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rabbit polyclonal anti atg16l1 antibody  (Cell Signaling Technology Inc)


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    Cell Signaling Technology Inc rabbit polyclonal anti atg16l1 antibody
    Autophagy is impaired in WABS patient and DDX11 KO RPE-1 cells. (A) Fibroblasts from three different WABS patients and from a healthy donor (HF1) were left in full medium or serum starved for 16 h. LC3-II level was assessed by western blot analysis. TUBA/tubulin was used as a loading control. (B) Graphs show the quantifications of LC3-II:TUBA. (C,D) fibroblasts from three different WABS patients and from a healthy donor (HF1) were treated with BAF A 1 for 16 h. LC3-II level was assessed by western blot analysis. TUBA was used as loading control. Graphs show the quantifications of LC3-II:TUBA. (E) RNA-seq analysis shows no difference in transcripts per kilobase million/TPM of autophagy genes ( <t>ATG16L1</t> , ATG5 , ATG12 , BECN1 , SQSTM1 ) between control (CTRL) and DDX11 KO ( DDX11 KO) RPE-1 cells. (F) Western blot analysis revealed no difference in the expression level of the indicated autophagy proteins between control (CTRL) and DDX11 KO ( DDX11 KO) RPE-1 cells. TUBA was used as loading control. (G,H) control (CTRL) and DDX11 KO ( DDX11 KO) RPE-1 cells were grown in full-medium (FM) or serum starved for 16 h (NO FBS). LC3-II was analyzed by western blot. TUBA was used as loading control. Graphs show the quantifications of LC3-II:TUBA. (I,J) Control (CTRL) and DDX11 KO ( DDX11 KO) RPE-1 cells were treated with DMSO or BAF A 1 for 6 h. LC3-II level was analyzed by western blot. TUBA was used as loading control. Graphs show the quantifications of LC3-II:TUBA. (K–M) DDX11 KO (KO) RPE-1 cells were transfected with a vector expressing DDX11-Flag wild-type protein (WT) or the DDX11-Flag K50R helicase-dead mutant (K50R) to perform rescue experiments. After 24 h, control (CTRL), DDX11 KO ( DDX11 KO) and DDX11-complemented DDX11 KO (KO + WT and KO + K50R) RPE-1 cells were treated with DMSO or BAF A 1 for 6 h. LC3 level (LC3-I and LC3-II) was analyzed by western blot. Graphs show the quantifications of LC3-II:TUBA. (N) Control (CTRL) and DDX11 KO ( DDX11 KO) RPE-1 cells were transfected with a vector expressing EGFP-HTT-74Q for 48 h to assess HTT mutant protein clearance. (O) Graph shows increased percentage of HTT-positive cells in the DDX11 KO RPE-1 line. Number of cells counted n = 50 in triplicates. Scale bar: 10 µm.
    Rabbit Polyclonal Anti Atg16l1 Antibody, 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/product/rabbit+polyclonal+anti+atg16l1+antibody/pmc12674475-240-44-56
    Average 86 stars, based on 1 article reviews
    rabbit polyclonal anti atg16l1 antibody - by Bioz Stars, 2026-10
    86/100 stars

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    1) Product Images from "Evidence of an unprecedented cytoplasmic function of DDX11, the Warsaw breakage syndrome DNA helicase, in regulating autophagy"

    Article Title: Evidence of an unprecedented cytoplasmic function of DDX11, the Warsaw breakage syndrome DNA helicase, in regulating autophagy

    Journal: Autophagy

    doi: 10.1080/15548627.2025.2507617

    Autophagy is impaired in WABS patient and DDX11 KO RPE-1 cells. (A) Fibroblasts from three different WABS patients and from a healthy donor (HF1) were left in full medium or serum starved for 16 h. LC3-II level was assessed by western blot analysis. TUBA/tubulin was used as a loading control. (B) Graphs show the quantifications of LC3-II:TUBA. (C,D) fibroblasts from three different WABS patients and from a healthy donor (HF1) were treated with BAF A 1 for 16 h. LC3-II level was assessed by western blot analysis. TUBA was used as loading control. Graphs show the quantifications of LC3-II:TUBA. (E) RNA-seq analysis shows no difference in transcripts per kilobase million/TPM of autophagy genes ( ATG16L1 , ATG5 , ATG12 , BECN1 , SQSTM1 ) between control (CTRL) and DDX11 KO ( DDX11 KO) RPE-1 cells. (F) Western blot analysis revealed no difference in the expression level of the indicated autophagy proteins between control (CTRL) and DDX11 KO ( DDX11 KO) RPE-1 cells. TUBA was used as loading control. (G,H) control (CTRL) and DDX11 KO ( DDX11 KO) RPE-1 cells were grown in full-medium (FM) or serum starved for 16 h (NO FBS). LC3-II was analyzed by western blot. TUBA was used as loading control. Graphs show the quantifications of LC3-II:TUBA. (I,J) Control (CTRL) and DDX11 KO ( DDX11 KO) RPE-1 cells were treated with DMSO or BAF A 1 for 6 h. LC3-II level was analyzed by western blot. TUBA was used as loading control. Graphs show the quantifications of LC3-II:TUBA. (K–M) DDX11 KO (KO) RPE-1 cells were transfected with a vector expressing DDX11-Flag wild-type protein (WT) or the DDX11-Flag K50R helicase-dead mutant (K50R) to perform rescue experiments. After 24 h, control (CTRL), DDX11 KO ( DDX11 KO) and DDX11-complemented DDX11 KO (KO + WT and KO + K50R) RPE-1 cells were treated with DMSO or BAF A 1 for 6 h. LC3 level (LC3-I and LC3-II) was analyzed by western blot. Graphs show the quantifications of LC3-II:TUBA. (N) Control (CTRL) and DDX11 KO ( DDX11 KO) RPE-1 cells were transfected with a vector expressing EGFP-HTT-74Q for 48 h to assess HTT mutant protein clearance. (O) Graph shows increased percentage of HTT-positive cells in the DDX11 KO RPE-1 line. Number of cells counted n = 50 in triplicates. Scale bar: 10 µm.
    Figure Legend Snippet: Autophagy is impaired in WABS patient and DDX11 KO RPE-1 cells. (A) Fibroblasts from three different WABS patients and from a healthy donor (HF1) were left in full medium or serum starved for 16 h. LC3-II level was assessed by western blot analysis. TUBA/tubulin was used as a loading control. (B) Graphs show the quantifications of LC3-II:TUBA. (C,D) fibroblasts from three different WABS patients and from a healthy donor (HF1) were treated with BAF A 1 for 16 h. LC3-II level was assessed by western blot analysis. TUBA was used as loading control. Graphs show the quantifications of LC3-II:TUBA. (E) RNA-seq analysis shows no difference in transcripts per kilobase million/TPM of autophagy genes ( ATG16L1 , ATG5 , ATG12 , BECN1 , SQSTM1 ) between control (CTRL) and DDX11 KO ( DDX11 KO) RPE-1 cells. (F) Western blot analysis revealed no difference in the expression level of the indicated autophagy proteins between control (CTRL) and DDX11 KO ( DDX11 KO) RPE-1 cells. TUBA was used as loading control. (G,H) control (CTRL) and DDX11 KO ( DDX11 KO) RPE-1 cells were grown in full-medium (FM) or serum starved for 16 h (NO FBS). LC3-II was analyzed by western blot. TUBA was used as loading control. Graphs show the quantifications of LC3-II:TUBA. (I,J) Control (CTRL) and DDX11 KO ( DDX11 KO) RPE-1 cells were treated with DMSO or BAF A 1 for 6 h. LC3-II level was analyzed by western blot. TUBA was used as loading control. Graphs show the quantifications of LC3-II:TUBA. (K–M) DDX11 KO (KO) RPE-1 cells were transfected with a vector expressing DDX11-Flag wild-type protein (WT) or the DDX11-Flag K50R helicase-dead mutant (K50R) to perform rescue experiments. After 24 h, control (CTRL), DDX11 KO ( DDX11 KO) and DDX11-complemented DDX11 KO (KO + WT and KO + K50R) RPE-1 cells were treated with DMSO or BAF A 1 for 6 h. LC3 level (LC3-I and LC3-II) was analyzed by western blot. Graphs show the quantifications of LC3-II:TUBA. (N) Control (CTRL) and DDX11 KO ( DDX11 KO) RPE-1 cells were transfected with a vector expressing EGFP-HTT-74Q for 48 h to assess HTT mutant protein clearance. (O) Graph shows increased percentage of HTT-positive cells in the DDX11 KO RPE-1 line. Number of cells counted n = 50 in triplicates. Scale bar: 10 µm.

