Figure 2. DPM1 modulates DSP... | Rockefeller University Press

Figure 2.

$DPM1 modulates DSP localization and cytoskeletal organization. (a) Immunostaining of DSP and DSG2 in HaCaT keratinocytes. Merge indicates overlap between DSP, DSG2, and nuclei stained with DAPI. Scale bar: 10 µm distance. (b) Quantification of the number of DSP puncta over the respective length of cell membrane (µm) from individual cells (represented by individual dots), N = 3. Unpaired Student’s t test. (c) Immunostaining of DSP and DSG2 in primary human keratinocytes. Scale bar: 10 µm distance. Panel shows representatives of three biological replicates. (d) Immunostaining of DSP in 3D-RHE of control (sgNT1) and sgDPM1 conditions. White dashed line indicates insert membrane. Magenta dashed rectangles mark regions magnified on the right (zoomed 2× of original image). Representative of three biological replicates. Scale bar: 10 µm distance. (e–g) Keratin staining depicted by pan-cytokeratin in control and DPM1 KD HaCaT keratinocytes. Analysis done by quantifying the keratin staining intensity (A.U.) across cell borders spanning a distance of 10 µm. Peak width was calculated between the two highest points from the distribution plot profile graph. Scale bar: 10 µm distance. 30 individual cells were quantified from three independent biological replicates. Unpaired Student’s t test. (h–j) F-actin stained by phalloidin in control and DPM1 KD HaCaT keratinocytes. Analysis done by quantifying the F-actin staining intensity (A.U.) across cell borders spanning a distance of 10 µm. The width of the peaks was calculated between the baseline values from distribution plot profile graphs. Scale bar: 10 µm distance. 30 individual cells were analyzed from three independent biological replicates. Unpaired Student’s t test. (k) Schematic of the AFM setup to measure cellular elasticity. Graph shows cellular elasticity, indicated by Young’s modulus (kPa). Each dot represents single cells from three biological replicates. Unpaired Student’s t test. (l and m) Kymographs and mobile fraction analysis derived from FRAP assays used to measure DSP stability at cell–cell contact sites (N = 3). Scale bar on the y-axis = 1 µm. Each dot represents one biological replicate. Unpaired Student’s t test.$

DPM1 modulates DSP localization and cytoskeletal organization. (a) Immunostaining of DSP and DSG2 in HaCaT keratinocytes. Merge indicates overlap between DSP, DSG2, and nuclei stained with DAPI. Scale bar: 10 µm distance. (b) Quantification of the number of DSP puncta over the respective length of cell membrane (µm) from individual cells (represented by individual dots), N = 3. Unpaired Student’s t test. (c) Immunostaining of DSP and DSG2 in primary human keratinocytes. Scale bar: 10 µm distance. Panel shows representatives of three biological replicates. (d) Immunostaining of DSP in 3D-RHE of control (sgNT1) and sgDPM1 conditions. White dashed line indicates insert membrane. Magenta dashed rectangles mark regions magnified on the right (zoomed 2× of original image). Representative of three biological replicates. Scale bar: 10 µm distance. (e–g) Keratin staining depicted by pan-cytokeratin in control and DPM1 KD HaCaT keratinocytes. Analysis done by quantifying the keratin staining intensity (A.U.) across cell borders spanning a distance of 10 µm. Peak width was calculated between the two highest points from the distribution plot profile graph. Scale bar: 10 µm distance. 30 individual cells were quantified from three independent biological replicates. Unpaired Student’s t test. (h–j) F-actin stained by phalloidin in control and DPM1 KD HaCaT keratinocytes. Analysis done by quantifying the F-actin staining intensity (A.U.) across cell borders spanning a distance of 10 µm. The width of the peaks was calculated between the baseline values from distribution plot profile graphs. Scale bar: 10 µm distance. 30 individual cells were analyzed from three independent biological replicates. Unpaired Student’s t test. (k) Schematic of the AFM setup to measure cellular elasticity. Graph shows cellular elasticity, indicated by Young’s modulus (kPa). Each dot represents single cells from three biological replicates. Unpaired Student’s t test. (l and m) Kymographs and mobile fraction analysis derived from FRAP assays used to measure DSP stability at cell–cell contact sites (N = 3). Scale bar on the y-axis = 1 µm. Each dot represents one biological replicate. Unpaired Student’s t test.

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