gfp mouse mab  (Cell Signaling Technology Inc)


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    Cell Signaling Technology Inc gfp mouse mab
    Gfp Mouse Mab, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/gfp mouse mab/product/Cell Signaling Technology Inc
    Average 86 stars, based on 1 article reviews
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    anti gfp 4b10 mouse monoclonal  (Cell Signaling Technology Inc)


    Bioz Manufacturer Symbol Cell Signaling Technology Inc manufactures this product  
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    Cell Signaling Technology Inc anti gfp 4b10 mouse monoclonal
    ( A ) Western blot images of CKPC-1 cells stably expressing <t>CYRI-B-p17-GFP</t> or GFP. KPC-1 and KPC-2 cell lines were used as control. Untransfected CKPC cells were also used as a control. Membranes were probed <t>for</t> <t>anti-GFP</t> (bottom blot) and anti-CYRI-B (top blot). GAPDH was used as loading control. Molecular weights are displayed on the side. ( B ) Representative immunofluorescence images of CKPC Cyrib knockout (KO) and rescued cells. Cells were seeded on fibronectin-coated coverslips, fixed and stained for F-actin (magenta), ArpC2 (cyan), and DAPI for nuclei (yellow). Scale bars, 20 μm. Yellow dotted boxes show the sites for the magnified images. Red arrows show the positive area for ArpC2 staining at the leading edge. Scale bars, 5 μm. Graph shows manual quantification of the number of cells presenting with lamellipodia (purple) or other protrusions (green) from (B). Mean ± S.D; paired t-test was performed in n=4. p-Value as indicated. ( C ) Quantification of cell area per cell from (B) based on the F-actin staining. Scatter plot here is presented as super plot and every independent biological repeat is coloured differently. Mean ± SEM; paired t-test was performed in n=4. p-Value as indicated. ( D ) Manual quantification of the length of the cell periphery showing strong ArpC2 accumulation, normalised to the total cell periphery. Scatter plot here is presented as a super plot and every independent biological repeat is coloured differently. Mean ± SEM; unpaired t-test was performed in n=3 (from a total of 30 cells). p-Value as indicated. ( E ) Manual quantification of the relative intensity of ArpC2 on the plasma membrane to cytoplasmic average intensity. Scatter plot here is presented as a super plot and every independent biological repeat is coloured differently. Mean ± SEM; unpaired t-test was performed in n=3. p-Value as indicated. Figure 4—figure supplement 2—source data 1. Scans of original western blots unlabelled and labelled to support . Figure 4—figure supplement 2—source data 2. Number of cells presenting with lamellipodia or other protrusions in CYRI-B knockout (GFP) vs CYRI-B-GFP rescued (CYRI-B-GFP) cells. Excel data and Prism analysis to support . Figure 4—figure supplement 2—source data 3. Area per cell in CYRI-B knockout (GFP) vs CYRI-B-GFP rescued (CYRI-B-GFP) cells ( , ). Figure 4—figure supplement 2—source data 4. Percentage of the periphery staining positive for ArpC2 in CYRI-B knockout (GFP) vs CYRI-B-GFP rescued (CYRI-B-GFP) cells. Excel data and Prism analysis to support . Figure 4—figure supplement 2—source data 5. Plasma membrane to cytoplasm relative intensity of ArpC2 in CYRI-B knockout (GFP) vs CYRI-B-GFP rescued (CYRI-B-GFP) cells. Excel data and Prism analysis to support .
    Anti Gfp 4b10 Mouse Monoclonal, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/anti gfp 4b10 mouse monoclonal/product/Cell Signaling Technology Inc
    Average 86 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    anti gfp 4b10 mouse monoclonal - by Bioz Stars, 2024-07
    86/100 stars

    Images

    1) Product Images from "CYRI-B-mediated macropinocytosis drives metastasis via lysophosphatidic acid receptor uptake"

    Article Title: CYRI-B-mediated macropinocytosis drives metastasis via lysophosphatidic acid receptor uptake

    Journal: eLife

    doi: 10.7554/eLife.83712

    ( A ) Western blot images of CKPC-1 cells stably expressing CYRI-B-p17-GFP or GFP. KPC-1 and KPC-2 cell lines were used as control. Untransfected CKPC cells were also used as a control. Membranes were probed for anti-GFP (bottom blot) and anti-CYRI-B (top blot). GAPDH was used as loading control. Molecular weights are displayed on the side. ( B ) Representative immunofluorescence images of CKPC Cyrib knockout (KO) and rescued cells. Cells were seeded on fibronectin-coated coverslips, fixed and stained for F-actin (magenta), ArpC2 (cyan), and DAPI for nuclei (yellow). Scale bars, 20 μm. Yellow dotted boxes show the sites for the magnified images. Red arrows show the positive area for ArpC2 staining at the leading edge. Scale bars, 5 μm. Graph shows manual quantification of the number of cells presenting with lamellipodia (purple) or other protrusions (green) from (B). Mean ± S.D; paired t-test was performed in n=4. p-Value as indicated. ( C ) Quantification of cell area per cell from (B) based on the F-actin staining. Scatter plot here is presented as super plot and every independent biological repeat is coloured differently. Mean ± SEM; paired t-test was performed in n=4. p-Value as indicated. ( D ) Manual quantification of the length of the cell periphery showing strong ArpC2 accumulation, normalised to the total cell periphery. Scatter plot here is presented as a super plot and every independent biological repeat is coloured differently. Mean ± SEM; unpaired t-test was performed in n=3 (from a total of 30 cells). p-Value as indicated. ( E ) Manual quantification of the relative intensity of ArpC2 on the plasma membrane to cytoplasmic average intensity. Scatter plot here is presented as a super plot and every independent biological repeat is coloured differently. Mean ± SEM; unpaired t-test was performed in n=3. p-Value as indicated. Figure 4—figure supplement 2—source data 1. Scans of original western blots unlabelled and labelled to support . Figure 4—figure supplement 2—source data 2. Number of cells presenting with lamellipodia or other protrusions in CYRI-B knockout (GFP) vs CYRI-B-GFP rescued (CYRI-B-GFP) cells. Excel data and Prism analysis to support . Figure 4—figure supplement 2—source data 3. Area per cell in CYRI-B knockout (GFP) vs CYRI-B-GFP rescued (CYRI-B-GFP) cells ( , ). Figure 4—figure supplement 2—source data 4. Percentage of the periphery staining positive for ArpC2 in CYRI-B knockout (GFP) vs CYRI-B-GFP rescued (CYRI-B-GFP) cells. Excel data and Prism analysis to support . Figure 4—figure supplement 2—source data 5. Plasma membrane to cytoplasm relative intensity of ArpC2 in CYRI-B knockout (GFP) vs CYRI-B-GFP rescued (CYRI-B-GFP) cells. Excel data and Prism analysis to support .
    Figure Legend Snippet: ( A ) Western blot images of CKPC-1 cells stably expressing CYRI-B-p17-GFP or GFP. KPC-1 and KPC-2 cell lines were used as control. Untransfected CKPC cells were also used as a control. Membranes were probed for anti-GFP (bottom blot) and anti-CYRI-B (top blot). GAPDH was used as loading control. Molecular weights are displayed on the side. ( B ) Representative immunofluorescence images of CKPC Cyrib knockout (KO) and rescued cells. Cells were seeded on fibronectin-coated coverslips, fixed and stained for F-actin (magenta), ArpC2 (cyan), and DAPI for nuclei (yellow). Scale bars, 20 μm. Yellow dotted boxes show the sites for the magnified images. Red arrows show the positive area for ArpC2 staining at the leading edge. Scale bars, 5 μm. Graph shows manual quantification of the number of cells presenting with lamellipodia (purple) or other protrusions (green) from (B). Mean ± S.D; paired t-test was performed in n=4. p-Value as indicated. ( C ) Quantification of cell area per cell from (B) based on the F-actin staining. Scatter plot here is presented as super plot and every independent biological repeat is coloured differently. Mean ± SEM; paired t-test was performed in n=4. p-Value as indicated. ( D ) Manual quantification of the length of the cell periphery showing strong ArpC2 accumulation, normalised to the total cell periphery. Scatter plot here is presented as a super plot and every independent biological repeat is coloured differently. Mean ± SEM; unpaired t-test was performed in n=3 (from a total of 30 cells). p-Value as indicated. ( E ) Manual quantification of the relative intensity of ArpC2 on the plasma membrane to cytoplasmic average intensity. Scatter plot here is presented as a super plot and every independent biological repeat is coloured differently. Mean ± SEM; unpaired t-test was performed in n=3. p-Value as indicated. Figure 4—figure supplement 2—source data 1. Scans of original western blots unlabelled and labelled to support . Figure 4—figure supplement 2—source data 2. Number of cells presenting with lamellipodia or other protrusions in CYRI-B knockout (GFP) vs CYRI-B-GFP rescued (CYRI-B-GFP) cells. Excel data and Prism analysis to support . Figure 4—figure supplement 2—source data 3. Area per cell in CYRI-B knockout (GFP) vs CYRI-B-GFP rescued (CYRI-B-GFP) cells ( , ). Figure 4—figure supplement 2—source data 4. Percentage of the periphery staining positive for ArpC2 in CYRI-B knockout (GFP) vs CYRI-B-GFP rescued (CYRI-B-GFP) cells. Excel data and Prism analysis to support . Figure 4—figure supplement 2—source data 5. Plasma membrane to cytoplasm relative intensity of ArpC2 in CYRI-B knockout (GFP) vs CYRI-B-GFP rescued (CYRI-B-GFP) cells. Excel data and Prism analysis to support .

    Techniques Used: Western Blot, Stable Transfection, Expressing, Control, Immunofluorescence, Knock-Out, Staining, Membrane


    Figure Legend Snippet:

    Techniques Used: Transfection, Construct, Isolation, Staining, Sequencing, Plasmid Preparation, RNAscope, CRISPR, SYBR Green Assay, Blocking Assay, Software

    anti gfp mouse monoclonal antibody  (Cell Signaling Technology Inc)


    Bioz Manufacturer Symbol Cell Signaling Technology Inc manufactures this product  
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    Cell Signaling Technology Inc anti gfp mouse monoclonal antibody
    ( A ) Scheme for expression of Cp OGA WT or Cp OGA DM in various Drosophila brain structures using different Gal4 drivers. ( B ) Immunostaining of adult Drosophila brains. Brains were stained with anti- O- GlcNAc antibody RL2 (red) to assess O- GlcNAcylation level, <t>and</t> <t>anti-GFP</t> (green) antibody to validate tissue-specific expression of Cp OGA. Nuclei were stained with DAPI (blue). Scale bar: 100 μm. ( C ) Quantification of fluorescent intensity of O- GlcNAc staining in Cp OGA WT or Cp OGA DM expressed brains. n = 4. ( D ) Immunostaining of adult Drosophila brains. Outlined areas indicate the cell bodies of Kenyon cells in mushroom body. Scale bar: 100 μm. ( E ) Quantification of relative fluorescent intensity of O- GlcNAc staining in Cp OGA WT or Cp OGA DM expressed brain structures. n = 8. ( F ) A compilation of performance index in learning test of the indicated flies expressing either Cp OGA WT or Cp OGA DM . n = 6-8. ( G ) A compilation of learning performance index of flies expressing Cp OGA WT or Cp OGA DM only in the mushroom body at adult stage. n = 6. Each datapoint represents an independent experiment with approximately 200 flies. p- values were determined by unpaired t -test, and the stars indicate significant differences (***p<0.001, **p<0.01 and ns, not significant, p≥0.05). Error bars represent SD. Figure 1—source data 1. Excel spreadsheet containing source data used to generate .
    Anti Gfp Mouse Monoclonal 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/result/anti gfp mouse monoclonal antibody/product/Cell Signaling Technology Inc
    Average 86 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    anti gfp mouse monoclonal antibody - by Bioz Stars, 2024-07
    86/100 stars

    Images

    1) Product Images from "Tissue-specific O- GlcNAcylation profiling identifies substrates in translational machinery in Drosophila mushroom body contributing to olfactory learning"

    Article Title: Tissue-specific O- GlcNAcylation profiling identifies substrates in translational machinery in Drosophila mushroom body contributing to olfactory learning

    Journal: eLife

    doi: 10.7554/eLife.91269

    ( A ) Scheme for expression of Cp OGA WT or Cp OGA DM in various Drosophila brain structures using different Gal4 drivers. ( B ) Immunostaining of adult Drosophila brains. Brains were stained with anti- O- GlcNAc antibody RL2 (red) to assess O- GlcNAcylation level, and anti-GFP (green) antibody to validate tissue-specific expression of Cp OGA. Nuclei were stained with DAPI (blue). Scale bar: 100 μm. ( C ) Quantification of fluorescent intensity of O- GlcNAc staining in Cp OGA WT or Cp OGA DM expressed brains. n = 4. ( D ) Immunostaining of adult Drosophila brains. Outlined areas indicate the cell bodies of Kenyon cells in mushroom body. Scale bar: 100 μm. ( E ) Quantification of relative fluorescent intensity of O- GlcNAc staining in Cp OGA WT or Cp OGA DM expressed brain structures. n = 8. ( F ) A compilation of performance index in learning test of the indicated flies expressing either Cp OGA WT or Cp OGA DM . n = 6-8. ( G ) A compilation of learning performance index of flies expressing Cp OGA WT or Cp OGA DM only in the mushroom body at adult stage. n = 6. Each datapoint represents an independent experiment with approximately 200 flies. p- values were determined by unpaired t -test, and the stars indicate significant differences (***p<0.001, **p<0.01 and ns, not significant, p≥0.05). Error bars represent SD. Figure 1—source data 1. Excel spreadsheet containing source data used to generate .
    Figure Legend Snippet: ( A ) Scheme for expression of Cp OGA WT or Cp OGA DM in various Drosophila brain structures using different Gal4 drivers. ( B ) Immunostaining of adult Drosophila brains. Brains were stained with anti- O- GlcNAc antibody RL2 (red) to assess O- GlcNAcylation level, and anti-GFP (green) antibody to validate tissue-specific expression of Cp OGA. Nuclei were stained with DAPI (blue). Scale bar: 100 μm. ( C ) Quantification of fluorescent intensity of O- GlcNAc staining in Cp OGA WT or Cp OGA DM expressed brains. n = 4. ( D ) Immunostaining of adult Drosophila brains. Outlined areas indicate the cell bodies of Kenyon cells in mushroom body. Scale bar: 100 μm. ( E ) Quantification of relative fluorescent intensity of O- GlcNAc staining in Cp OGA WT or Cp OGA DM expressed brain structures. n = 8. ( F ) A compilation of performance index in learning test of the indicated flies expressing either Cp OGA WT or Cp OGA DM . n = 6-8. ( G ) A compilation of learning performance index of flies expressing Cp OGA WT or Cp OGA DM only in the mushroom body at adult stage. n = 6. Each datapoint represents an independent experiment with approximately 200 flies. p- values were determined by unpaired t -test, and the stars indicate significant differences (***p<0.001, **p<0.01 and ns, not significant, p≥0.05). Error bars represent SD. Figure 1—source data 1. Excel spreadsheet containing source data used to generate .

    Techniques Used: Expressing, Immunostaining, Staining

    ( A ) Heatmaps showing the mRNA levels (upper) and the normalized O- GlcNAc levels (lower) of the identified ribosomal candidates in different brain regions. ( B ) Immunoprecipitation of ribosomes using FLAG-tagged RpL13A. The expression of RpL13A-FLAG was validated by immunoblotting with anti-FLAG antibody. Ribosomal proteins were visualized using silver staining, and O- GlcNAcylation of ribosomes was analyzed by immunoblotting with anti- O- GlcNAc antibody RL2. ( C ) A compilation of the performance index of the indicated flies in the learning test. Learning defect of flies expressing Cp OGA WT was corrected by selective expression of dMyc in mushroom body. n = 6-7. Each datapoint represents an independent experiment with approximately 200 flies. ( D ) Ex vivo measurement of protein synthesis in mushroom body using the O-propargyl-puromycin (OPP) assay. Brains from the indicated flies were stained with anti-GFP (green) antibody to validate Cp OGA expression, and OPP (gray) to quantify protein synthesis. Nuclei were visualized with DAPI (blue). Outlined areas indicate the cell bodies of Kenyon cells of mushroom body. Scale bar: 100 μm. ( E ) Quantification of relative OPP fluorescent intensity in mushroom body regions. n = 8-12. p -values were determined by unpaired t -test, the stars indicate significant differences (***p<0.001, **p<0.01, *p<0.05). Error bars represent SD. Figure 4—source data 1. Raw data of all western blots for . Figure 4—source data 2. Complete and uncropped membranes of all western blots for . Figure 4—source data 3. Excel spreadsheet containing source data used to generate .
    Figure Legend Snippet: ( A ) Heatmaps showing the mRNA levels (upper) and the normalized O- GlcNAc levels (lower) of the identified ribosomal candidates in different brain regions. ( B ) Immunoprecipitation of ribosomes using FLAG-tagged RpL13A. The expression of RpL13A-FLAG was validated by immunoblotting with anti-FLAG antibody. Ribosomal proteins were visualized using silver staining, and O- GlcNAcylation of ribosomes was analyzed by immunoblotting with anti- O- GlcNAc antibody RL2. ( C ) A compilation of the performance index of the indicated flies in the learning test. Learning defect of flies expressing Cp OGA WT was corrected by selective expression of dMyc in mushroom body. n = 6-7. Each datapoint represents an independent experiment with approximately 200 flies. ( D ) Ex vivo measurement of protein synthesis in mushroom body using the O-propargyl-puromycin (OPP) assay. Brains from the indicated flies were stained with anti-GFP (green) antibody to validate Cp OGA expression, and OPP (gray) to quantify protein synthesis. Nuclei were visualized with DAPI (blue). Outlined areas indicate the cell bodies of Kenyon cells of mushroom body. Scale bar: 100 μm. ( E ) Quantification of relative OPP fluorescent intensity in mushroom body regions. n = 8-12. p -values were determined by unpaired t -test, the stars indicate significant differences (***p<0.001, **p<0.01, *p<0.05). Error bars represent SD. Figure 4—source data 1. Raw data of all western blots for . Figure 4—source data 2. Complete and uncropped membranes of all western blots for . Figure 4—source data 3. Excel spreadsheet containing source data used to generate .

    Techniques Used: Immunoprecipitation, Expressing, Western Blot, Silver Staining, Ex Vivo, Staining


    Figure Legend Snippet:

    Techniques Used: Silver Staining, Protease Inhibitor, Magnetic Beads, SYBR Green Assay, Sequencing, Modification, Software, Microscopy

    gfp 4b10 mouse mab  (Cell Signaling Technology Inc)


    Bioz Manufacturer Symbol Cell Signaling Technology Inc manufactures this product  
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    Cell Signaling Technology Inc gfp 4b10 mouse mab
    Gfp 4b10 Mouse Mab, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/gfp 4b10 mouse mab/product/Cell Signaling Technology Inc
    Average 86 stars, based on 1 article reviews
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    gfp 4b10 mouse mab - by Bioz Stars, 2024-07
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    gfp mouse mab  (Cell Signaling Technology Inc)


    Bioz Manufacturer Symbol Cell Signaling Technology Inc manufactures this product  
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    Cell Signaling Technology Inc gfp mouse mab
    Gfp Mouse Mab, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/gfp mouse mab/product/Cell Signaling Technology Inc
    Average 86 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    gfp mouse mab - by Bioz Stars, 2024-07
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    gfp 4b10 mouse mab  (Cell Signaling Technology Inc)


    Bioz Manufacturer Symbol Cell Signaling Technology Inc manufactures this product  
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    Cell Signaling Technology Inc gfp 4b10 mouse mab
    Gfp 4b10 Mouse Mab, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/gfp 4b10 mouse mab/product/Cell Signaling Technology Inc
    Average 86 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    gfp 4b10 mouse mab - by Bioz Stars, 2024-07
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    gfp mouse monoclonal primary antibody  (Cell Signaling Technology Inc)


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    Cell Signaling Technology Inc gfp mouse monoclonal primary antibody
    ( A ) Construct design to generate HEK293 FRT stable cells harboring the KCNJ13 W53X allele. ( B ) Chromatogram generated from HEK293 FRT stable cells showing the W53X codon marked in the red box and the downward black arrow indicating the specific nucleotide change (G>A).( C ) Schematic of the hKCNJ13 locus highlighting the mutation c.158G>A (blue box marked with asterisk) and position of the W53X targeting sgRNA (black line) with TGG PAM (red line). ( D ) Base-editing efficiencies are shown as the percentages of sequencing reads with the corrected WT allele (and no other silent changes, bystander edits, or indels) in HEK293 W53X cells following electroporation of ABE8e protein+sgRNA (RNP) or ABE8e mRNA+sgRNA ( n = 3). Markers (diamonds) represent the individual biological replicates ( n = 3), and error bars represent SEM by 2-tailed Student’s t test. ( E ) Percentages of sequencing reads with indels in ABE8e RNP– and ABE8e mRNA–treated stable cells ( n = 3). Markers (diamonds) represent the individual biological replicates ( n = 3), and error bars represent SEM by 2-tailed Student’s t test. ( F <t>)</t> <t>Kir7.1</t> expression in ABE8e mRNA–treated cells assessed by immunocytochemistry. <t>GFP</t> primary antibody was used to enhance the endogenous signal. DAPI was used to stain the nucleus. Scale bars: 50 μm. White arrows mark membrane localization in cells.
    Gfp Mouse Monoclonal Primary 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/result/gfp mouse monoclonal primary antibody/product/Cell Signaling Technology Inc
    Average 86 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    gfp mouse monoclonal primary antibody - by Bioz Stars, 2024-07
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    Images

    1) Product Images from "Nonviral base editing of KCNJ13 mutation preserves vision in a model of inherited retinal channelopathy"

    Article Title: Nonviral base editing of KCNJ13 mutation preserves vision in a model of inherited retinal channelopathy

    Journal: The Journal of Clinical Investigation

    doi: 10.1172/JCI171356

    ( A ) Construct design to generate HEK293 FRT stable cells harboring the KCNJ13 W53X allele. ( B ) Chromatogram generated from HEK293 FRT stable cells showing the W53X codon marked in the red box and the downward black arrow indicating the specific nucleotide change (G>A).( C ) Schematic of the hKCNJ13 locus highlighting the mutation c.158G>A (blue box marked with asterisk) and position of the W53X targeting sgRNA (black line) with TGG PAM (red line). ( D ) Base-editing efficiencies are shown as the percentages of sequencing reads with the corrected WT allele (and no other silent changes, bystander edits, or indels) in HEK293 W53X cells following electroporation of ABE8e protein+sgRNA (RNP) or ABE8e mRNA+sgRNA ( n = 3). Markers (diamonds) represent the individual biological replicates ( n = 3), and error bars represent SEM by 2-tailed Student’s t test. ( E ) Percentages of sequencing reads with indels in ABE8e RNP– and ABE8e mRNA–treated stable cells ( n = 3). Markers (diamonds) represent the individual biological replicates ( n = 3), and error bars represent SEM by 2-tailed Student’s t test. ( F ) Kir7.1 expression in ABE8e mRNA–treated cells assessed by immunocytochemistry. GFP primary antibody was used to enhance the endogenous signal. DAPI was used to stain the nucleus. Scale bars: 50 μm. White arrows mark membrane localization in cells.
    Figure Legend Snippet: ( A ) Construct design to generate HEK293 FRT stable cells harboring the KCNJ13 W53X allele. ( B ) Chromatogram generated from HEK293 FRT stable cells showing the W53X codon marked in the red box and the downward black arrow indicating the specific nucleotide change (G>A).( C ) Schematic of the hKCNJ13 locus highlighting the mutation c.158G>A (blue box marked with asterisk) and position of the W53X targeting sgRNA (black line) with TGG PAM (red line). ( D ) Base-editing efficiencies are shown as the percentages of sequencing reads with the corrected WT allele (and no other silent changes, bystander edits, or indels) in HEK293 W53X cells following electroporation of ABE8e protein+sgRNA (RNP) or ABE8e mRNA+sgRNA ( n = 3). Markers (diamonds) represent the individual biological replicates ( n = 3), and error bars represent SEM by 2-tailed Student’s t test. ( E ) Percentages of sequencing reads with indels in ABE8e RNP– and ABE8e mRNA–treated stable cells ( n = 3). Markers (diamonds) represent the individual biological replicates ( n = 3), and error bars represent SEM by 2-tailed Student’s t test. ( F ) Kir7.1 expression in ABE8e mRNA–treated cells assessed by immunocytochemistry. GFP primary antibody was used to enhance the endogenous signal. DAPI was used to stain the nucleus. Scale bars: 50 μm. White arrows mark membrane localization in cells.

