Supplementary MaterialsFigure 1source data 1: Percentage of double-edged polygons in ZO-1-positive honeycomb. polygons in ZO-1-positive honeycomb statistically analyzed in Figure 1E.DOI: http://dx.doi.org/10.7554/eLife.19593.006 elife-19593-fig1-data3.xlsx (59K) DOI:?10.7554/eLife.19593.006 Figure 2source data 1: Fluorescent intensity of Dsg1-EC at single- and double-edged polygons. Fluorescent intensity of Dsg1-EC at the center of ZO-1-positive polygons in ETA-induced cellular sheet statistically analyzed in Figure 2D.DOI: http://dx.doi.org/10.7554/eLife.19593.015 elife-19593-fig2-data1.xlsx (83K) DOI:?10.7554/eLife.19593.015 Abstract In multicellular organisms, cells adopt various shapes, from Radiprodil flattened sheets of endothelium to dendritic neurons, that allow the cells to function effectively. Here, we elucidated the unique shape of cells in the cornified stratified epithelia of the mammalian epidermis that allows them to achieve homeostasis of the tight junction (TJ) barrier. Using intimate in vivo 3D imaging, we found that the basic shape of TJ-bearing cells is a flattened Kelvin’s tetrakaidecahedron (f-TKD), an optimal shape for filling space. In vivo live imaging further elucidated the dynamic replacement of TJs on the edges of f-TKD cells that enables the TJ-bearing cells to Radiprodil translocate across the TJ barrier. We propose a spatiotemporal orchestration model of f-TKD cell turnover, where in the classic context of ‘form follows function’, cell shape provides a fundamental basis for the barrier homeostasis and physical strength of cornified stratified epithelia. DOI: http://dx.doi.org/10.7554/eLife.19593.001 image of ZO-1-positive honeycomb in mouse-ear epidermis showing double-edged polygons (*) and single-edged polygons (#). (C) Regularity in the size of the ZO-1-positive polygons represented in (B) and Figure 1figure supplement 1, shown by the mean SEM [error bars] (one-way ANOVA multiple comparison test). (D) 3D image of a ZO-1-positive double-edged polygon in view (top) and 90-rotated side view of the yellow-dotted rectangle (bottom). Upper exterior polygon, yellow arrowheads; lower interior polygon, white arrows. See Video 1. (E) Regularity of relative Z-axis position. Boxplots show the median, minimum, maximum, and interquartile range (one-way ANOVA multiple comparison test) for the ZO-1-positive polygons represented in Figure 1figure supplement 2. (F) In vivo live images of Venus in the ear of ZO-1-Venus mice (left column) and their schematics (right column). Yellow arrowheads and green edges, edges of a Venus-positive polygon; white arrows and purple edges, edges of a newly appearing Venus-positive polygon; black arrows, Venus-positive edges connecting each vertex of the two polygons. See Video 4. