and NSP13 of SARS-CoV-2 Cooperate to Modulate Inflammatory Cell Death Activation.
    Article Snippet: .. Plasmid Constructs: Expression plasmid for SARS-CoV-2 spike (Addgene, 158761) was a gift fromGerald Pao,[94] and expression plasmids for SARS-CoV-2 membrane protein (Addgene, 141386) and envelope protein (Addgene, 141385) were a gift from Nevan Krogan. ..

    Membrane:

    Article Title: The NSP5, ORF6 and NSP13 of SARS-CoV-2 Cooperate to Modulate Inflammatory Cell Death Activation.
    Article Snippet: .. Plasmid Constructs: Expression plasmid for SARS-CoV-2 spike (Addgene, 158761) was a gift fromGerald Pao,[94] and expression plasmids for SARS-CoV-2 membrane protein (Addgene, 141386) and envelope protein (Addgene, 141385) were a gift from Nevan Krogan. ..



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    (A) HEp-2 cells were “mock” infected (NI) or infected with RSV for 24 h, at MOI 1 and processed for IP using M antibodies. Input (1 %), unbound (1 %) and bound (30 %) fractions were loaded on SDS-PAGE and a Western blot was performed using antibodies indicated on the left. Representative of three independent experiments. (B) <t>HEp-2-Drebrin-YFP</t> and HEp-2-YFP cells were infected with RSV for 24 h, at MOI 1 and processed for IP using anti-GFP nanobodies. Input (0,5 %), unbound (0,5 %) and bound (30 %) fractions were loaded on SDS-PAGE and a Western blot was performed using antibodies indicated on the left. Drebrin-YFP and YFP corresponding bands were indicated by an arrow on the left. Representative of three independent experiments.
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    Image Search Results


    (A) HEp-2 cells were “mock” infected (NI) or infected with RSV for 24 h, at MOI 1 and processed for IP using M antibodies. Input (1 %), unbound (1 %) and bound (30 %) fractions were loaded on SDS-PAGE and a Western blot was performed using antibodies indicated on the left. Representative of three independent experiments. (B) HEp-2-Drebrin-YFP and HEp-2-YFP cells were infected with RSV for 24 h, at MOI 1 and processed for IP using anti-GFP nanobodies. Input (0,5 %), unbound (0,5 %) and bound (30 %) fractions were loaded on SDS-PAGE and a Western blot was performed using antibodies indicated on the left. Drebrin-YFP and YFP corresponding bands were indicated by an arrow on the left. Representative of three independent experiments.

    Journal: bioRxiv

    Article Title: The actin-binding protein drebrin interacts with the respiratory syncytial virus matrix protein and promotes virus particle release

    doi: 10.64898/2026.10.05.756730

    Figure Lengend Snippet: (A) HEp-2 cells were “mock” infected (NI) or infected with RSV for 24 h, at MOI 1 and processed for IP using M antibodies. Input (1 %), unbound (1 %) and bound (30 %) fractions were loaded on SDS-PAGE and a Western blot was performed using antibodies indicated on the left. Representative of three independent experiments. (B) HEp-2-Drebrin-YFP and HEp-2-YFP cells were infected with RSV for 24 h, at MOI 1 and processed for IP using anti-GFP nanobodies. Input (0,5 %), unbound (0,5 %) and bound (30 %) fractions were loaded on SDS-PAGE and a Western blot was performed using antibodies indicated on the left. Drebrin-YFP and YFP corresponding bands were indicated by an arrow on the left. Representative of three independent experiments.

    Article Snippet: YFP-tagged DBN (drebrin-YFP) construct has been provided from addgene (#40359).

    Techniques: Infection, SDS Page, Western Blot

    Drebrin promotes viral particle release . (A) Schematic representation of the release assay. HEp-2 cells were transfected with control or specific siRNAs and seeded onto Transwell inserts for 48h (step 1). Cells were then infected with RSV-mCherry at an MOI of 1 (step 2). After washes to remove non-internalized virus, the infected “donor” cells were then placed in contact with target cells (step 3). The number of infected “donor” and “target” cells were quantified based on mCherry fluorescence using the Incucyte S3 system at 20h (step4) and 38h (step 5) post-infection respectively. RSV release efficiency was calculated as the ratio of infected target cells to infected donor cells and normalized to the control siRNA condition. (B) HEp-2 cells were transfected with si NT or drebrin-specific siRNAs (si drebrin) and release assay was performed as described in A. Date are mean ± SD of 6 independent experiments, each point represents mean of triplicates. Y axis represents viral release efficiency (ratio of infected target cells to infected donor cells) normalized to the control siRNA condition. (C) release assay was performed with HEp-2-YFP and HEp-2-Drebrin-YFP cells. Data are mean ± SD of 4 independent experiments done in triplicates. Y axis represents viral release efficiency (ratio of infected target cells to infected donor cells) normalized to the control siRNA condition. (B-C) A one-sample t-test was performed. * : P < 0.05; ** : P < 0.001.

    Journal: bioRxiv

    Article Title: The actin-binding protein drebrin interacts with the respiratory syncytial virus matrix protein and promotes virus particle release

    doi: 10.64898/2026.10.05.756730

    Figure Lengend Snippet: Drebrin promotes viral particle release . (A) Schematic representation of the release assay. HEp-2 cells were transfected with control or specific siRNAs and seeded onto Transwell inserts for 48h (step 1). Cells were then infected with RSV-mCherry at an MOI of 1 (step 2). After washes to remove non-internalized virus, the infected “donor” cells were then placed in contact with target cells (step 3). The number of infected “donor” and “target” cells were quantified based on mCherry fluorescence using the Incucyte S3 system at 20h (step4) and 38h (step 5) post-infection respectively. RSV release efficiency was calculated as the ratio of infected target cells to infected donor cells and normalized to the control siRNA condition. (B) HEp-2 cells were transfected with si NT or drebrin-specific siRNAs (si drebrin) and release assay was performed as described in A. Date are mean ± SD of 6 independent experiments, each point represents mean of triplicates. Y axis represents viral release efficiency (ratio of infected target cells to infected donor cells) normalized to the control siRNA condition. (C) release assay was performed with HEp-2-YFP and HEp-2-Drebrin-YFP cells. Data are mean ± SD of 4 independent experiments done in triplicates. Y axis represents viral release efficiency (ratio of infected target cells to infected donor cells) normalized to the control siRNA condition. (B-C) A one-sample t-test was performed. * : P < 0.05; ** : P < 0.001.

    Article Snippet: YFP-tagged DBN (drebrin-YFP) construct has been provided from addgene (#40359).

    Techniques: Release Assay, Transfection, Control, Infection, Virus, Fluorescence

    (A) diagrams of drebrin mutants are shown. ADFH, actin-depolymerizing factor homology domain. ABD1, actin binding domain 1. ABD2, actin binding domain 2. IDD intrinsically disordered domain. Constructs are YFP-tagged at the C-terminus. Numbering is for human drebrin E. (B) HEp-2 cells lines were established with the indicated plasmids and cells were infected with RSV for 24h at MOI 1 and processed for IP using anti-GFP nanobodies. Input (0,5 %), unbound (0,5 %) and bound (30 %) fractions were loaded on SDS-PAGE and Western blotting using antibodies indicated on the left. Representative of three independent experiments. (C) release assay was performed as in with cells expressing indicated plasmids. Data are mean ± SD of 6 independent experiments done in triplicates. Y axis represents viral release efficiency (ratio of infected target cells to infected donor cells) normalized to the control siRNA condition. One-Way ANOVA with Dunnett’s multiple comparisons test and Geisser-Greenhouse correction was performed, ns : P > 0.05, ** : P < 0.01, **** : P < 0.0001.

    Journal: bioRxiv

    Article Title: The actin-binding protein drebrin interacts with the respiratory syncytial virus matrix protein and promotes virus particle release

    doi: 10.64898/2026.10.05.756730

    Figure Lengend Snippet: (A) diagrams of drebrin mutants are shown. ADFH, actin-depolymerizing factor homology domain. ABD1, actin binding domain 1. ABD2, actin binding domain 2. IDD intrinsically disordered domain. Constructs are YFP-tagged at the C-terminus. Numbering is for human drebrin E. (B) HEp-2 cells lines were established with the indicated plasmids and cells were infected with RSV for 24h at MOI 1 and processed for IP using anti-GFP nanobodies. Input (0,5 %), unbound (0,5 %) and bound (30 %) fractions were loaded on SDS-PAGE and Western blotting using antibodies indicated on the left. Representative of three independent experiments. (C) release assay was performed as in with cells expressing indicated plasmids. Data are mean ± SD of 6 independent experiments done in triplicates. Y axis represents viral release efficiency (ratio of infected target cells to infected donor cells) normalized to the control siRNA condition. One-Way ANOVA with Dunnett’s multiple comparisons test and Geisser-Greenhouse correction was performed, ns : P > 0.05, ** : P < 0.01, **** : P < 0.0001.

    Article Snippet: YFP-tagged DBN (drebrin-YFP) construct has been provided from addgene (#40359).

    Techniques: Binding Assay, Construct, Infection, SDS Page, Western Blot, Release Assay, Expressing, Control

    (A) Drebrin-YFP HEp-2 cells were uninfected or infected with RSV at MOI 1 for 24 h. Proteins associated with drebrin-YFP were affinity-purified using GFP nanobody-coupled agarose beads, identified by mass spectrometry, and quantified based on spectral counts obtained for each sample. A log2 fold change was calculated by comparing RSV-infected and uninfected conditions. Only proteins with at least two spectral counts in at least two biological replicates in either condition were included in the analysis. Statistical significance was assessed using a beta-binomial test adapted for spectral count data. Results are presented as a volcano plot. The horizontal dashed gray line indicates the statistical significance threshold ( P < 0.05), whereas the vertical dashed gray lines indicate a twofold difference between infected and uninfected samples. Cellular proteins used in C are indicated in blue. Viral RSV proteins are indicated in black. Results from three independent biological replicates (B) Left: Heat map of RT-qPCR analysis comparing mRNA levels of 17 candidate proteins in RSV-infected and uninfected cells. Blue indicates > 3-fold upregulation and yellow < 3-fold. Right: Heat map of viral replication analysis comparing RSV-mCherry signal in RSV-infected cells treated with interactors-specific siRNA and si NT. Blue indicates replication >140%, brown replication < 60% and yellow 60% < replication >140%. Complete data are shown in Fig S8. (C) HEp-2 cells were depleted with the indicated siRNAs and viral release assay was performed and compared with control conditions. One-Way ANOVA with Dunnett’s multiple comparisons test and Geisser-Greenhouse correction was performed relative to si NT. *** : P <0.001, ns : P >0,05.

    Journal: bioRxiv

    Article Title: The actin-binding protein drebrin interacts with the respiratory syncytial virus matrix protein and promotes virus particle release

    doi: 10.64898/2026.10.05.756730

    Figure Lengend Snippet: (A) Drebrin-YFP HEp-2 cells were uninfected or infected with RSV at MOI 1 for 24 h. Proteins associated with drebrin-YFP were affinity-purified using GFP nanobody-coupled agarose beads, identified by mass spectrometry, and quantified based on spectral counts obtained for each sample. A log2 fold change was calculated by comparing RSV-infected and uninfected conditions. Only proteins with at least two spectral counts in at least two biological replicates in either condition were included in the analysis. Statistical significance was assessed using a beta-binomial test adapted for spectral count data. Results are presented as a volcano plot. The horizontal dashed gray line indicates the statistical significance threshold ( P < 0.05), whereas the vertical dashed gray lines indicate a twofold difference between infected and uninfected samples. Cellular proteins used in C are indicated in blue. Viral RSV proteins are indicated in black. Results from three independent biological replicates (B) Left: Heat map of RT-qPCR analysis comparing mRNA levels of 17 candidate proteins in RSV-infected and uninfected cells. Blue indicates > 3-fold upregulation and yellow < 3-fold. Right: Heat map of viral replication analysis comparing RSV-mCherry signal in RSV-infected cells treated with interactors-specific siRNA and si NT. Blue indicates replication >140%, brown replication < 60% and yellow 60% < replication >140%. Complete data are shown in Fig S8. (C) HEp-2 cells were depleted with the indicated siRNAs and viral release assay was performed and compared with control conditions. One-Way ANOVA with Dunnett’s multiple comparisons test and Geisser-Greenhouse correction was performed relative to si NT. *** : P <0.001, ns : P >0,05.

    Article Snippet: YFP-tagged DBN (drebrin-YFP) construct has been provided from addgene (#40359).

    Techniques: Infection, Affinity Purification, Mass Spectrometry, Analysis, Quantitative RT-PCR, Release Assay, Control, Significance Assay