    Techniques Used: Construct, Generated, Mutagenesis, Sequencing, Electroporation, Expressing, Immunocytochemistry, Staining, Membrane

    gfp mouse monoclonal primary antibody  (Cell Signaling Technology Inc)


    Bioz Manufacturer Symbol Cell Signaling Technology Inc manufactures this product  
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    Cell Signaling Technology Inc gfp mouse monoclonal primary antibody
    Gfp Mouse Monoclonal Primary 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/result/gfp mouse monoclonal primary antibody/product/Cell Signaling Technology Inc
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    gfp mouse monoclonal primary antibody - by Bioz Stars, 2024-07
    86/100 stars

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    mouse monoclonal anti gfp  (Cell Signaling Technology Inc)


    Bioz Manufacturer Symbol Cell Signaling Technology Inc manufactures this product  
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    Cell Signaling Technology Inc mouse monoclonal anti gfp
    (A) Schematic of Mup1 sorting. (B) <t>Mup1-GFP</t> localization after methionine stimulation. (C) Western blotting analysis of Mup1 sorting. Cell lysates were analyzed by immunoblotting <t>using</t> <t>anti-GFP</t> and anti-G6PDH antibodies. Vacuole delivery of Mup1-GFP yields protease-resistant GFP fragments (GFP’). (D) Quantification of Mup1-GFP processing. (E) GFP-CPS localization. (F) Quantification of GFP-CPS localization of each category. (G) Schematic of retromer-mediated Vps10 recycling. (H) Vps10-GFP localization in WT, vps35 Δ (retromer), and vps13 Δ cells. (I) Quantification of Vps10-GFP localization. Scale bar: 1 µm.
    Mouse Monoclonal Anti Gfp, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/mouse monoclonal anti gfp/product/Cell Signaling Technology Inc
    Average 86 stars, based on 1 article reviews
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    Images

    1) Product Images from "A role for Vps13-mediated lipid transfer at the ER-endosome contact site in ESCRT-mediated sorting"

    Article Title: A role for Vps13-mediated lipid transfer at the ER-endosome contact site in ESCRT-mediated sorting

    Journal: bioRxiv

    doi: 10.1101/2023.07.06.547997

    (A) Schematic of Mup1 sorting. (B) Mup1-GFP localization after methionine stimulation. (C) Western blotting analysis of Mup1 sorting. Cell lysates were analyzed by immunoblotting using anti-GFP and anti-G6PDH antibodies. Vacuole delivery of Mup1-GFP yields protease-resistant GFP fragments (GFP’). (D) Quantification of Mup1-GFP processing. (E) GFP-CPS localization. (F) Quantification of GFP-CPS localization of each category. (G) Schematic of retromer-mediated Vps10 recycling. (H) Vps10-GFP localization in WT, vps35 Δ (retromer), and vps13 Δ cells. (I) Quantification of Vps10-GFP localization. Scale bar: 1 µm.
    Figure Legend Snippet: (A) Schematic of Mup1 sorting. (B) Mup1-GFP localization after methionine stimulation. (C) Western blotting analysis of Mup1 sorting. Cell lysates were analyzed by immunoblotting using anti-GFP and anti-G6PDH antibodies. Vacuole delivery of Mup1-GFP yields protease-resistant GFP fragments (GFP’). (D) Quantification of Mup1-GFP processing. (E) GFP-CPS localization. (F) Quantification of GFP-CPS localization of each category. (G) Schematic of retromer-mediated Vps10 recycling. (H) Vps10-GFP localization in WT, vps35 Δ (retromer), and vps13 Δ cells. (I) Quantification of Vps10-GFP localization. Scale bar: 1 µm.

    Techniques Used: Western Blot

    (A) Localization of Vps13-GFP. Scale bar: 1 µm. (B) Quantification of Vps13-GFP colocalizing with mCherry-Vps21. (C) Localization of Vps13 ΔC -GFP (residues 1-1851). Scale bar: 1 µm. (D) Quantification of Vps13-GFP puncta localization. (E) The localization of Vps13 N -GFP (residues 1-39). Scale bar: 1 µm. (F) Line scan analysis for the region highlighted by the yellow dash line in E. (G) Vps13-GFP localization at the ER-endosome contact site. Vps13-GFP, DsRed-HDEL (ER), and Mup1-BFP (endosome) expressing cells were stimulated with methionine. Scale bar: 1 µm. (H) Live cell-imaging analysis of the ER (GFP-HDEL) and endosome (mCherry-Vps21). Scale bar: 500 nm. (I) ER diameters per 100 nm interval starting at the endosome contact along 500 nm length of ER. (J and K) Mup1-GFP processing in vps13 mutants after methionine stimulation. Cell lysates were analyzed by immunoblotting using anti-GFP and anti-G6PDH antibodies.
    Figure Legend Snippet: (A) Localization of Vps13-GFP. Scale bar: 1 µm. (B) Quantification of Vps13-GFP colocalizing with mCherry-Vps21. (C) Localization of Vps13 ΔC -GFP (residues 1-1851). Scale bar: 1 µm. (D) Quantification of Vps13-GFP puncta localization. (E) The localization of Vps13 N -GFP (residues 1-39). Scale bar: 1 µm. (F) Line scan analysis for the region highlighted by the yellow dash line in E. (G) Vps13-GFP localization at the ER-endosome contact site. Vps13-GFP, DsRed-HDEL (ER), and Mup1-BFP (endosome) expressing cells were stimulated with methionine. Scale bar: 1 µm. (H) Live cell-imaging analysis of the ER (GFP-HDEL) and endosome (mCherry-Vps21). Scale bar: 500 nm. (I) ER diameters per 100 nm interval starting at the endosome contact along 500 nm length of ER. (J and K) Mup1-GFP processing in vps13 mutants after methionine stimulation. Cell lysates were analyzed by immunoblotting using anti-GFP and anti-G6PDH antibodies.

    Techniques Used: Expressing, Live Cell Imaging, Western Blot

    (A) Schematic of ESCRT-mediated sorting at the endosome. (B) Can1-FKBP processing after rapamycin treatments. Cell lysates were analyzed by immunoblotting using anti-GFP and anti-G6PDH antibodies. (C) GFP-Vps27 (ESCRT-0) localization. (D) Snf7-GFP (ESCRT-III) localization. Scale bar: 1 µm.
    Figure Legend Snippet: (A) Schematic of ESCRT-mediated sorting at the endosome. (B) Can1-FKBP processing after rapamycin treatments. Cell lysates were analyzed by immunoblotting using anti-GFP and anti-G6PDH antibodies. (C) GFP-Vps27 (ESCRT-0) localization. (D) Snf7-GFP (ESCRT-III) localization. Scale bar: 1 µm.

    Techniques Used: Western Blot

    (A) The ubiquitination of Mup1-GFP. Mup1-GFP expressing cells were stimulated with methionine for 15 min and then immunoprecipitated under denatured conditions. The immunoprecipitated (IP) products were analyzed by immunoblotting using anti-GFP and anti-ubiquitin antibodies. (B) Schematic of a rapamycin-dependent degradation system for Can1. (C) Can1-FKBP localization. Rapamycin-insensitive mutant cells ( tor1-1 , fpr1 Δ) expressing Can1-GFP-2xFKBP (Can1-FKBP) and FRB-3xUb were treated with rapamycin. (D) Quantification of Can-FKBP sorting. (E) Vps4-GFP localization. (F) Quantification of endosomal Vps4-GFP localization. (G) Schematic of Mup1-pHluorin-mCherry sorting. (H) Mup1-pHluorin-mCherry localization. Mup1-pHluorin-mCherry expressing vam7 ts and vam7 t vps13 Δ cells were incubated at 37°C and stimulated with methionine. (I) pHluorin fluorescence of Mup1-pHluorin-mCherry. Scale bar: 1 µm.
    Figure Legend Snippet: (A) The ubiquitination of Mup1-GFP. Mup1-GFP expressing cells were stimulated with methionine for 15 min and then immunoprecipitated under denatured conditions. The immunoprecipitated (IP) products were analyzed by immunoblotting using anti-GFP and anti-ubiquitin antibodies. (B) Schematic of a rapamycin-dependent degradation system for Can1. (C) Can1-FKBP localization. Rapamycin-insensitive mutant cells ( tor1-1 , fpr1 Δ) expressing Can1-GFP-2xFKBP (Can1-FKBP) and FRB-3xUb were treated with rapamycin. (D) Quantification of Can-FKBP sorting. (E) Vps4-GFP localization. (F) Quantification of endosomal Vps4-GFP localization. (G) Schematic of Mup1-pHluorin-mCherry sorting. (H) Mup1-pHluorin-mCherry localization. Mup1-pHluorin-mCherry expressing vam7 ts and vam7 t vps13 Δ cells were incubated at 37°C and stimulated with methionine. (I) pHluorin fluorescence of Mup1-pHluorin-mCherry. Scale bar: 1 µm.