    Techniques Used: Western Blot, Control, RNA Sequencing, Expressing, Transfection, Plasmid Preparation, Mutagenesis

    DDX11 regulates ATG16L1 localization. (A,B) Control (CTRL), DDX11 KO ( DDX11 KO) and DDX11-complemented DDX11 KO ( DDX11 KO + WT DDX11 ) RPE-1 cells were transfected with ATG16L1-GFP for 24 h. Confocal imaging shows accumulation of ATG16L1 in the perinuclear area that was reverted by re-expressing the DDX11-Flag wild-type protein. Total cells analyzed: n = 50 per experiment, performed in triplicates. Scale bar: 10 µm. (C) Co-immunoprecipitation experiment using an anti-ATG16 antibody in control (CTRL) and DDX11 KO RPE-1 cells. Western blot analysis shows co-immunoprecipitation of ATG5-ATG12 in both control and DDX11 KO ( DDX11 KO) RPE-1 cells indicating that, in the absence of DDX11, the ATG12–ATG5-ATG16L1 complex formation is not affected. Asterisks denote the IgG heavy chains. (D) Control (CTRL), DDX11 KO ( DDX11 KO) and DDX11-complemented DDX11 KO ( DDX11 KO + WT DDX11 ) RPE-1 cells were transfected with a vector expressing GFP-ATG16L1 for 48 h. Co-immunoprecipitation experiment was performed using anti-GFP antibody conjugated beads. The indicated proteins were detected by western blot of the pulled-down sample using specific antibodies. (E) Control (CTRL), DDX11 KO ( DDX11 KO) and DDX11-complemented DDX11 KO ( DDX11 KO + WT DDX11 ) RPE-1 cells were transfected with GFP-ATG16L1 and mCherry-LC3 for 24 h. Then, cells were processed as described for panel A. Confocal imaging reveals that ATG16L1 and LC3 do not colocalize in DDX11 KO cells. This phenotype is reversed by re-expressing the DDX11-Flag wild-type protein. Scale bar: 10 µm.
    Figure Legend Snippet: DDX11 regulates ATG16L1 localization. (A,B) Control (CTRL), DDX11 KO ( DDX11 KO) and DDX11-complemented DDX11 KO ( DDX11 KO + WT DDX11 ) RPE-1 cells were transfected with ATG16L1-GFP for 24 h. Confocal imaging shows accumulation of ATG16L1 in the perinuclear area that was reverted by re-expressing the DDX11-Flag wild-type protein. Total cells analyzed: n = 50 per experiment, performed in triplicates. Scale bar: 10 µm. (C) Co-immunoprecipitation experiment using an anti-ATG16 antibody in control (CTRL) and DDX11 KO RPE-1 cells. Western blot analysis shows co-immunoprecipitation of ATG5-ATG12 in both control and DDX11 KO ( DDX11 KO) RPE-1 cells indicating that, in the absence of DDX11, the ATG12–ATG5-ATG16L1 complex formation is not affected. Asterisks denote the IgG heavy chains. (D) Control (CTRL), DDX11 KO ( DDX11 KO) and DDX11-complemented DDX11 KO ( DDX11 KO + WT DDX11 ) RPE-1 cells were transfected with a vector expressing GFP-ATG16L1 for 48 h. Co-immunoprecipitation experiment was performed using anti-GFP antibody conjugated beads. The indicated proteins were detected by western blot of the pulled-down sample using specific antibodies. (E) Control (CTRL), DDX11 KO ( DDX11 KO) and DDX11-complemented DDX11 KO ( DDX11 KO + WT DDX11 ) RPE-1 cells were transfected with GFP-ATG16L1 and mCherry-LC3 for 24 h. Then, cells were processed as described for panel A. Confocal imaging reveals that ATG16L1 and LC3 do not colocalize in DDX11 KO cells. This phenotype is reversed by re-expressing the DDX11-Flag wild-type protein. Scale bar: 10 µm.

    Techniques Used: Control, Transfection, Imaging, Expressing, Immunoprecipitation, Western Blot, Plasmid Preparation

    Schematic model describing a putative role of DDX11 in autophagy pathway regulation. DDX11 exerts its function as a DNA helicase in the nuclear compartment. Within the cytoplasm, DDX11 cooperates with SQSTM1 and ATG16L1 to promote phagophore formation as well as the autophagic flux (bold green and blue dotted arrows, respectively; left side ). Absence of DDX11 affects intracellular localization of the ATG16L1 autophagy precursor, which in turn impairs LC3 lipidation and autophagosome fusion with lysosomal compartment, ultimately reducing the autophagic flux (thin green and blue dotted arrows, respectively; right side ).
    Figure Legend Snippet: Schematic model describing a putative role of DDX11 in autophagy pathway regulation. DDX11 exerts its function as a DNA helicase in the nuclear compartment. Within the cytoplasm, DDX11 cooperates with SQSTM1 and ATG16L1 to promote phagophore formation as well as the autophagic flux (bold green and blue dotted arrows, respectively; left side ). Absence of DDX11 affects intracellular localization of the ATG16L1 autophagy precursor, which in turn impairs LC3 lipidation and autophagosome fusion with lysosomal compartment, ultimately reducing the autophagic flux (thin green and blue dotted arrows, respectively; right side ).

    Techniques Used:

    Related Articles

    Western Blot:

    Article Title: Evidence of an unprecedented cytoplasmic function of DDX11, the Warsaw breakage syndrome DNA helicase, in regulating autophagy
    Article Snippet: The following antibodies were used: mouse monoclonal anti DDX11 (Santa Cruz Biotechnology, sc271711); mouse monoclonal anti-Flag antibody (Merck, F1804); rabbit polyclonal anti-LC3 (Novus Biologicals, NB100–2220); mouse monoclonal anti-LAMP1 (Cell Signaling Technology, 15665); rabbit polyclonal anti-SQSTM1 (MBL International, PM045); mouse monoclonal anti-TUBA/α-tubulin antibody (Merck, T6199); rabbit polyclonal anti-ATG16L1 antibody (MBL Life Sciences, PM040); rabbit monoclonal anti-ATG5 antibody (Cell Signaling Technology, 129945); rabbit polyclonal anti-BECN1/BECLIN1 antibody (Cell Signaling Technology, 3738S); rabbit polyclonal anti-AKT antibody (Cell Signaling Technology, 9272S); rabbit polyclonal anti-phospho-AKT (Ser473) antibody (Cell Signaling Technology, 9271 L); rabbit polyclonal anti-BRIP1/FANCJ antibody (Novus Biologicals, NBP1–31883); mouse monoclonal anti-histone H3 antibody (Abcam, ab24834); rabbit monoclonal anti-GFP antibody (Cell Signaling Technology, 2956); mouse and rabbit HRP-conjugated secondary antibody (Merck, A9044 and 12–348, respectively); Alexa Fluor 488 anti-rabbit antibody, Alexa Fluor 555 anti-mouse antibody and Alexa Fluor 633 anti-mouse antibody (Thermo Fisher Scientific, A-11008, A-21137, A-21146, respectively).

    Control:

    Article Title: Evidence of an unprecedented cytoplasmic function of DDX11, the Warsaw breakage syndrome DNA helicase, in regulating autophagy
    Article Snippet: The following antibodies were used: mouse monoclonal anti DDX11 (Santa Cruz Biotechnology, sc271711); mouse monoclonal anti-Flag antibody (Merck, F1804); rabbit polyclonal anti-LC3 (Novus Biologicals, NB100–2220); mouse monoclonal anti-LAMP1 (Cell Signaling Technology, 15665); rabbit polyclonal anti-SQSTM1 (MBL International, PM045); mouse monoclonal anti-TUBA/α-tubulin antibody (Merck, T6199); rabbit polyclonal anti-ATG16L1 antibody (MBL Life Sciences, PM040); rabbit monoclonal anti-ATG5 antibody (Cell Signaling Technology, 129945); rabbit polyclonal anti-BECN1/BECLIN1 antibody (Cell Signaling Technology, 3738S); rabbit polyclonal anti-AKT antibody (Cell Signaling Technology, 9272S); rabbit polyclonal anti-phospho-AKT (Ser473) antibody (Cell Signaling Technology, 9271 L); rabbit polyclonal anti-BRIP1/FANCJ antibody (Novus Biologicals, NBP1–31883); mouse monoclonal anti-histone H3 antibody (Abcam, ab24834); rabbit monoclonal anti-GFP antibody (Cell Signaling Technology, 2956); mouse and rabbit HRP-conjugated secondary antibody (Merck, A9044 and 12–348, respectively); Alexa Fluor 488 anti-rabbit antibody, Alexa Fluor 555 anti-mouse antibody and Alexa Fluor 633 anti-mouse antibody (Thermo Fisher Scientific, A-11008, A-21137, A-21146, respectively).

    RNA Sequencing:

    Article Title: Evidence of an unprecedented cytoplasmic function of DDX11, the Warsaw breakage syndrome DNA helicase, in regulating autophagy
    Article Snippet: The following antibodies were used: mouse monoclonal anti DDX11 (Santa Cruz Biotechnology, sc271711); mouse monoclonal anti-Flag antibody (Merck, F1804); rabbit polyclonal anti-LC3 (Novus Biologicals, NB100–2220); mouse monoclonal anti-LAMP1 (Cell Signaling Technology, 15665); rabbit polyclonal anti-SQSTM1 (MBL International, PM045); mouse monoclonal anti-TUBA/α-tubulin antibody (Merck, T6199); rabbit polyclonal anti-ATG16L1 antibody (MBL Life Sciences, PM040); rabbit monoclonal anti-ATG5 antibody (Cell Signaling Technology, 129945); rabbit polyclonal anti-BECN1/BECLIN1 antibody (Cell Signaling Technology, 3738S); rabbit polyclonal anti-AKT antibody (Cell Signaling Technology, 9272S); rabbit polyclonal anti-phospho-AKT (Ser473) antibody (Cell Signaling Technology, 9271 L); rabbit polyclonal anti-BRIP1/FANCJ antibody (Novus Biologicals, NBP1–31883); mouse monoclonal anti-histone H3 antibody (Abcam, ab24834); rabbit monoclonal anti-GFP antibody (Cell Signaling Technology, 2956); mouse and rabbit HRP-conjugated secondary antibody (Merck, A9044 and 12–348, respectively); Alexa Fluor 488 anti-rabbit antibody, Alexa Fluor 555 anti-mouse antibody and Alexa Fluor 633 anti-mouse antibody (Thermo Fisher Scientific, A-11008, A-21137, A-21146, respectively).

