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a Schematic of the viral CRISPR/Cas9 approach to <t>disrupt</t> <t>DLK</t> and/or <t>LZK</t> in retinal cells. b Retinae stained with phosphorylated c-Jun and mCherry at 10 weeks post tamoxifen following treatment with sgLacZ/sgGFP, or sgDLK/sgLZK on the opposite eye. Inlays are of boxed areas and scale bar is 5 µm in boxed areas. c Representative images of phosphorylated c-Jun immunostaining in retinal flatmounts following administration of sgLacZ/sgGFP, sgDLK/sgDLK, sgLZK/sgLZK, or sgDLK/sgLZK. d Overview of retinas with locations of cleaved caspase-3+ cells in the GCL indicated by black X in viral-treated eyes. e Density of phosphorylated c-Jun cells within the GCL following sgDLK/sgDLK administration. ** p = 0.0080. Myrf fl/fl n = 3, Myrf ΔiSox10 n = 4 mice. f Density of phosphorylated c-Jun positive cells in the GCL following sgLZK/sgLZK administration. ns p = 0.0766. Myrf fl/fl n = 4, Myrf ΔiSox10 n = 5 mice. g Density of phosphorylated c-Jun cells within the GCL following sgDLK/sgLZK administration. * p = 0.0148. Myrf fl/fl n = 3, Myrf ΔiSox10 n = 5 mice. h Cleaved-caspase-3+ cells within the GCL following sgDLK/sgDLK administration. * p = 0.0254. Myrf fl/fl n = 2, Myrf ΔiSox10 n = 5 mice. i The density of cleaved-caspase-3+ cells within the GCL after sgLZK/sgLZK administration. ns P = 0.8530. Myrf fl/fl n = 3, Myrf ΔiSox10 n = 5 mice. j Cleaved-caspase-3+ cells within the GCL following sgDLK/sgLZK administration. * p = 0.0125. Myrf fl/fl n = 3, Myrf ΔiSox10 n = 5 mice. Scale bars are 50 µm in ( b , c ) and 500 µm in ( d ). Connected lines indicate retinae from the same mouse in ( e – j ). Paired Student’s t test with Holm-Šidák correction for multiple comparisons was used from ( e – j ). All statistical tests are two-sided. ns not statistically significant. Error bars are SEM. Source data for this Figure are provided as a Source Data file. a created in BioRender. Duncan (2023) BioRender.com/e99a533.
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a Schematic of the viral CRISPR/Cas9 approach to disrupt DLK and/or LZK in retinal cells. b Retinae stained with phosphorylated c-Jun and mCherry at 10 weeks post tamoxifen following treatment with sgLacZ/sgGFP, or sgDLK/sgLZK on the opposite eye. Inlays are of boxed areas and scale bar is 5 µm in boxed areas. c Representative images of phosphorylated c-Jun immunostaining in retinal flatmounts following administration of sgLacZ/sgGFP, sgDLK/sgDLK, sgLZK/sgLZK, or sgDLK/sgLZK. d Overview of retinas with locations of cleaved caspase-3+ cells in the GCL indicated by black X in viral-treated eyes. e Density of phosphorylated c-Jun cells within the GCL following sgDLK/sgDLK administration. ** p = 0.0080. Myrf fl/fl n = 3, Myrf ΔiSox10 n = 4 mice. f Density of phosphorylated c-Jun positive cells in the GCL following sgLZK/sgLZK administration. ns p = 0.0766. Myrf fl/fl n = 4, Myrf ΔiSox10 n = 5 mice. g Density of phosphorylated c-Jun cells within the GCL following sgDLK/sgLZK administration. * p = 0.0148. Myrf fl/fl n = 3, Myrf ΔiSox10 n = 5 mice. h Cleaved-caspase-3+ cells within the GCL following sgDLK/sgDLK administration. * p = 0.0254. Myrf fl/fl n = 2, Myrf ΔiSox10 n = 5 mice. i The density of cleaved-caspase-3+ cells within the GCL after sgLZK/sgLZK administration. ns P = 0.8530. Myrf fl/fl n = 3, Myrf ΔiSox10 n = 5 mice. j Cleaved-caspase-3+ cells within the GCL following sgDLK/sgLZK administration. * p = 0.0125. Myrf fl/fl n = 3, Myrf ΔiSox10 n = 5 mice. Scale bars are 50 µm in ( b , c ) and 500 µm in ( d ). Connected lines indicate retinae from the same mouse in ( e – j ). Paired Student’s t test with Holm-Šidák correction for multiple comparisons was used from ( e – j ). All statistical tests are two-sided. ns not statistically significant. Error bars are SEM. Source data for this Figure are provided as a Source Data file. a created in BioRender. Duncan (2023) BioRender.com/e99a533.

Journal: Nature Communications

Article Title: Remyelination protects neurons from DLK-mediated neurodegeneration

doi: 10.1038/s41467-024-53429-5

Figure Lengend Snippet: a Schematic of the viral CRISPR/Cas9 approach to disrupt DLK and/or LZK in retinal cells. b Retinae stained with phosphorylated c-Jun and mCherry at 10 weeks post tamoxifen following treatment with sgLacZ/sgGFP, or sgDLK/sgLZK on the opposite eye. Inlays are of boxed areas and scale bar is 5 µm in boxed areas. c Representative images of phosphorylated c-Jun immunostaining in retinal flatmounts following administration of sgLacZ/sgGFP, sgDLK/sgDLK, sgLZK/sgLZK, or sgDLK/sgLZK. d Overview of retinas with locations of cleaved caspase-3+ cells in the GCL indicated by black X in viral-treated eyes. e Density of phosphorylated c-Jun cells within the GCL following sgDLK/sgDLK administration. ** p = 0.0080. Myrf fl/fl n = 3, Myrf ΔiSox10 n = 4 mice. f Density of phosphorylated c-Jun positive cells in the GCL following sgLZK/sgLZK administration. ns p = 0.0766. Myrf fl/fl n = 4, Myrf ΔiSox10 n = 5 mice. g Density of phosphorylated c-Jun cells within the GCL following sgDLK/sgLZK administration. * p = 0.0148. Myrf fl/fl n = 3, Myrf ΔiSox10 n = 5 mice. h Cleaved-caspase-3+ cells within the GCL following sgDLK/sgDLK administration. * p = 0.0254. Myrf fl/fl n = 2, Myrf ΔiSox10 n = 5 mice. i The density of cleaved-caspase-3+ cells within the GCL after sgLZK/sgLZK administration. ns P = 0.8530. Myrf fl/fl n = 3, Myrf ΔiSox10 n = 5 mice. j Cleaved-caspase-3+ cells within the GCL following sgDLK/sgLZK administration. * p = 0.0125. Myrf fl/fl n = 3, Myrf ΔiSox10 n = 5 mice. Scale bars are 50 µm in ( b , c ) and 500 µm in ( d ). Connected lines indicate retinae from the same mouse in ( e – j ). Paired Student’s t test with Holm-Šidák correction for multiple comparisons was used from ( e – j ). All statistical tests are two-sided. ns not statistically significant. Error bars are SEM. Source data for this Figure are provided as a Source Data file. a created in BioRender. Duncan (2023) BioRender.com/e99a533.

Article Snippet: Plasmids generated in this study for DLK or LZK knockout have been deposited at Addgene (Plasmid #208834, 208835, 208836, 208837) and AAV2-hSyn1-Cre is available upon request.

Techniques: CRISPR, Staining, Immunostaining