    Techniques Used: Expressing, Immunoprecipitation, Western Blot, Mutagenesis, Incubation, Fluorescence

    mouse monoclonal anti gfp  (Cell Signaling Technology Inc)


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    Cell Signaling Technology Inc mouse monoclonal anti gfp
    Mouse Monoclonal Anti Gfp, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Cell Signaling Technology Inc gfp mouse mab
    Gfp Mouse Mab, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Cell Signaling Technology Inc anti gfp 4b10 mouse monoclonal
    ( A ) Western blot images of CKPC-1 cells stably expressing <t>CYRI-B-p17-GFP</t> or GFP. KPC-1 and KPC-2 cell lines were used as control. Untransfected CKPC cells were also used as a control. Membranes were probed <t>for</t> <t>anti-GFP</t> (bottom blot) and anti-CYRI-B (top blot). GAPDH was used as loading control. Molecular weights are displayed on the side. ( B ) Representative immunofluorescence images of CKPC Cyrib knockout (KO) and rescued cells. Cells were seeded on fibronectin-coated coverslips, fixed and stained for F-actin (magenta), ArpC2 (cyan), and DAPI for nuclei (yellow). Scale bars, 20 μm. Yellow dotted boxes show the sites for the magnified images. Red arrows show the positive area for ArpC2 staining at the leading edge. Scale bars, 5 μm. Graph shows manual quantification of the number of cells presenting with lamellipodia (purple) or other protrusions (green) from (B). Mean ± S.D; paired t-test was performed in n=4. p-Value as indicated. ( C ) Quantification of cell area per cell from (B) based on the F-actin staining. Scatter plot here is presented as super plot and every independent biological repeat is coloured differently. Mean ± SEM; paired t-test was performed in n=4. p-Value as indicated. ( D ) Manual quantification of the length of the cell periphery showing strong ArpC2 accumulation, normalised to the total cell periphery. Scatter plot here is presented as a super plot and every independent biological repeat is coloured differently. Mean ± SEM; unpaired t-test was performed in n=3 (from a total of 30 cells). p-Value as indicated. ( E ) Manual quantification of the relative intensity of ArpC2 on the plasma membrane to cytoplasmic average intensity. Scatter plot here is presented as a super plot and every independent biological repeat is coloured differently. Mean ± SEM; unpaired t-test was performed in n=3. p-Value as indicated. Figure 4—figure supplement 2—source data 1. Scans of original western blots unlabelled and labelled to support . Figure 4—figure supplement 2—source data 2. Number of cells presenting with lamellipodia or other protrusions in CYRI-B knockout (GFP) vs CYRI-B-GFP rescued (CYRI-B-GFP) cells. Excel data and Prism analysis to support . Figure 4—figure supplement 2—source data 3. Area per cell in CYRI-B knockout (GFP) vs CYRI-B-GFP rescued (CYRI-B-GFP) cells ( , ). Figure 4—figure supplement 2—source data 4. Percentage of the periphery staining positive for ArpC2 in CYRI-B knockout (GFP) vs CYRI-B-GFP rescued (CYRI-B-GFP) cells. Excel data and Prism analysis to support . Figure 4—figure supplement 2—source data 5. Plasma membrane to cytoplasm relative intensity of ArpC2 in CYRI-B knockout (GFP) vs CYRI-B-GFP rescued (CYRI-B-GFP) cells. Excel data and Prism analysis to support .
    Anti Gfp 4b10 Mouse Monoclonal, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Cell Signaling Technology Inc anti gfp mouse monoclonal antibody
    ( A ) Scheme for expression of Cp OGA WT or Cp OGA DM in various Drosophila brain structures using different Gal4 drivers. ( B ) Immunostaining of adult Drosophila brains. Brains were stained with anti- O- GlcNAc antibody RL2 (red) to assess O- GlcNAcylation level, <t>and</t> <t>anti-GFP</t> (green) antibody to validate tissue-specific expression of Cp OGA. Nuclei were stained with DAPI (blue). Scale bar: 100 μm. ( C ) Quantification of fluorescent intensity of O- GlcNAc staining in Cp OGA WT or Cp OGA DM expressed brains. n = 4. ( D ) Immunostaining of adult Drosophila brains. Outlined areas indicate the cell bodies of Kenyon cells in mushroom body. Scale bar: 100 μm. ( E ) Quantification of relative fluorescent intensity of O- GlcNAc staining in Cp OGA WT or Cp OGA DM expressed brain structures. n = 8. ( F ) A compilation of performance index in learning test of the indicated flies expressing either Cp OGA WT or Cp OGA DM . n = 6-8. ( G ) A compilation of learning performance index of flies expressing Cp OGA WT or Cp OGA DM only in the mushroom body at adult stage. n = 6. Each datapoint represents an independent experiment with approximately 200 flies. p- values were determined by unpaired t -test, and the stars indicate significant differences (***p<0.001, **p<0.01 and ns, not significant, p≥0.05). Error bars represent SD. Figure 1—source data 1. Excel spreadsheet containing source data used to generate .
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    Cell Signaling Technology Inc gfp 4b10 mouse mab
    ( A ) Scheme for expression of Cp OGA WT or Cp OGA DM in various Drosophila brain structures using different Gal4 drivers. ( B ) Immunostaining of adult Drosophila brains. Brains were stained with anti- O- GlcNAc antibody RL2 (red) to assess O- GlcNAcylation level, <t>and</t> <t>anti-GFP</t> (green) antibody to validate tissue-specific expression of Cp OGA. Nuclei were stained with DAPI (blue). Scale bar: 100 μm. ( C ) Quantification of fluorescent intensity of O- GlcNAc staining in Cp OGA WT or Cp OGA DM expressed brains. n = 4. ( D ) Immunostaining of adult Drosophila brains. Outlined areas indicate the cell bodies of Kenyon cells in mushroom body. Scale bar: 100 μm. ( E ) Quantification of relative fluorescent intensity of O- GlcNAc staining in Cp OGA WT or Cp OGA DM expressed brain structures. n = 8. ( F ) A compilation of performance index in learning test of the indicated flies expressing either Cp OGA WT or Cp OGA DM . n = 6-8. ( G ) A compilation of learning performance index of flies expressing Cp OGA WT or Cp OGA DM only in the mushroom body at adult stage. n = 6. Each datapoint represents an independent experiment with approximately 200 flies. p- values were determined by unpaired t -test, and the stars indicate significant differences (***p<0.001, **p<0.01 and ns, not significant, p≥0.05). Error bars represent SD. Figure 1—source data 1. Excel spreadsheet containing source data used to generate .
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    Cell Signaling Technology Inc gfp mouse monoclonal primary antibody
    ( A ) Construct design to generate HEK293 FRT stable cells harboring the KCNJ13 W53X allele. ( B ) Chromatogram generated from HEK293 FRT stable cells showing the W53X codon marked in the red box and the downward black arrow indicating the specific nucleotide change (G>A).( C ) Schematic of the hKCNJ13 locus highlighting the mutation c.158G>A (blue box marked with asterisk) and position of the W53X targeting sgRNA (black line) with TGG PAM (red line). ( D ) Base-editing efficiencies are shown as the percentages of sequencing reads with the corrected WT allele (and no other silent changes, bystander edits, or indels) in HEK293 W53X cells following electroporation of ABE8e protein+sgRNA (RNP) or ABE8e mRNA+sgRNA ( n = 3). Markers (diamonds) represent the individual biological replicates ( n = 3), and error bars represent SEM by 2-tailed Student’s t test. ( E ) Percentages of sequencing reads with indels in ABE8e RNP– and ABE8e mRNA–treated stable cells ( n = 3). Markers (diamonds) represent the individual biological replicates ( n = 3), and error bars represent SEM by 2-tailed Student’s t test. ( F <t>)</t> <t>Kir7.1</t> expression in ABE8e mRNA–treated cells assessed by immunocytochemistry. <t>GFP</t> primary antibody was used to enhance the endogenous signal. DAPI was used to stain the nucleus. Scale bars: 50 μm. White arrows mark membrane localization in cells.
    Gfp Mouse Monoclonal Primary 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
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    Cell Signaling Technology Inc mouse monoclonal anti gfp
    (A) Schematic of Mup1 sorting. (B) <t>Mup1-GFP</t> localization after methionine stimulation. (C) Western blotting analysis of Mup1 sorting. Cell lysates were analyzed by immunoblotting <t>using</t> <t>anti-GFP</t> and anti-G6PDH antibodies. Vacuole delivery of Mup1-GFP yields protease-resistant GFP fragments (GFP’). (D) Quantification of Mup1-GFP processing. (E) GFP-CPS localization. (F) Quantification of GFP-CPS localization of each category. (G) Schematic of retromer-mediated Vps10 recycling. (H) Vps10-GFP localization in WT, vps35 Δ (retromer), and vps13 Δ cells. (I) Quantification of Vps10-GFP localization. Scale bar: 1 µm.
    Mouse Monoclonal Anti Gfp, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Image Search Results