    Expressing:

    Article Title: Evidence of an unprecedented cytoplasmic function of DDX11, the Warsaw breakage syndrome DNA helicase, in regulating autophagy
    Article Snippet: The following antibodies were used: mouse monoclonal anti DDX11 (Santa Cruz Biotechnology, sc271711); mouse monoclonal anti-Flag antibody (Merck, F1804); rabbit polyclonal anti-LC3 (Novus Biologicals, NB100–2220); mouse monoclonal anti-LAMP1 (Cell Signaling Technology, 15665); rabbit polyclonal anti-SQSTM1 (MBL International, PM045); mouse monoclonal anti-TUBA/α-tubulin antibody (Merck, T6199); rabbit polyclonal anti-ATG16L1 antibody (MBL Life Sciences, PM040); rabbit monoclonal anti-ATG5 antibody (Cell Signaling Technology, 129945); rabbit polyclonal anti-BECN1/BECLIN1 antibody (Cell Signaling Technology, 3738S); rabbit polyclonal anti-AKT antibody (Cell Signaling Technology, 9272S); rabbit polyclonal anti-phospho-AKT (Ser473) antibody (Cell Signaling Technology, 9271 L); rabbit polyclonal anti-BRIP1/FANCJ antibody (Novus Biologicals, NBP1–31883); mouse monoclonal anti-histone H3 antibody (Abcam, ab24834); rabbit monoclonal anti-GFP antibody (Cell Signaling Technology, 2956); mouse and rabbit HRP-conjugated secondary antibody (Merck, A9044 and 12–348, respectively); Alexa Fluor 488 anti-rabbit antibody, Alexa Fluor 555 anti-mouse antibody and Alexa Fluor 633 anti-mouse antibody (Thermo Fisher Scientific, A-11008, A-21137, A-21146, respectively).

    Transfection:

    Article Title: Evidence of an unprecedented cytoplasmic function of DDX11, the Warsaw breakage syndrome DNA helicase, in regulating autophagy
    Article Snippet: The following antibodies were used: mouse monoclonal anti DDX11 (Santa Cruz Biotechnology, sc271711); mouse monoclonal anti-Flag antibody (Merck, F1804); rabbit polyclonal anti-LC3 (Novus Biologicals, NB100–2220); mouse monoclonal anti-LAMP1 (Cell Signaling Technology, 15665); rabbit polyclonal anti-SQSTM1 (MBL International, PM045); mouse monoclonal anti-TUBA/α-tubulin antibody (Merck, T6199); rabbit polyclonal anti-ATG16L1 antibody (MBL Life Sciences, PM040); rabbit monoclonal anti-ATG5 antibody (Cell Signaling Technology, 129945); rabbit polyclonal anti-BECN1/BECLIN1 antibody (Cell Signaling Technology, 3738S); rabbit polyclonal anti-AKT antibody (Cell Signaling Technology, 9272S); rabbit polyclonal anti-phospho-AKT (Ser473) antibody (Cell Signaling Technology, 9271 L); rabbit polyclonal anti-BRIP1/FANCJ antibody (Novus Biologicals, NBP1–31883); mouse monoclonal anti-histone H3 antibody (Abcam, ab24834); rabbit monoclonal anti-GFP antibody (Cell Signaling Technology, 2956); mouse and rabbit HRP-conjugated secondary antibody (Merck, A9044 and 12–348, respectively); Alexa Fluor 488 anti-rabbit antibody, Alexa Fluor 555 anti-mouse antibody and Alexa Fluor 633 anti-mouse antibody (Thermo Fisher Scientific, A-11008, A-21137, A-21146, respectively).

    Plasmid Preparation:

    Article Title: Evidence of an unprecedented cytoplasmic function of DDX11, the Warsaw breakage syndrome DNA helicase, in regulating autophagy
    Article Snippet: The following antibodies were used: mouse monoclonal anti DDX11 (Santa Cruz Biotechnology, sc271711); mouse monoclonal anti-Flag antibody (Merck, F1804); rabbit polyclonal anti-LC3 (Novus Biologicals, NB100–2220); mouse monoclonal anti-LAMP1 (Cell Signaling Technology, 15665); rabbit polyclonal anti-SQSTM1 (MBL International, PM045); mouse monoclonal anti-TUBA/α-tubulin antibody (Merck, T6199); rabbit polyclonal anti-ATG16L1 antibody (MBL Life Sciences, PM040); rabbit monoclonal anti-ATG5 antibody (Cell Signaling Technology, 129945); rabbit polyclonal anti-BECN1/BECLIN1 antibody (Cell Signaling Technology, 3738S); rabbit polyclonal anti-AKT antibody (Cell Signaling Technology, 9272S); rabbit polyclonal anti-phospho-AKT (Ser473) antibody (Cell Signaling Technology, 9271 L); rabbit polyclonal anti-BRIP1/FANCJ antibody (Novus Biologicals, NBP1–31883); mouse monoclonal anti-histone H3 antibody (Abcam, ab24834); rabbit monoclonal anti-GFP antibody (Cell Signaling Technology, 2956); mouse and rabbit HRP-conjugated secondary antibody (Merck, A9044 and 12–348, respectively); Alexa Fluor 488 anti-rabbit antibody, Alexa Fluor 555 anti-mouse antibody and Alexa Fluor 633 anti-mouse antibody (Thermo Fisher Scientific, A-11008, A-21137, A-21146, respectively).

    Mutagenesis:

    Article Title: Evidence of an unprecedented cytoplasmic function of DDX11, the Warsaw breakage syndrome DNA helicase, in regulating autophagy
    Article Snippet: The following antibodies were used: mouse monoclonal anti DDX11 (Santa Cruz Biotechnology, sc271711); mouse monoclonal anti-Flag antibody (Merck, F1804); rabbit polyclonal anti-LC3 (Novus Biologicals, NB100–2220); mouse monoclonal anti-LAMP1 (Cell Signaling Technology, 15665); rabbit polyclonal anti-SQSTM1 (MBL International, PM045); mouse monoclonal anti-TUBA/α-tubulin antibody (Merck, T6199); rabbit polyclonal anti-ATG16L1 antibody (MBL Life Sciences, PM040); rabbit monoclonal anti-ATG5 antibody (Cell Signaling Technology, 129945); rabbit polyclonal anti-BECN1/BECLIN1 antibody (Cell Signaling Technology, 3738S); rabbit polyclonal anti-AKT antibody (Cell Signaling Technology, 9272S); rabbit polyclonal anti-phospho-AKT (Ser473) antibody (Cell Signaling Technology, 9271 L); rabbit polyclonal anti-BRIP1/FANCJ antibody (Novus Biologicals, NBP1–31883); mouse monoclonal anti-histone H3 antibody (Abcam, ab24834); rabbit monoclonal anti-GFP antibody (Cell Signaling Technology, 2956); mouse and rabbit HRP-conjugated secondary antibody (Merck, A9044 and 12–348, respectively); Alexa Fluor 488 anti-rabbit antibody, Alexa Fluor 555 anti-mouse antibody and Alexa Fluor 633 anti-mouse antibody (Thermo Fisher Scientific, A-11008, A-21137, A-21146, respectively).

    Imaging:

    Article Title: Evidence of an unprecedented cytoplasmic function of DDX11, the Warsaw breakage syndrome DNA helicase, in regulating autophagy
    Article Snippet: The following antibodies were used: mouse monoclonal anti DDX11 (Santa Cruz Biotechnology, sc271711); mouse monoclonal anti-Flag antibody (Merck, F1804); rabbit polyclonal anti-LC3 (Novus Biologicals, NB100–2220); mouse monoclonal anti-LAMP1 (Cell Signaling Technology, 15665); rabbit polyclonal anti-SQSTM1 (MBL International, PM045); mouse monoclonal anti-TUBA/α-tubulin antibody (Merck, T6199); rabbit polyclonal anti-ATG16L1 antibody (MBL Life Sciences, PM040); rabbit monoclonal anti-ATG5 antibody (Cell Signaling Technology, 129945); rabbit polyclonal anti-BECN1/BECLIN1 antibody (Cell Signaling Technology, 3738S); rabbit polyclonal anti-AKT antibody (Cell Signaling Technology, 9272S); rabbit polyclonal anti-phospho-AKT (Ser473) antibody (Cell Signaling Technology, 9271 L); rabbit polyclonal anti-BRIP1/FANCJ antibody (Novus Biologicals, NBP1–31883); mouse monoclonal anti-histone H3 antibody (Abcam, ab24834); rabbit monoclonal anti-GFP antibody (Cell Signaling Technology, 2956); mouse and rabbit HRP-conjugated secondary antibody (Merck, A9044 and 12–348, respectively); Alexa Fluor 488 anti-rabbit antibody, Alexa Fluor 555 anti-mouse antibody and Alexa Fluor 633 anti-mouse antibody (Thermo Fisher Scientific, A-11008, A-21137, A-21146, respectively).

    Immunoprecipitation:

    Article Title: Evidence of an unprecedented cytoplasmic function of DDX11, the Warsaw breakage syndrome DNA helicase, in regulating autophagy
    Article Snippet: The following antibodies were used: mouse monoclonal anti DDX11 (Santa Cruz Biotechnology, sc271711); mouse monoclonal anti-Flag antibody (Merck, F1804); rabbit polyclonal anti-LC3 (Novus Biologicals, NB100–2220); mouse monoclonal anti-LAMP1 (Cell Signaling Technology, 15665); rabbit polyclonal anti-SQSTM1 (MBL International, PM045); mouse monoclonal anti-TUBA/α-tubulin antibody (Merck, T6199); rabbit polyclonal anti-ATG16L1 antibody (MBL Life Sciences, PM040); rabbit monoclonal anti-ATG5 antibody (Cell Signaling Technology, 129945); rabbit polyclonal anti-BECN1/BECLIN1 antibody (Cell Signaling Technology, 3738S); rabbit polyclonal anti-AKT antibody (Cell Signaling Technology, 9272S); rabbit polyclonal anti-phospho-AKT (Ser473) antibody (Cell Signaling Technology, 9271 L); rabbit polyclonal anti-BRIP1/FANCJ antibody (Novus Biologicals, NBP1–31883); mouse monoclonal anti-histone H3 antibody (Abcam, ab24834); rabbit monoclonal anti-GFP antibody (Cell Signaling Technology, 2956); mouse and rabbit HRP-conjugated secondary antibody (Merck, A9044 and 12–348, respectively); Alexa Fluor 488 anti-rabbit antibody, Alexa Fluor 555 anti-mouse antibody and Alexa Fluor 633 anti-mouse antibody (Thermo Fisher Scientific, A-11008, A-21137, A-21146, respectively).



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    rabbit polyclonal anti atg16l1 antibody  (Cell Signaling Technology Inc)
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    Cell Signaling Technology Inc rabbit polyclonal anti atg16l1 antibody
    Autophagy is impaired in WABS patient and DDX11 KO RPE-1 cells. (A) Fibroblasts from three different WABS patients and from a healthy donor (HF1) were left in full medium or serum starved for 16 h. LC3-II level was assessed by western blot analysis. TUBA/tubulin was used as a loading control. (B) Graphs show the quantifications of LC3-II:TUBA. (C,D) fibroblasts from three different WABS patients and from a healthy donor (HF1) were treated with BAF A 1 for 16 h. LC3-II level was assessed by western blot analysis. TUBA was used as loading control. Graphs show the quantifications of LC3-II:TUBA. (E) RNA-seq analysis shows no difference in transcripts per kilobase million/TPM of autophagy genes ( <t>ATG16L1</t> , ATG5 , ATG12 , BECN1 , SQSTM1 ) between control (CTRL) and DDX11 KO ( DDX11 KO) RPE-1 cells. (F) Western blot analysis revealed no difference in the expression level of the indicated autophagy proteins between control (CTRL) and DDX11 KO ( DDX11 KO) RPE-1 cells. TUBA was used as loading control. (G,H) control (CTRL) and DDX11 KO ( DDX11 KO) RPE-1 cells were grown in full-medium (FM) or serum starved for 16 h (NO FBS). LC3-II was analyzed by western blot. TUBA was used as loading control. Graphs show the quantifications of LC3-II:TUBA. (I,J) Control (CTRL) and DDX11 KO ( DDX11 KO) RPE-1 cells were treated with DMSO or BAF A 1 for 6 h. LC3-II level was analyzed by western blot. TUBA was used as loading control. Graphs show the quantifications of LC3-II:TUBA. (K–M) DDX11 KO (KO) RPE-1 cells were transfected with a vector expressing DDX11-Flag wild-type protein (WT) or the DDX11-Flag K50R helicase-dead mutant (K50R) to perform rescue experiments. After 24 h, control (CTRL), DDX11 KO ( DDX11 KO) and DDX11-complemented DDX11 KO (KO + WT and KO + K50R) RPE-1 cells were treated with DMSO or BAF A 1 for 6 h. LC3 level (LC3-I and LC3-II) was analyzed by western blot. Graphs show the quantifications of LC3-II:TUBA. (N) Control (CTRL) and DDX11 KO ( DDX11 KO) RPE-1 cells were transfected with a vector expressing EGFP-HTT-74Q for 48 h to assess HTT mutant protein clearance. (O) Graph shows increased percentage of HTT-positive cells in the DDX11 KO RPE-1 line. Number of cells counted n = 50 in triplicates. Scale bar: 10 µm.
    Rabbit Polyclonal Anti Atg16l1 Antibody, 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/product/rabbit+polyclonal+anti+atg16l1+antibody/pmc12674475-240-44-56
    Average 86 stars, based on 1 article reviews
    rabbit polyclonal anti atg16l1 antibody - by Bioz Stars, 2026-10
    86/100 stars
    		  Buy from Supplier
    antibody rabbit polyclonal anti-atg16l1  (MBL International)
    90
    MBL International antibody rabbit polyclonal anti-atg16l1
    Autophagy is impaired in WABS patient and DDX11 KO RPE-1 cells. (A) Fibroblasts from three different WABS patients and from a healthy donor (HF1) were left in full medium or serum starved for 16 h. LC3-II level was assessed by western blot analysis. TUBA/tubulin was used as a loading control. (B) Graphs show the quantifications of LC3-II:TUBA. (C,D) fibroblasts from three different WABS patients and from a healthy donor (HF1) were treated with BAF A 1 for 16 h. LC3-II level was assessed by western blot analysis. TUBA was used as loading control. Graphs show the quantifications of LC3-II:TUBA. (E) RNA-seq analysis shows no difference in transcripts per kilobase million/TPM of autophagy genes ( <t>ATG16L1</t> , ATG5 , ATG12 , BECN1 , SQSTM1 ) between control (CTRL) and DDX11 KO ( DDX11 KO) RPE-1 cells. (F) Western blot analysis revealed no difference in the expression level of the indicated autophagy proteins between control (CTRL) and DDX11 KO ( DDX11 KO) RPE-1 cells. TUBA was used as loading control. (G,H) control (CTRL) and DDX11 KO ( DDX11 KO) RPE-1 cells were grown in full-medium (FM) or serum starved for 16 h (NO FBS). LC3-II was analyzed by western blot. TUBA was used as loading control. Graphs show the quantifications of LC3-II:TUBA. (I,J) Control (CTRL) and DDX11 KO ( DDX11 KO) RPE-1 cells were treated with DMSO or BAF A 1 for 6 h. LC3-II level was analyzed by western blot. TUBA was used as loading control. Graphs show the quantifications of LC3-II:TUBA. (K–M) DDX11 KO (KO) RPE-1 cells were transfected with a vector expressing DDX11-Flag wild-type protein (WT) or the DDX11-Flag K50R helicase-dead mutant (K50R) to perform rescue experiments. After 24 h, control (CTRL), DDX11 KO ( DDX11 KO) and DDX11-complemented DDX11 KO (KO + WT and KO + K50R) RPE-1 cells were treated with DMSO or BAF A 1 for 6 h. LC3 level (LC3-I and LC3-II) was analyzed by western blot. Graphs show the quantifications of LC3-II:TUBA. (N) Control (CTRL) and DDX11 KO ( DDX11 KO) RPE-1 cells were transfected with a vector expressing EGFP-HTT-74Q for 48 h to assess HTT mutant protein clearance. (O) Graph shows increased percentage of HTT-positive cells in the DDX11 KO RPE-1 line. Number of cells counted n = 50 in triplicates. Scale bar: 10 µm.
    Antibody Rabbit Polyclonal Anti Atg16l1, supplied by MBL International, used in various techniques. Bioz Stars score: 90/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    rabbit polyclonal anti-atg16l1  (MBL Life science)
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    Autophagy is impaired in WABS patient and DDX11 KO RPE-1 cells. (A) Fibroblasts from three different WABS patients and from a healthy donor (HF1) were left in full medium or serum starved for 16 h. LC3-II level was assessed by western blot analysis. TUBA/tubulin was used as a loading control. (B) Graphs show the quantifications of LC3-II:TUBA. (C,D) fibroblasts from three different WABS patients and from a healthy donor (HF1) were treated with BAF A 1 for 16 h. LC3-II level was assessed by western blot analysis. TUBA was used as loading control. Graphs show the quantifications of LC3-II:TUBA. (E) RNA-seq analysis shows no difference in transcripts per kilobase million/TPM of autophagy genes ( <t>ATG16L1</t> , ATG5 , ATG12 , BECN1 , SQSTM1 ) between control (CTRL) and DDX11 KO ( DDX11 KO) RPE-1 cells. (F) Western blot analysis revealed no difference in the expression level of the indicated autophagy proteins between control (CTRL) and DDX11 KO ( DDX11 KO) RPE-1 cells. TUBA was used as loading control. (G,H) control (CTRL) and DDX11 KO ( DDX11 KO) RPE-1 cells were grown in full-medium (FM) or serum starved for 16 h (NO FBS). LC3-II was analyzed by western blot. TUBA was used as loading control. Graphs show the quantifications of LC3-II:TUBA. (I,J) Control (CTRL) and DDX11 KO ( DDX11 KO) RPE-1 cells were treated with DMSO or BAF A 1 for 6 h. LC3-II level was analyzed by western blot. TUBA was used as loading control. Graphs show the quantifications of LC3-II:TUBA. (K–M) DDX11 KO (KO) RPE-1 cells were transfected with a vector expressing DDX11-Flag wild-type protein (WT) or the DDX11-Flag K50R helicase-dead mutant (K50R) to perform rescue experiments. After 24 h, control (CTRL), DDX11 KO ( DDX11 KO) and DDX11-complemented DDX11 KO (KO + WT and KO + K50R) RPE-1 cells were treated with DMSO or BAF A 1 for 6 h. LC3 level (LC3-I and LC3-II) was analyzed by western blot. Graphs show the quantifications of LC3-II:TUBA. (N) Control (CTRL) and DDX11 KO ( DDX11 KO) RPE-1 cells were transfected with a vector expressing EGFP-HTT-74Q for 48 h to assess HTT mutant protein clearance. (O) Graph shows increased percentage of HTT-positive cells in the DDX11 KO RPE-1 line. Number of cells counted n = 50 in triplicates. Scale bar: 10 µm.
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    rabbit polyclonal anti atg16l1  (Cell Signaling Technology Inc)
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    Autophagy is impaired in WABS patient and DDX11 KO RPE-1 cells. (A) Fibroblasts from three different WABS patients and from a healthy donor (HF1) were left in full medium or serum starved for 16 h. LC3-II level was assessed by western blot analysis. TUBA/tubulin was used as a loading control. (B) Graphs show the quantifications of LC3-II:TUBA. (C,D) fibroblasts from three different WABS patients and from a healthy donor (HF1) were treated with BAF A 1 for 16 h. LC3-II level was assessed by western blot analysis. TUBA was used as loading control. Graphs show the quantifications of LC3-II:TUBA. (E) RNA-seq analysis shows no difference in transcripts per kilobase million/TPM of autophagy genes ( <t>ATG16L1</t> , ATG5 , ATG12 , BECN1 , SQSTM1 ) between control (CTRL) and DDX11 KO ( DDX11 KO) RPE-1 cells. (F) Western blot analysis revealed no difference in the expression level of the indicated autophagy proteins between control (CTRL) and DDX11 KO ( DDX11 KO) RPE-1 cells. TUBA was used as loading control. (G,H) control (CTRL) and DDX11 KO ( DDX11 KO) RPE-1 cells were grown in full-medium (FM) or serum starved for 16 h (NO FBS). LC3-II was analyzed by western blot. TUBA was used as loading control. Graphs show the quantifications of LC3-II:TUBA. (I,J) Control (CTRL) and DDX11 KO ( DDX11 KO) RPE-1 cells were treated with DMSO or BAF A 1 for 6 h. LC3-II level was analyzed by western blot. TUBA was used as loading control. Graphs show the quantifications of LC3-II:TUBA. (K–M) DDX11 KO (KO) RPE-1 cells were transfected with a vector expressing DDX11-Flag wild-type protein (WT) or the DDX11-Flag K50R helicase-dead mutant (K50R) to perform rescue experiments. After 24 h, control (CTRL), DDX11 KO ( DDX11 KO) and DDX11-complemented DDX11 KO (KO + WT and KO + K50R) RPE-1 cells were treated with DMSO or BAF A 1 for 6 h. LC3 level (LC3-I and LC3-II) was analyzed by western blot. Graphs show the quantifications of LC3-II:TUBA. (N) Control (CTRL) and DDX11 KO ( DDX11 KO) RPE-1 cells were transfected with a vector expressing EGFP-HTT-74Q for 48 h to assess HTT mutant protein clearance. (O) Graph shows increased percentage of HTT-positive cells in the DDX11 KO RPE-1 line. Number of cells counted n = 50 in triplicates. Scale bar: 10 µm.
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    polyclonal rabbit anti-atg16l1 antibody (catalog no.: pm040, rrid: ab_1278757)  (MBL Life science)
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    <t>ATG16L1</t> is required for bafilomycin A1 (Baf)-induced TDP-43 secretion. A and B , ATG16L1 WT or KO HeLa cells transfected with the mCherry-LC3 construct were exposed to 100 nM Baf for 24 h. Representative confocal live-cell images of ATG16L1 ( A ) WT or ( B ) KO cells transfected with the mCherry-LC3 ( red ) construct exposed to Baf. The plasma membrane (PM) was stained with PlasMem Bright Green ( green ). The image surrounded by a yellow square was magnified. Scale bar represents 5 μm. C – F , ATG16L1 WT or KO HeLa cells were exposed to 100 nM Baf for 24 h. The cell, P1, and P2 fractions were prepared as shown in the section. C , representative immunoblots (ATG16L1, TDP-43, TSG101, LC3, and α-tubulin) of the cell, P1, and P2 fractions are shown. The dashed square indicates bands of cleaved TDP-43 used for quantification. D – F , densitometric data on ( D ) full-length TDP-43, ( E ) cleaved TDP-43, and ( F ) TSG101 in the cell, P1, and P2 fractions were calculated from immunoblotting results. Relative signal intensities normalized against ATG16L1 WT cells are shown. In bar graphs, data are presented as means ± SD (N = 3). ∗ indicates p < 0.05 by the two-tailed unpaired t test. LC3, microtubule-associated proteins 1A/1B light chain 3B; TDP-43, transactive response DNA-binding protein 43 kDa; TSG101, tumor susceptibility gene 101 protein.
    Polyclonal Rabbit Anti Atg16l1 Antibody (Catalog No.: Pm040, Rrid: Ab 1278757), supplied by MBL Life science, used in various techniques. Bioz Stars score: 90/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    <t>ATG16L1</t> is required for bafilomycin A1 (Baf)-induced TDP-43 secretion. A and B , ATG16L1 WT or KO HeLa cells transfected with the mCherry-LC3 construct were exposed to 100 nM Baf for 24 h. Representative confocal live-cell images of ATG16L1 ( A ) WT or ( B ) KO cells transfected with the mCherry-LC3 ( red ) construct exposed to Baf. The plasma membrane (PM) was stained with PlasMem Bright Green ( green ). The image surrounded by a yellow square was magnified. Scale bar represents 5 μm. C – F , ATG16L1 WT or KO HeLa cells were exposed to 100 nM Baf for 24 h. The cell, P1, and P2 fractions were prepared as shown in the section. C , representative immunoblots (ATG16L1, TDP-43, TSG101, LC3, and α-tubulin) of the cell, P1, and P2 fractions are shown. The dashed square indicates bands of cleaved TDP-43 used for quantification. D – F , densitometric data on ( D ) full-length TDP-43, ( E ) cleaved TDP-43, and ( F ) TSG101 in the cell, P1, and P2 fractions were calculated from immunoblotting results. Relative signal intensities normalized against ATG16L1 WT cells are shown. In bar graphs, data are presented as means ± SD (N = 3). ∗ indicates p < 0.05 by the two-tailed unpaired t test. LC3, microtubule-associated proteins 1A/1B light chain 3B; TDP-43, transactive response DNA-binding protein 43 kDa; TSG101, tumor susceptibility gene 101 protein.
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    <t>ATG16L1</t> is required for bafilomycin A1 (Baf)-induced TDP-43 secretion. A and B , ATG16L1 WT or KO HeLa cells transfected with the mCherry-LC3 construct were exposed to 100 nM Baf for 24 h. Representative confocal live-cell images of ATG16L1 ( A ) WT or ( B ) KO cells transfected with the mCherry-LC3 ( red ) construct exposed to Baf. The plasma membrane (PM) was stained with PlasMem Bright Green ( green ). The image surrounded by a yellow square was magnified. Scale bar represents 5 μm. C – F , ATG16L1 WT or KO HeLa cells were exposed to 100 nM Baf for 24 h. The cell, P1, and P2 fractions were prepared as shown in the section. C , representative immunoblots (ATG16L1, TDP-43, TSG101, LC3, and α-tubulin) of the cell, P1, and P2 fractions are shown. The dashed square indicates bands of cleaved TDP-43 used for quantification. D – F , densitometric data on ( D ) full-length TDP-43, ( E ) cleaved TDP-43, and ( F ) TSG101 in the cell, P1, and P2 fractions were calculated from immunoblotting results. Relative signal intensities normalized against ATG16L1 WT cells are shown. In bar graphs, data are presented as means ± SD (N = 3). ∗ indicates p < 0.05 by the two-tailed unpaired t test. LC3, microtubule-associated proteins 1A/1B light chain 3B; TDP-43, transactive response DNA-binding protein 43 kDa; TSG101, tumor susceptibility gene 101 protein.
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    Image Search Results


    Autophagy is impaired in WABS patient and DDX11 KO RPE-1 cells. (A) Fibroblasts from three different WABS patients and from a healthy donor (HF1) were left in full medium or serum starved for 16 h. LC3-II level was assessed by western blot analysis. TUBA/tubulin was used as a loading control. (B) Graphs show the quantifications of LC3-II:TUBA. (C,D) fibroblasts from three different WABS patients and from a healthy donor (HF1) were treated with BAF A 1 for 16 h. LC3-II level was assessed by western blot analysis. TUBA was used as loading control. Graphs show the quantifications of LC3-II:TUBA. (E) RNA-seq analysis shows no difference in transcripts per kilobase million/TPM of autophagy genes ( ATG16L1 , ATG5 , ATG12 , BECN1 , SQSTM1 ) between control (CTRL) and DDX11 KO ( DDX11 KO) RPE-1 cells. (F) Western blot analysis revealed no difference in the expression level of the indicated autophagy proteins between control (CTRL) and DDX11 KO ( DDX11 KO) RPE-1 cells. TUBA was used as loading control. (G,H) control (CTRL) and DDX11 KO ( DDX11 KO) RPE-1 cells were grown in full-medium (FM) or serum starved for 16 h (NO FBS). LC3-II was analyzed by western blot. TUBA was used as loading control. Graphs show the quantifications of LC3-II:TUBA. (I,J) Control (CTRL) and DDX11 KO ( DDX11 KO) RPE-1 cells were treated with DMSO or BAF A 1 for 6 h. LC3-II level was analyzed by western blot. TUBA was used as loading control. Graphs show the quantifications of LC3-II:TUBA. (K–M) DDX11 KO (KO) RPE-1 cells were transfected with a vector expressing DDX11-Flag wild-type protein (WT) or the DDX11-Flag K50R helicase-dead mutant (K50R) to perform rescue experiments. After 24 h, control (CTRL), DDX11 KO ( DDX11 KO) and DDX11-complemented DDX11 KO (KO + WT and KO + K50R) RPE-1 cells were treated with DMSO or BAF A 1 for 6 h. LC3 level (LC3-I and LC3-II) was analyzed by western blot. Graphs show the quantifications of LC3-II:TUBA. (N) Control (CTRL) and DDX11 KO ( DDX11 KO) RPE-1 cells were transfected with a vector expressing EGFP-HTT-74Q for 48 h to assess HTT mutant protein clearance. (O) Graph shows increased percentage of HTT-positive cells in the DDX11 KO RPE-1 line. Number of cells counted n = 50 in triplicates. Scale bar: 10 µm.

