Fig. 1: Caveolae localization and modulation in human epidermis and 2D co-culture.
From: Coupling of melanocyte signaling and mechanics by caveolae is required for human skin pigmentation
a IFM images of melanocytes and keratinocytes co-cultured for 1 day and immunolabelled for Cav1 (caveolin-1) or Cavin1 (top or bottom, respectively; green) and HMB45 (premelanosome protein PMEL, red). Cav1 or Cavin1 polarization, arrowheads; melanocytes, asterisks. The boxed regions mark the zoomed area. Bars, 10 µm. b Quantification of Cav1 or Cavin1 mean fluorescent intensity in boxes depicted in a. c, d Quantification of the frequency of melanocytes with Cav1 or Cavin1 polarization [as in a, arrowheads] in mono- or co-cultures (c), or maintained in different media (d). e Conventional 2D EM from human skin tissue fixed chemically (top) or immobilized by high-pressure freezing (HPF, bottom); plasma membranes of keratinocytes (red) and of melanocytes (blue), plasma membrane invaginations with morphological features of caveolae (arrowheads). The boxed regions mark the zoomed area. Bars: (main) 1 µm; (zoom) 100 nm. f 3D-model reconstruction electron tomography of the melanocyte–keratinocyte interface in human skin epidermis; plasma membrane of melanocytes (green) and of keratinocytes (blue), limiting membrane of melanosomes (red), melanin (black), caveolae (white) in single (arrowhead) and clustered structures (arrow) and keratin bundles (white arrowheads). See also Supplementary Video 1 and Supplementary Fig. 1f. g, h Quantification of the number of caveolae [as identified in e] per 10 µm of plasma membrane at the indicated interface (g) and of individual cell types at melanocyte–keratinocyte interface (h) during 3D-HRPE formation. i Immunoblot analysis and quantification of Cav1 protein levels (normalized to Calnexin) in melanocytes daily exposed to UV-B radiations (ultraviolet-B; 11 mJ/cm2, 3 days). Asterisk, Cav1 full-length protein and truncated form (upper and lower bands, respectively). Values are mean ± s.e.m. b 12 cells. c, d 150 cells, three independent experiments. g, h 1 experiment. i Three independent experiments. b Two-tailed unpaired t-test with Welch’s correction; c, d one-way ANOVA with Tukey’s multiple-comparison test; g, h comparison between interface/cells at the same time point: two-tailed unpaired t-test with Welch’s correction; comparison between time points from the same cell type: one-way ANOVA with Tukey’s multiple-comparison test; i one-way ANOVA with Sidak’s multiple-comparison test. See also Supplementary Table 1.