    ( A ) Western blot images of CKPC-1 cells stably expressing CYRI-B-p17-GFP or GFP. KPC-1 and KPC-2 cell lines were used as control. Untransfected CKPC cells were also used as a control. Membranes were probed for anti-GFP (bottom blot) and anti-CYRI-B (top blot). GAPDH was used as loading control. Molecular weights are displayed on the side. ( B ) Representative immunofluorescence images of CKPC Cyrib knockout (KO) and rescued cells. Cells were seeded on fibronectin-coated coverslips, fixed and stained for F-actin (magenta), ArpC2 (cyan), and DAPI for nuclei (yellow). Scale bars, 20 μm. Yellow dotted boxes show the sites for the magnified images. Red arrows show the positive area for ArpC2 staining at the leading edge. Scale bars, 5 μm. Graph shows manual quantification of the number of cells presenting with lamellipodia (purple) or other protrusions (green) from (B). Mean ± S.D; paired t-test was performed in n=4. p-Value as indicated. ( C ) Quantification of cell area per cell from (B) based on the F-actin staining. Scatter plot here is presented as super plot and every independent biological repeat is coloured differently. Mean ± SEM; paired t-test was performed in n=4. p-Value as indicated. ( D ) Manual quantification of the length of the cell periphery showing strong ArpC2 accumulation, normalised to the total cell periphery. Scatter plot here is presented as a super plot and every independent biological repeat is coloured differently. Mean ± SEM; unpaired t-test was performed in n=3 (from a total of 30 cells). p-Value as indicated. ( E ) Manual quantification of the relative intensity of ArpC2 on the plasma membrane to cytoplasmic average intensity. Scatter plot here is presented as a super plot and every independent biological repeat is coloured differently. Mean ± SEM; unpaired t-test was performed in n=3. p-Value as indicated. Figure 4—figure supplement 2—source data 1. Scans of original western blots unlabelled and labelled to support . Figure 4—figure supplement 2—source data 2. Number of cells presenting with lamellipodia or other protrusions in CYRI-B knockout (GFP) vs CYRI-B-GFP rescued (CYRI-B-GFP) cells. Excel data and Prism analysis to support . Figure 4—figure supplement 2—source data 3. Area per cell in CYRI-B knockout (GFP) vs CYRI-B-GFP rescued (CYRI-B-GFP) cells ( , ). Figure 4—figure supplement 2—source data 4. Percentage of the periphery staining positive for ArpC2 in CYRI-B knockout (GFP) vs CYRI-B-GFP rescued (CYRI-B-GFP) cells. Excel data and Prism analysis to support . Figure 4—figure supplement 2—source data 5. Plasma membrane to cytoplasm relative intensity of ArpC2 in CYRI-B knockout (GFP) vs CYRI-B-GFP rescued (CYRI-B-GFP) cells. Excel data and Prism analysis to support .

    Journal: eLife

    Article Title: CYRI-B-mediated macropinocytosis drives metastasis via lysophosphatidic acid receptor uptake

    doi: 10.7554/eLife.83712

    Figure Lengend Snippet: ( A ) Western blot images of CKPC-1 cells stably expressing CYRI-B-p17-GFP or GFP. KPC-1 and KPC-2 cell lines were used as control. Untransfected CKPC cells were also used as a control. Membranes were probed for anti-GFP (bottom blot) and anti-CYRI-B (top blot). GAPDH was used as loading control. Molecular weights are displayed on the side. ( B ) Representative immunofluorescence images of CKPC Cyrib knockout (KO) and rescued cells. Cells were seeded on fibronectin-coated coverslips, fixed and stained for F-actin (magenta), ArpC2 (cyan), and DAPI for nuclei (yellow). Scale bars, 20 μm. Yellow dotted boxes show the sites for the magnified images. Red arrows show the positive area for ArpC2 staining at the leading edge. Scale bars, 5 μm. Graph shows manual quantification of the number of cells presenting with lamellipodia (purple) or other protrusions (green) from (B). Mean ± S.D; paired t-test was performed in n=4. p-Value as indicated. ( C ) Quantification of cell area per cell from (B) based on the F-actin staining. Scatter plot here is presented as super plot and every independent biological repeat is coloured differently. Mean ± SEM; paired t-test was performed in n=4. p-Value as indicated. ( D ) Manual quantification of the length of the cell periphery showing strong ArpC2 accumulation, normalised to the total cell periphery. Scatter plot here is presented as a super plot and every independent biological repeat is coloured differently. Mean ± SEM; unpaired t-test was performed in n=3 (from a total of 30 cells). p-Value as indicated. ( E ) Manual quantification of the relative intensity of ArpC2 on the plasma membrane to cytoplasmic average intensity. Scatter plot here is presented as a super plot and every independent biological repeat is coloured differently. Mean ± SEM; unpaired t-test was performed in n=3. p-Value as indicated. Figure 4—figure supplement 2—source data 1. Scans of original western blots unlabelled and labelled to support . Figure 4—figure supplement 2—source data 2. Number of cells presenting with lamellipodia or other protrusions in CYRI-B knockout (GFP) vs CYRI-B-GFP rescued (CYRI-B-GFP) cells. Excel data and Prism analysis to support . Figure 4—figure supplement 2—source data 3. Area per cell in CYRI-B knockout (GFP) vs CYRI-B-GFP rescued (CYRI-B-GFP) cells ( , ). Figure 4—figure supplement 2—source data 4. Percentage of the periphery staining positive for ArpC2 in CYRI-B knockout (GFP) vs CYRI-B-GFP rescued (CYRI-B-GFP) cells. Excel data and Prism analysis to support . Figure 4—figure supplement 2—source data 5. Plasma membrane to cytoplasm relative intensity of ArpC2 in CYRI-B knockout (GFP) vs CYRI-B-GFP rescued (CYRI-B-GFP) cells. Excel data and Prism analysis to support .

    Article Snippet: Antibody , Anti-GFP (4B10) (Mouse monoclonal) , Cell Signaling , CAT. #2955 , WB: 1:1000.

    Techniques: Western Blot, Stable Transfection, Expressing, Control, Immunofluorescence, Knock-Out, Staining, Membrane

    Journal: eLife

    Article Title: CYRI-B-mediated macropinocytosis drives metastasis via lysophosphatidic acid receptor uptake

    doi: 10.7554/eLife.83712

    Figure Lengend Snippet:

    Article Snippet: Antibody , Anti-GFP (4B10) (Mouse monoclonal) , Cell Signaling , CAT. #2955 , WB: 1:1000.

    Techniques: Transfection, Construct, Isolation, Staining, Sequencing, Plasmid Preparation, RNAscope, CRISPR, SYBR Green Assay, Blocking Assay, Software

    ( A ) Scheme for expression of Cp OGA WT or Cp OGA DM in various Drosophila brain structures using different Gal4 drivers. ( B ) Immunostaining of adult Drosophila brains. Brains were stained with anti- O- GlcNAc antibody RL2 (red) to assess O- GlcNAcylation level, and anti-GFP (green) antibody to validate tissue-specific expression of Cp OGA. Nuclei were stained with DAPI (blue). Scale bar: 100 μm. ( C ) Quantification of fluorescent intensity of O- GlcNAc staining in Cp OGA WT or Cp OGA DM expressed brains. n = 4. ( D ) Immunostaining of adult Drosophila brains. Outlined areas indicate the cell bodies of Kenyon cells in mushroom body. Scale bar: 100 μm. ( E ) Quantification of relative fluorescent intensity of O- GlcNAc staining in Cp OGA WT or Cp OGA DM expressed brain structures. n = 8. ( F ) A compilation of performance index in learning test of the indicated flies expressing either Cp OGA WT or Cp OGA DM . n = 6-8. ( G ) A compilation of learning performance index of flies expressing Cp OGA WT or Cp OGA DM only in the mushroom body at adult stage. n = 6. Each datapoint represents an independent experiment with approximately 200 flies. p- values were determined by unpaired t -test, and the stars indicate significant differences (***p<0.001, **p<0.01 and ns, not significant, p≥0.05). Error bars represent SD. Figure 1—source data 1. Excel spreadsheet containing source data used to generate .

    Journal: eLife

    Article Title: Tissue-specific O- GlcNAcylation profiling identifies substrates in translational machinery in Drosophila mushroom body contributing to olfactory learning

    doi: 10.7554/eLife.91269

    Figure Lengend Snippet: ( A ) Scheme for expression of Cp OGA WT or Cp OGA DM in various Drosophila brain structures using different Gal4 drivers. ( B ) Immunostaining of adult Drosophila brains. Brains were stained with anti- O- GlcNAc antibody RL2 (red) to assess O- GlcNAcylation level, and anti-GFP (green) antibody to validate tissue-specific expression of Cp OGA. Nuclei were stained with DAPI (blue). Scale bar: 100 μm. ( C ) Quantification of fluorescent intensity of O- GlcNAc staining in Cp OGA WT or Cp OGA DM expressed brains. n = 4. ( D ) Immunostaining of adult Drosophila brains. Outlined areas indicate the cell bodies of Kenyon cells in mushroom body. Scale bar: 100 μm. ( E ) Quantification of relative fluorescent intensity of O- GlcNAc staining in Cp OGA WT or Cp OGA DM expressed brain structures. n = 8. ( F ) A compilation of performance index in learning test of the indicated flies expressing either Cp OGA WT or Cp OGA DM . n = 6-8. ( G ) A compilation of learning performance index of flies expressing Cp OGA WT or Cp OGA DM only in the mushroom body at adult stage. n = 6. Each datapoint represents an independent experiment with approximately 200 flies. p- values were determined by unpaired t -test, and the stars indicate significant differences (***p<0.001, **p<0.01 and ns, not significant, p≥0.05). Error bars represent SD. Figure 1—source data 1. Excel spreadsheet containing source data used to generate .

    Article Snippet: Antibody , Anti-GFP Mouse Monoclonal Antibody (4B10) , Cell Signaling Technology , Cat# 2955, RRID: AB_1196614 , WB (1:1000) IF (1:200).

    Techniques: Expressing, Immunostaining, Staining

    ( A ) Heatmaps showing the mRNA levels (upper) and the normalized O- GlcNAc levels (lower) of the identified ribosomal candidates in different brain regions. ( B ) Immunoprecipitation of ribosomes using FLAG-tagged RpL13A. The expression of RpL13A-FLAG was validated by immunoblotting with anti-FLAG antibody. Ribosomal proteins were visualized using silver staining, and O- GlcNAcylation of ribosomes was analyzed by immunoblotting with anti- O- GlcNAc antibody RL2. ( C ) A compilation of the performance index of the indicated flies in the learning test. Learning defect of flies expressing Cp OGA WT was corrected by selective expression of dMyc in mushroom body. n = 6-7. Each datapoint represents an independent experiment with approximately 200 flies. ( D ) Ex vivo measurement of protein synthesis in mushroom body using the O-propargyl-puromycin (OPP) assay. Brains from the indicated flies were stained with anti-GFP (green) antibody to validate Cp OGA expression, and OPP (gray) to quantify protein synthesis. Nuclei were visualized with DAPI (blue). Outlined areas indicate the cell bodies of Kenyon cells of mushroom body. Scale bar: 100 μm. ( E ) Quantification of relative OPP fluorescent intensity in mushroom body regions. n = 8-12. p -values were determined by unpaired t -test, the stars indicate significant differences (***p<0.001, **p<0.01, *p<0.05). Error bars represent SD. Figure 4—source data 1. Raw data of all western blots for . Figure 4—source data 2. Complete and uncropped membranes of all western blots for . Figure 4—source data 3. Excel spreadsheet containing source data used to generate .

    Journal: eLife

    Article Title: Tissue-specific O- GlcNAcylation profiling identifies substrates in translational machinery in Drosophila mushroom body contributing to olfactory learning

    doi: 10.7554/eLife.91269

    Figure Lengend Snippet: ( A ) Heatmaps showing the mRNA levels (upper) and the normalized O- GlcNAc levels (lower) of the identified ribosomal candidates in different brain regions. ( B ) Immunoprecipitation of ribosomes using FLAG-tagged RpL13A. The expression of RpL13A-FLAG was validated by immunoblotting with anti-FLAG antibody. Ribosomal proteins were visualized using silver staining, and O- GlcNAcylation of ribosomes was analyzed by immunoblotting with anti- O- GlcNAc antibody RL2. ( C ) A compilation of the performance index of the indicated flies in the learning test. Learning defect of flies expressing Cp OGA WT was corrected by selective expression of dMyc in mushroom body. n = 6-7. Each datapoint represents an independent experiment with approximately 200 flies. ( D ) Ex vivo measurement of protein synthesis in mushroom body using the O-propargyl-puromycin (OPP) assay. Brains from the indicated flies were stained with anti-GFP (green) antibody to validate Cp OGA expression, and OPP (gray) to quantify protein synthesis. Nuclei were visualized with DAPI (blue). Outlined areas indicate the cell bodies of Kenyon cells of mushroom body. Scale bar: 100 μm. ( E ) Quantification of relative OPP fluorescent intensity in mushroom body regions. n = 8-12. p -values were determined by unpaired t -test, the stars indicate significant differences (***p<0.001, **p<0.01, *p<0.05). Error bars represent SD. Figure 4—source data 1. Raw data of all western blots for . Figure 4—source data 2. Complete and uncropped membranes of all western blots for . Figure 4—source data 3. Excel spreadsheet containing source data used to generate .

    Article Snippet: Antibody , Anti-GFP Mouse Monoclonal Antibody (4B10) , Cell Signaling Technology , Cat# 2955, RRID: AB_1196614 , WB (1:1000) IF (1:200).

    Techniques: Immunoprecipitation, Expressing, Western Blot, Silver Staining, Ex Vivo, Staining

    Journal: eLife

    Article Title: Tissue-specific O- GlcNAcylation profiling identifies substrates in translational machinery in Drosophila mushroom body contributing to olfactory learning

    doi: 10.7554/eLife.91269

    Figure Lengend Snippet:

    Article Snippet: Antibody , Anti-GFP Mouse Monoclonal Antibody (4B10) , Cell Signaling Technology , Cat# 2955, RRID: AB_1196614 , WB (1:1000) IF (1:200).

    Techniques: Silver Staining, Protease Inhibitor, Magnetic Beads, SYBR Green Assay, Sequencing, Modification, Software, Microscopy

    ( A ) Construct design to generate HEK293 FRT stable cells harboring the KCNJ13 W53X allele. ( B ) Chromatogram generated from HEK293 FRT stable cells showing the W53X codon marked in the red box and the downward black arrow indicating the specific nucleotide change (G>A).( C ) Schematic of the hKCNJ13 locus highlighting the mutation c.158G>A (blue box marked with asterisk) and position of the W53X targeting sgRNA (black line) with TGG PAM (red line). ( D ) Base-editing efficiencies are shown as the percentages of sequencing reads with the corrected WT allele (and no other silent changes, bystander edits, or indels) in HEK293 W53X cells following electroporation of ABE8e protein+sgRNA (RNP) or ABE8e mRNA+sgRNA ( n = 3). Markers (diamonds) represent the individual biological replicates ( n = 3), and error bars represent SEM by 2-tailed Student’s t test. ( E ) Percentages of sequencing reads with indels in ABE8e RNP– and ABE8e mRNA–treated stable cells ( n = 3). Markers (diamonds) represent the individual biological replicates ( n = 3), and error bars represent SEM by 2-tailed Student’s t test. ( F ) Kir7.1 expression in ABE8e mRNA–treated cells assessed by immunocytochemistry. GFP primary antibody was used to enhance the endogenous signal. DAPI was used to stain the nucleus. Scale bars: 50 μm. White arrows mark membrane localization in cells.

    Journal: The Journal of Clinical Investigation

    Article Title: Nonviral base editing of KCNJ13 mutation preserves vision in a model of inherited retinal channelopathy

    doi: 10.1172/JCI171356

    Figure Lengend Snippet: ( A ) Construct design to generate HEK293 FRT stable cells harboring the KCNJ13 W53X allele. ( B ) Chromatogram generated from HEK293 FRT stable cells showing the W53X codon marked in the red box and the downward black arrow indicating the specific nucleotide change (G>A).( C ) Schematic of the hKCNJ13 locus highlighting the mutation c.158G>A (blue box marked with asterisk) and position of the W53X targeting sgRNA (black line) with TGG PAM (red line). ( D ) Base-editing efficiencies are shown as the percentages of sequencing reads with the corrected WT allele (and no other silent changes, bystander edits, or indels) in HEK293 W53X cells following electroporation of ABE8e protein+sgRNA (RNP) or ABE8e mRNA+sgRNA ( n = 3). Markers (diamonds) represent the individual biological replicates ( n = 3), and error bars represent SEM by 2-tailed Student’s t test. ( E ) Percentages of sequencing reads with indels in ABE8e RNP– and ABE8e mRNA–treated stable cells ( n = 3). Markers (diamonds) represent the individual biological replicates ( n = 3), and error bars represent SEM by 2-tailed Student’s t test. ( F ) Kir7.1 expression in ABE8e mRNA–treated cells assessed by immunocytochemistry. GFP primary antibody was used to enhance the endogenous signal. DAPI was used to stain the nucleus. Scale bars: 50 μm. White arrows mark membrane localization in cells.

    Article Snippet: As the protein is GFP tagged, GFP mouse monoclonal primary antibody (Cell Signaling, 2955, 1:250) was used to enhance the Kir7.1 protein expression for its detection in the cells.

    Techniques: Construct, Generated, Mutagenesis, Sequencing, Electroporation, Expressing, Immunocytochemistry, Staining, Membrane

    (A) Schematic of Mup1 sorting. (B) Mup1-GFP localization after methionine stimulation. (C) Western blotting analysis of Mup1 sorting. Cell lysates were analyzed by immunoblotting using anti-GFP and anti-G6PDH antibodies. Vacuole delivery of Mup1-GFP yields protease-resistant GFP fragments (GFP’). (D) Quantification of Mup1-GFP processing. (E) GFP-CPS localization. (F) Quantification of GFP-CPS localization of each category. (G) Schematic of retromer-mediated Vps10 recycling. (H) Vps10-GFP localization in WT, vps35 Δ (retromer), and vps13 Δ cells. (I) Quantification of Vps10-GFP localization. Scale bar: 1 µm.