    Journal: Autophagy

    Article Title: Evidence of an unprecedented cytoplasmic function of DDX11, the Warsaw breakage syndrome DNA helicase, in regulating autophagy

    doi: 10.1080/15548627.2025.2507617

    Figure Lengend Snippet: Autophagy is impaired in WABS patient and DDX11 KO RPE-1 cells. (A) Fibroblasts from three different WABS patients and from a healthy donor (HF1) were left in full medium or serum starved for 16 h. LC3-II level was assessed by western blot analysis. TUBA/tubulin was used as a loading control. (B) Graphs show the quantifications of LC3-II:TUBA. (C,D) fibroblasts from three different WABS patients and from a healthy donor (HF1) were treated with BAF A 1 for 16 h. LC3-II level was assessed by western blot analysis. TUBA was used as loading control. Graphs show the quantifications of LC3-II:TUBA. (E) RNA-seq analysis shows no difference in transcripts per kilobase million/TPM of autophagy genes ( ATG16L1 , ATG5 , ATG12 , BECN1 , SQSTM1 ) between control (CTRL) and DDX11 KO ( DDX11 KO) RPE-1 cells. (F) Western blot analysis revealed no difference in the expression level of the indicated autophagy proteins between control (CTRL) and DDX11 KO ( DDX11 KO) RPE-1 cells. TUBA was used as loading control. (G,H) control (CTRL) and DDX11 KO ( DDX11 KO) RPE-1 cells were grown in full-medium (FM) or serum starved for 16 h (NO FBS). LC3-II was analyzed by western blot. TUBA was used as loading control. Graphs show the quantifications of LC3-II:TUBA. (I,J) Control (CTRL) and DDX11 KO ( DDX11 KO) RPE-1 cells were treated with DMSO or BAF A 1 for 6 h. LC3-II level was analyzed by western blot. TUBA was used as loading control. Graphs show the quantifications of LC3-II:TUBA. (K–M) DDX11 KO (KO) RPE-1 cells were transfected with a vector expressing DDX11-Flag wild-type protein (WT) or the DDX11-Flag K50R helicase-dead mutant (K50R) to perform rescue experiments. After 24 h, control (CTRL), DDX11 KO ( DDX11 KO) and DDX11-complemented DDX11 KO (KO + WT and KO + K50R) RPE-1 cells were treated with DMSO or BAF A 1 for 6 h. LC3 level (LC3-I and LC3-II) was analyzed by western blot. Graphs show the quantifications of LC3-II:TUBA. (N) Control (CTRL) and DDX11 KO ( DDX11 KO) RPE-1 cells were transfected with a vector expressing EGFP-HTT-74Q for 48 h to assess HTT mutant protein clearance. (O) Graph shows increased percentage of HTT-positive cells in the DDX11 KO RPE-1 line. Number of cells counted n = 50 in triplicates. Scale bar: 10 µm.

    Article Snippet: The following antibodies were used: mouse monoclonal anti DDX11 (Santa Cruz Biotechnology, sc271711); mouse monoclonal anti-Flag antibody (Merck, F1804); rabbit polyclonal anti-LC3 (Novus Biologicals, NB100–2220); mouse monoclonal anti-LAMP1 (Cell Signaling Technology, 15665); rabbit polyclonal anti-SQSTM1 (MBL International, PM045); mouse monoclonal anti-TUBA/α-tubulin antibody (Merck, T6199); rabbit polyclonal anti-ATG16L1 antibody (MBL Life Sciences, PM040); rabbit monoclonal anti-ATG5 antibody (Cell Signaling Technology, 129945); rabbit polyclonal anti-BECN1/BECLIN1 antibody (Cell Signaling Technology, 3738S); rabbit polyclonal anti-AKT antibody (Cell Signaling Technology, 9272S); rabbit polyclonal anti-phospho-AKT (Ser473) antibody (Cell Signaling Technology, 9271 L); rabbit polyclonal anti-BRIP1/FANCJ antibody (Novus Biologicals, NBP1–31883); mouse monoclonal anti-histone H3 antibody (Abcam, ab24834); rabbit monoclonal anti-GFP antibody (Cell Signaling Technology, 2956); mouse and rabbit HRP-conjugated secondary antibody (Merck, A9044 and 12–348, respectively); Alexa Fluor 488 anti-rabbit antibody, Alexa Fluor 555 anti-mouse antibody and Alexa Fluor 633 anti-mouse antibody (Thermo Fisher Scientific, A-11008, A-21137, A-21146, respectively).

    Techniques: Western Blot, Control, RNA Sequencing, Expressing, Transfection, Plasmid Preparation, Mutagenesis

    DDX11 regulates ATG16L1 localization. (A,B) Control (CTRL), DDX11 KO ( DDX11 KO) and DDX11-complemented DDX11 KO ( DDX11 KO + WT DDX11 ) RPE-1 cells were transfected with ATG16L1-GFP for 24 h. Confocal imaging shows accumulation of ATG16L1 in the perinuclear area that was reverted by re-expressing the DDX11-Flag wild-type protein. Total cells analyzed: n = 50 per experiment, performed in triplicates. Scale bar: 10 µm. (C) Co-immunoprecipitation experiment using an anti-ATG16 antibody in control (CTRL) and DDX11 KO RPE-1 cells. Western blot analysis shows co-immunoprecipitation of ATG5-ATG12 in both control and DDX11 KO ( DDX11 KO) RPE-1 cells indicating that, in the absence of DDX11, the ATG12–ATG5-ATG16L1 complex formation is not affected. Asterisks denote the IgG heavy chains. (D) Control (CTRL), DDX11 KO ( DDX11 KO) and DDX11-complemented DDX11 KO ( DDX11 KO + WT DDX11 ) RPE-1 cells were transfected with a vector expressing GFP-ATG16L1 for 48 h. Co-immunoprecipitation experiment was performed using anti-GFP antibody conjugated beads. The indicated proteins were detected by western blot of the pulled-down sample using specific antibodies. (E) Control (CTRL), DDX11 KO ( DDX11 KO) and DDX11-complemented DDX11 KO ( DDX11 KO + WT DDX11 ) RPE-1 cells were transfected with GFP-ATG16L1 and mCherry-LC3 for 24 h. Then, cells were processed as described for panel A. Confocal imaging reveals that ATG16L1 and LC3 do not colocalize in DDX11 KO cells. This phenotype is reversed by re-expressing the DDX11-Flag wild-type protein. Scale bar: 10 µm.

    Journal: Autophagy

    Article Title: Evidence of an unprecedented cytoplasmic function of DDX11, the Warsaw breakage syndrome DNA helicase, in regulating autophagy

    doi: 10.1080/15548627.2025.2507617

    Figure Lengend Snippet: DDX11 regulates ATG16L1 localization. (A,B) Control (CTRL), DDX11 KO ( DDX11 KO) and DDX11-complemented DDX11 KO ( DDX11 KO + WT DDX11 ) RPE-1 cells were transfected with ATG16L1-GFP for 24 h. Confocal imaging shows accumulation of ATG16L1 in the perinuclear area that was reverted by re-expressing the DDX11-Flag wild-type protein. Total cells analyzed: n = 50 per experiment, performed in triplicates. Scale bar: 10 µm. (C) Co-immunoprecipitation experiment using an anti-ATG16 antibody in control (CTRL) and DDX11 KO RPE-1 cells. Western blot analysis shows co-immunoprecipitation of ATG5-ATG12 in both control and DDX11 KO ( DDX11 KO) RPE-1 cells indicating that, in the absence of DDX11, the ATG12–ATG5-ATG16L1 complex formation is not affected. Asterisks denote the IgG heavy chains. (D) Control (CTRL), DDX11 KO ( DDX11 KO) and DDX11-complemented DDX11 KO ( DDX11 KO + WT DDX11 ) RPE-1 cells were transfected with a vector expressing GFP-ATG16L1 for 48 h. Co-immunoprecipitation experiment was performed using anti-GFP antibody conjugated beads. The indicated proteins were detected by western blot of the pulled-down sample using specific antibodies. (E) Control (CTRL), DDX11 KO ( DDX11 KO) and DDX11-complemented DDX11 KO ( DDX11 KO + WT DDX11 ) RPE-1 cells were transfected with GFP-ATG16L1 and mCherry-LC3 for 24 h. Then, cells were processed as described for panel A. Confocal imaging reveals that ATG16L1 and LC3 do not colocalize in DDX11 KO cells. This phenotype is reversed by re-expressing the DDX11-Flag wild-type protein. Scale bar: 10 µm.

    Article Snippet: The following antibodies were used: mouse monoclonal anti DDX11 (Santa Cruz Biotechnology, sc271711); mouse monoclonal anti-Flag antibody (Merck, F1804); rabbit polyclonal anti-LC3 (Novus Biologicals, NB100–2220); mouse monoclonal anti-LAMP1 (Cell Signaling Technology, 15665); rabbit polyclonal anti-SQSTM1 (MBL International, PM045); mouse monoclonal anti-TUBA/α-tubulin antibody (Merck, T6199); rabbit polyclonal anti-ATG16L1 antibody (MBL Life Sciences, PM040); rabbit monoclonal anti-ATG5 antibody (Cell Signaling Technology, 129945); rabbit polyclonal anti-BECN1/BECLIN1 antibody (Cell Signaling Technology, 3738S); rabbit polyclonal anti-AKT antibody (Cell Signaling Technology, 9272S); rabbit polyclonal anti-phospho-AKT (Ser473) antibody (Cell Signaling Technology, 9271 L); rabbit polyclonal anti-BRIP1/FANCJ antibody (Novus Biologicals, NBP1–31883); mouse monoclonal anti-histone H3 antibody (Abcam, ab24834); rabbit monoclonal anti-GFP antibody (Cell Signaling Technology, 2956); mouse and rabbit HRP-conjugated secondary antibody (Merck, A9044 and 12–348, respectively); Alexa Fluor 488 anti-rabbit antibody, Alexa Fluor 555 anti-mouse antibody and Alexa Fluor 633 anti-mouse antibody (Thermo Fisher Scientific, A-11008, A-21137, A-21146, respectively).

    Techniques: Control, Transfection, Imaging, Expressing, Immunoprecipitation, Western Blot, Plasmid Preparation

    Schematic model describing a putative role of DDX11 in autophagy pathway regulation. DDX11 exerts its function as a DNA helicase in the nuclear compartment. Within the cytoplasm, DDX11 cooperates with SQSTM1 and ATG16L1 to promote phagophore formation as well as the autophagic flux (bold green and blue dotted arrows, respectively; left side ). Absence of DDX11 affects intracellular localization of the ATG16L1 autophagy precursor, which in turn impairs LC3 lipidation and autophagosome fusion with lysosomal compartment, ultimately reducing the autophagic flux (thin green and blue dotted arrows, respectively; right side ).

    Journal: Autophagy

    Article Title: Evidence of an unprecedented cytoplasmic function of DDX11, the Warsaw breakage syndrome DNA helicase, in regulating autophagy

    doi: 10.1080/15548627.2025.2507617

    Figure Lengend Snippet: Schematic model describing a putative role of DDX11 in autophagy pathway regulation. DDX11 exerts its function as a DNA helicase in the nuclear compartment. Within the cytoplasm, DDX11 cooperates with SQSTM1 and ATG16L1 to promote phagophore formation as well as the autophagic flux (bold green and blue dotted arrows, respectively; left side ). Absence of DDX11 affects intracellular localization of the ATG16L1 autophagy precursor, which in turn impairs LC3 lipidation and autophagosome fusion with lysosomal compartment, ultimately reducing the autophagic flux (thin green and blue dotted arrows, respectively; right side ).

    Article Snippet: The following antibodies were used: mouse monoclonal anti DDX11 (Santa Cruz Biotechnology, sc271711); mouse monoclonal anti-Flag antibody (Merck, F1804); rabbit polyclonal anti-LC3 (Novus Biologicals, NB100–2220); mouse monoclonal anti-LAMP1 (Cell Signaling Technology, 15665); rabbit polyclonal anti-SQSTM1 (MBL International, PM045); mouse monoclonal anti-TUBA/α-tubulin antibody (Merck, T6199); rabbit polyclonal anti-ATG16L1 antibody (MBL Life Sciences, PM040); rabbit monoclonal anti-ATG5 antibody (Cell Signaling Technology, 129945); rabbit polyclonal anti-BECN1/BECLIN1 antibody (Cell Signaling Technology, 3738S); rabbit polyclonal anti-AKT antibody (Cell Signaling Technology, 9272S); rabbit polyclonal anti-phospho-AKT (Ser473) antibody (Cell Signaling Technology, 9271 L); rabbit polyclonal anti-BRIP1/FANCJ antibody (Novus Biologicals, NBP1–31883); mouse monoclonal anti-histone H3 antibody (Abcam, ab24834); rabbit monoclonal anti-GFP antibody (Cell Signaling Technology, 2956); mouse and rabbit HRP-conjugated secondary antibody (Merck, A9044 and 12–348, respectively); Alexa Fluor 488 anti-rabbit antibody, Alexa Fluor 555 anti-mouse antibody and Alexa Fluor 633 anti-mouse antibody (Thermo Fisher Scientific, A-11008, A-21137, A-21146, respectively).

    Techniques:

    ATG16L1 is required for bafilomycin A1 (Baf)-induced TDP-43 secretion. A and B , ATG16L1 WT or KO HeLa cells transfected with the mCherry-LC3 construct were exposed to 100 nM Baf for 24 h. Representative confocal live-cell images of ATG16L1 ( A ) WT or ( B ) KO cells transfected with the mCherry-LC3 ( red ) construct exposed to Baf. The plasma membrane (PM) was stained with PlasMem Bright Green ( green ). The image surrounded by a yellow square was magnified. Scale bar represents 5 μm. C – F , ATG16L1 WT or KO HeLa cells were exposed to 100 nM Baf for 24 h. The cell, P1, and P2 fractions were prepared as shown in the section. C , representative immunoblots (ATG16L1, TDP-43, TSG101, LC3, and α-tubulin) of the cell, P1, and P2 fractions are shown. The dashed square indicates bands of cleaved TDP-43 used for quantification. D – F , densitometric data on ( D ) full-length TDP-43, ( E ) cleaved TDP-43, and ( F ) TSG101 in the cell, P1, and P2 fractions were calculated from immunoblotting results. Relative signal intensities normalized against ATG16L1 WT cells are shown. In bar graphs, data are presented as means ± SD (N = 3). ∗ indicates p < 0.05 by the two-tailed unpaired t test. LC3, microtubule-associated proteins 1A/1B light chain 3B; TDP-43, transactive response DNA-binding protein 43 kDa; TSG101, tumor susceptibility gene 101 protein.

    Journal: The Journal of Biological Chemistry

    Article Title: Dysregulation of the progranulin-driven autophagy-lysosomal pathway mediates secretion of the nuclear protein TDP-43

    doi: 10.1016/j.jbc.2023.105272

    Figure Lengend Snippet: ATG16L1 is required for bafilomycin A1 (Baf)-induced TDP-43 secretion. A and B , ATG16L1 WT or KO HeLa cells transfected with the mCherry-LC3 construct were exposed to 100 nM Baf for 24 h. Representative confocal live-cell images of ATG16L1 ( A ) WT or ( B ) KO cells transfected with the mCherry-LC3 ( red ) construct exposed to Baf. The plasma membrane (PM) was stained with PlasMem Bright Green ( green ). The image surrounded by a yellow square was magnified. Scale bar represents 5 μm. C – F , ATG16L1 WT or KO HeLa cells were exposed to 100 nM Baf for 24 h. The cell, P1, and P2 fractions were prepared as shown in the section. C , representative immunoblots (ATG16L1, TDP-43, TSG101, LC3, and α-tubulin) of the cell, P1, and P2 fractions are shown. The dashed square indicates bands of cleaved TDP-43 used for quantification. D – F , densitometric data on ( D ) full-length TDP-43, ( E ) cleaved TDP-43, and ( F ) TSG101 in the cell, P1, and P2 fractions were calculated from immunoblotting results. Relative signal intensities normalized against ATG16L1 WT cells are shown. In bar graphs, data are presented as means ± SD (N = 3). ∗ indicates p < 0.05 by the two-tailed unpaired t test. LC3, microtubule-associated proteins 1A/1B light chain 3B; TDP-43, transactive response DNA-binding protein 43 kDa; TSG101, tumor susceptibility gene 101 protein.

    Article Snippet: A monoclonal mouse anti-TDP-43 antibody (catalog no.: 60019-2-AP, Research Resource Identifier [RRID]: AB_2200520 ), polyclonal rabbit anti-C-terminal TDP-43 antibody (catalog no.: 12892-1-AP, RRID: AB_2200505 ), polyclonal rabbit anti-N-terminal TDP-43 antibody (catalog no.: 10782-2-AP, RRID: AB_615042 ), polyclonal rabbit anti-PGRN antibody (catalog no.: 18410-1-AP, RRID: AB_10598161 ), polyclonal rabbit anti-TSG101 antibody (catalog no.: 28283-1-AP, RRID: AB_2881104 ; catalog no.: 14497-1-AP, RRID: AB_2208090 ), polyclonal rabbit anti-ACTN4 antibody (catalog no.: 19096-1-AP, RRID: AB_10642150 ), polyclonal rabbit anti-ADAM10 antibody (catalog no.: 25900-1-AP, RRID: AB_2880291 ), and polyclonal rabbit anti-TFEB antibody (catalog no.: 13372-1-AP, RRID: AB_2919794 ) were purchased from ProteinTech; a monoclonal mouse anti-α-tubulin antibody (catalog no.: 70-102) and polyclonal rabbit anti-GFP antibody (catalog no.: 60-011) from BioAcademia; a monoclonal mouse anti-LC3 antibody (catalog no.: M186-3), normal rabbit immunoglobulin G (IgG) (catalog no.: PM035), polyclonal rabbit anti-Syntaxin-17 (human) antibody (catalog no.: PM076), and polyclonal rabbit anti-ATG16L1 antibody (catalog no.: PM040, RRID: AB_1278757 ) from MBL; a monoclonal mouse anti-p62 antibody (catalog no.: 610832, RRID: AB_398151 ) from BD Biosciences; a polyclonal sheep antimouse PGRN antibody (catalog no.: AF2557, RRID: AB_2114504 ) from R&D Systems; a monoclonal mouse anti-WIPI2 antibody (catalog no.: MCA5780GA, RRID: AB_10845951 ) from Bio-Rad; a biotin-conjugated horse antimouse IgG antibody (catalog no.: BA-2000, RRID: AB_2313581 ), biotin-conjugated goat anti-rabbit IgG antibody (catalog no.: BA-1000, RRID: AB_2313606 ), and biotin-conjugated rabbit antisheep IgG antibody (catalog no.: BA-6000, RRID: AB_2336217 ) from Vector Laboratories; and Alexa Fluor 568-conjugated donkey antimouse IgG (catalog no.: ab175472, RRID: AB_2636996 ), and Alexa Fluor 488–conjugated donkey antisheep IgG (catalog no.: ab150177, RRID: AB_2801320 ) from Abcam.

    Techniques: Transfection, Construct, Membrane, Staining, Western Blot, Two Tailed Test, Binding Assay

    The knockdown of GRN increases the secretion of TDP-43 and LC3-II in an autophagy-dependent manner. A and B , HeLa cells transfected with control or GRN siRNA for 48 h were stained with DALGreen and incubated for 2 h. A , representative live-cell images stained with DALGreen ( green ) are shown. Nuclei were stained with Hoechst 33342 ( blue ). Scale bar represents 50 μm. B , relative signal intensities normalized against cells transfected with control siRNA were calculated from three independent experiments including at least 78 cells. C and D , HeLa cells transfected with control or GRN siRNA for 48 h were stained with AcidiFluor ORANGE, and the acidity of lysosomes was compared. C , representative live-cell images stained with AcidiFluor ORANGE and photographed at a bright field are shown. Scale bar represents 100 μm. D , relative signal intensities normalized against cells transfected with control siRNA were calculated from three independent experiments including at least 627 cells. E – I , HeLa cells transfected with control or GRN siRNA were incubated for 72 h. The cell, P1, and P2 fractions were prepared as shown in the section. E , representative immunoblots (PGRN, TDP-43, TSG101, LC3, and α-tubulin) of the cell, P1, and P2 fractions are shown. The dashed square indicates the bands of cleaved TDP-43 used for quantification. F – I , densitometric data on ( F ) full-length TDP-43, ( G ) cleaved TDP-43, ( H ) TSG101, and ( I ) LC3-II were calculated from immunoblotting results. Relative signal intensities normalized against cells transfected with GRN siRNA are shown. J – M , ATG16L1 KO HeLa cells transfected with control or GRN siRNA were incubated for 72 h. The cell, P1, and P2 fractions were prepared as shown in the section. N is a sample from normal HeLa cells. J, representative immunoblots (PGRN, ATG16L1, TDP-43, TSG101, LC3, and α-tubulin) of the cell, P1, and P2 fractions are shown. The dashed square indicates the bands of cleaved TDP-43 used for quantification. K – M , densitometric data on ( K ) full-length TDP-43, ( L ) cleaved TDP-43, and ( M ) TSG101 were calculated from immunoblotting results. Relative signal intensities normalized against cells transfected with GRN siRNA are shown. In bar graphs, data are presented as means ± SD (N = 3). ∗ indicates p < 0.05 by the two-tailed unpaired t test or Mann–Whitney U test. LC3, microtubule-associated proteins 1A/1B light chain 3B; PGRN, progranulin; TDP-43, transactive response DNA-binding protein 43 kDa; TSG101, tumor susceptibility gene 101 protein.

    Journal: The Journal of Biological Chemistry

    Article Title: Dysregulation of the progranulin-driven autophagy-lysosomal pathway mediates secretion of the nuclear protein TDP-43

    doi: 10.1016/j.jbc.2023.105272

    Figure Lengend Snippet: The knockdown of GRN increases the secretion of TDP-43 and LC3-II in an autophagy-dependent manner. A and B , HeLa cells transfected with control or GRN siRNA for 48 h were stained with DALGreen and incubated for 2 h. A , representative live-cell images stained with DALGreen ( green ) are shown. Nuclei were stained with Hoechst 33342 ( blue ). Scale bar represents 50 μm. B , relative signal intensities normalized against cells transfected with control siRNA were calculated from three independent experiments including at least 78 cells. C and D , HeLa cells transfected with control or GRN siRNA for 48 h were stained with AcidiFluor ORANGE, and the acidity of lysosomes was compared. C , representative live-cell images stained with AcidiFluor ORANGE and photographed at a bright field are shown. Scale bar represents 100 μm. D , relative signal intensities normalized against cells transfected with control siRNA were calculated from three independent experiments including at least 627 cells. E – I , HeLa cells transfected with control or GRN siRNA were incubated for 72 h. The cell, P1, and P2 fractions were prepared as shown in the section. E , representative immunoblots (PGRN, TDP-43, TSG101, LC3, and α-tubulin) of the cell, P1, and P2 fractions are shown. The dashed square indicates the bands of cleaved TDP-43 used for quantification. F – I , densitometric data on ( F ) full-length TDP-43, ( G ) cleaved TDP-43, ( H ) TSG101, and ( I ) LC3-II were calculated from immunoblotting results. Relative signal intensities normalized against cells transfected with GRN siRNA are shown. J – M , ATG16L1 KO HeLa cells transfected with control or GRN siRNA were incubated for 72 h. The cell, P1, and P2 fractions were prepared as shown in the section. N is a sample from normal HeLa cells. J, representative immunoblots (PGRN, ATG16L1, TDP-43, TSG101, LC3, and α-tubulin) of the cell, P1, and P2 fractions are shown. The dashed square indicates the bands of cleaved TDP-43 used for quantification. K – M , densitometric data on ( K ) full-length TDP-43, ( L ) cleaved TDP-43, and ( M ) TSG101 were calculated from immunoblotting results. Relative signal intensities normalized against cells transfected with GRN siRNA are shown. In bar graphs, data are presented as means ± SD (N = 3). ∗ indicates p < 0.05 by the two-tailed unpaired t test or Mann–Whitney U test. LC3, microtubule-associated proteins 1A/1B light chain 3B; PGRN, progranulin; TDP-43, transactive response DNA-binding protein 43 kDa; TSG101, tumor susceptibility gene 101 protein.

    Article Snippet: A monoclonal mouse anti-TDP-43 antibody (catalog no.: 60019-2-AP, Research Resource Identifier [RRID]: AB_2200520 ), polyclonal rabbit anti-C-terminal TDP-43 antibody (catalog no.: 12892-1-AP, RRID: AB_2200505 ), polyclonal rabbit anti-N-terminal TDP-43 antibody (catalog no.: 10782-2-AP, RRID: AB_615042 ), polyclonal rabbit anti-PGRN antibody (catalog no.: 18410-1-AP, RRID: AB_10598161 ), polyclonal rabbit anti-TSG101 antibody (catalog no.: 28283-1-AP, RRID: AB_2881104 ; catalog no.: 14497-1-AP, RRID: AB_2208090 ), polyclonal rabbit anti-ACTN4 antibody (catalog no.: 19096-1-AP, RRID: AB_10642150 ), polyclonal rabbit anti-ADAM10 antibody (catalog no.: 25900-1-AP, RRID: AB_2880291 ), and polyclonal rabbit anti-TFEB antibody (catalog no.: 13372-1-AP, RRID: AB_2919794 ) were purchased from ProteinTech; a monoclonal mouse anti-α-tubulin antibody (catalog no.: 70-102) and polyclonal rabbit anti-GFP antibody (catalog no.: 60-011) from BioAcademia; a monoclonal mouse anti-LC3 antibody (catalog no.: M186-3), normal rabbit immunoglobulin G (IgG) (catalog no.: PM035), polyclonal rabbit anti-Syntaxin-17 (human) antibody (catalog no.: PM076), and polyclonal rabbit anti-ATG16L1 antibody (catalog no.: PM040, RRID: AB_1278757 ) from MBL; a monoclonal mouse anti-p62 antibody (catalog no.: 610832, RRID: AB_398151 ) from BD Biosciences; a polyclonal sheep antimouse PGRN antibody (catalog no.: AF2557, RRID: AB_2114504 ) from R&D Systems; a monoclonal mouse anti-WIPI2 antibody (catalog no.: MCA5780GA, RRID: AB_10845951 ) from Bio-Rad; a biotin-conjugated horse antimouse IgG antibody (catalog no.: BA-2000, RRID: AB_2313581 ), biotin-conjugated goat anti-rabbit IgG antibody (catalog no.: BA-1000, RRID: AB_2313606 ), and biotin-conjugated rabbit antisheep IgG antibody (catalog no.: BA-6000, RRID: AB_2336217 ) from Vector Laboratories; and Alexa Fluor 568-conjugated donkey antimouse IgG (catalog no.: ab175472, RRID: AB_2636996 ), and Alexa Fluor 488–conjugated donkey antisheep IgG (catalog no.: ab150177, RRID: AB_2801320 ) from Abcam.

    Techniques: Transfection, Staining, Incubation, Western Blot, Two Tailed Test, MANN-WHITNEY, Binding Assay