    Journal: bioRxiv

    Article Title: A role for Vps13-mediated lipid transfer at the ER-endosome contact site in ESCRT-mediated sorting

    doi: 10.1101/2023.07.06.547997

    Figure Lengend Snippet: (A) Schematic of Mup1 sorting. (B) Mup1-GFP localization after methionine stimulation. (C) Western blotting analysis of Mup1 sorting. Cell lysates were analyzed by immunoblotting using anti-GFP and anti-G6PDH antibodies. Vacuole delivery of Mup1-GFP yields protease-resistant GFP fragments (GFP’). (D) Quantification of Mup1-GFP processing. (E) GFP-CPS localization. (F) Quantification of GFP-CPS localization of each category. (G) Schematic of retromer-mediated Vps10 recycling. (H) Vps10-GFP localization in WT, vps35 Δ (retromer), and vps13 Δ cells. (I) Quantification of Vps10-GFP localization. Scale bar: 1 µm.

    Article Snippet: For immunoblotting, mouse monoclonal anti-GFP (B-2; Santa Cruz, Sc-9996), rabbit polyclonal anti-G6PDH (Sigma-Aldrich, SAB2100871), and anti-ubiquitin (P4D1; Cell Signaling, #3936) were used at dilution factors of 1:5000, 1:10,000 and 1:1000, respectively.

    Techniques: Western Blot

    (A) Localization of Vps13-GFP. Scale bar: 1 µm. (B) Quantification of Vps13-GFP colocalizing with mCherry-Vps21. (C) Localization of Vps13 ΔC -GFP (residues 1-1851). Scale bar: 1 µm. (D) Quantification of Vps13-GFP puncta localization. (E) The localization of Vps13 N -GFP (residues 1-39). Scale bar: 1 µm. (F) Line scan analysis for the region highlighted by the yellow dash line in E. (G) Vps13-GFP localization at the ER-endosome contact site. Vps13-GFP, DsRed-HDEL (ER), and Mup1-BFP (endosome) expressing cells were stimulated with methionine. Scale bar: 1 µm. (H) Live cell-imaging analysis of the ER (GFP-HDEL) and endosome (mCherry-Vps21). Scale bar: 500 nm. (I) ER diameters per 100 nm interval starting at the endosome contact along 500 nm length of ER. (J and K) Mup1-GFP processing in vps13 mutants after methionine stimulation. Cell lysates were analyzed by immunoblotting using anti-GFP and anti-G6PDH antibodies.

    Journal: bioRxiv

    Article Title: A role for Vps13-mediated lipid transfer at the ER-endosome contact site in ESCRT-mediated sorting

    doi: 10.1101/2023.07.06.547997

    Figure Lengend Snippet: (A) Localization of Vps13-GFP. Scale bar: 1 µm. (B) Quantification of Vps13-GFP colocalizing with mCherry-Vps21. (C) Localization of Vps13 ΔC -GFP (residues 1-1851). Scale bar: 1 µm. (D) Quantification of Vps13-GFP puncta localization. (E) The localization of Vps13 N -GFP (residues 1-39). Scale bar: 1 µm. (F) Line scan analysis for the region highlighted by the yellow dash line in E. (G) Vps13-GFP localization at the ER-endosome contact site. Vps13-GFP, DsRed-HDEL (ER), and Mup1-BFP (endosome) expressing cells were stimulated with methionine. Scale bar: 1 µm. (H) Live cell-imaging analysis of the ER (GFP-HDEL) and endosome (mCherry-Vps21). Scale bar: 500 nm. (I) ER diameters per 100 nm interval starting at the endosome contact along 500 nm length of ER. (J and K) Mup1-GFP processing in vps13 mutants after methionine stimulation. Cell lysates were analyzed by immunoblotting using anti-GFP and anti-G6PDH antibodies.

    Article Snippet: For immunoblotting, mouse monoclonal anti-GFP (B-2; Santa Cruz, Sc-9996), rabbit polyclonal anti-G6PDH (Sigma-Aldrich, SAB2100871), and anti-ubiquitin (P4D1; Cell Signaling, #3936) were used at dilution factors of 1:5000, 1:10,000 and 1:1000, respectively.

    Techniques: Expressing, Live Cell Imaging, Western Blot

    (A) Schematic of ESCRT-mediated sorting at the endosome. (B) Can1-FKBP processing after rapamycin treatments. Cell lysates were analyzed by immunoblotting using anti-GFP and anti-G6PDH antibodies. (C) GFP-Vps27 (ESCRT-0) localization. (D) Snf7-GFP (ESCRT-III) localization. Scale bar: 1 µm.

    Journal: bioRxiv

    Article Title: A role for Vps13-mediated lipid transfer at the ER-endosome contact site in ESCRT-mediated sorting

    doi: 10.1101/2023.07.06.547997

    Figure Lengend Snippet: (A) Schematic of ESCRT-mediated sorting at the endosome. (B) Can1-FKBP processing after rapamycin treatments. Cell lysates were analyzed by immunoblotting using anti-GFP and anti-G6PDH antibodies. (C) GFP-Vps27 (ESCRT-0) localization. (D) Snf7-GFP (ESCRT-III) localization. Scale bar: 1 µm.

    Article Snippet: For immunoblotting, mouse monoclonal anti-GFP (B-2; Santa Cruz, Sc-9996), rabbit polyclonal anti-G6PDH (Sigma-Aldrich, SAB2100871), and anti-ubiquitin (P4D1; Cell Signaling, #3936) were used at dilution factors of 1:5000, 1:10,000 and 1:1000, respectively.

    Techniques: Western Blot

    (A) The ubiquitination of Mup1-GFP. Mup1-GFP expressing cells were stimulated with methionine for 15 min and then immunoprecipitated under denatured conditions. The immunoprecipitated (IP) products were analyzed by immunoblotting using anti-GFP and anti-ubiquitin antibodies. (B) Schematic of a rapamycin-dependent degradation system for Can1. (C) Can1-FKBP localization. Rapamycin-insensitive mutant cells ( tor1-1 , fpr1 Δ) expressing Can1-GFP-2xFKBP (Can1-FKBP) and FRB-3xUb were treated with rapamycin. (D) Quantification of Can-FKBP sorting. (E) Vps4-GFP localization. (F) Quantification of endosomal Vps4-GFP localization. (G) Schematic of Mup1-pHluorin-mCherry sorting. (H) Mup1-pHluorin-mCherry localization. Mup1-pHluorin-mCherry expressing vam7 ts and vam7 t vps13 Δ cells were incubated at 37°C and stimulated with methionine. (I) pHluorin fluorescence of Mup1-pHluorin-mCherry. Scale bar: 1 µm.

    Journal: bioRxiv

    Article Title: A role for Vps13-mediated lipid transfer at the ER-endosome contact site in ESCRT-mediated sorting

    doi: 10.1101/2023.07.06.547997

    Figure Lengend Snippet: (A) The ubiquitination of Mup1-GFP. Mup1-GFP expressing cells were stimulated with methionine for 15 min and then immunoprecipitated under denatured conditions. The immunoprecipitated (IP) products were analyzed by immunoblotting using anti-GFP and anti-ubiquitin antibodies. (B) Schematic of a rapamycin-dependent degradation system for Can1. (C) Can1-FKBP localization. Rapamycin-insensitive mutant cells ( tor1-1 , fpr1 Δ) expressing Can1-GFP-2xFKBP (Can1-FKBP) and FRB-3xUb were treated with rapamycin. (D) Quantification of Can-FKBP sorting. (E) Vps4-GFP localization. (F) Quantification of endosomal Vps4-GFP localization. (G) Schematic of Mup1-pHluorin-mCherry sorting. (H) Mup1-pHluorin-mCherry localization. Mup1-pHluorin-mCherry expressing vam7 ts and vam7 t vps13 Δ cells were incubated at 37°C and stimulated with methionine. (I) pHluorin fluorescence of Mup1-pHluorin-mCherry. Scale bar: 1 µm.

    Article Snippet: For immunoblotting, mouse monoclonal anti-GFP (B-2; Santa Cruz, Sc-9996), rabbit polyclonal anti-G6PDH (Sigma-Aldrich, SAB2100871), and anti-ubiquitin (P4D1; Cell Signaling, #3936) were used at dilution factors of 1:5000, 1:10,000 and 1:1000, respectively.

    Techniques: Expressing, Immunoprecipitation, Western Blot, Mutagenesis, Incubation, Fluorescence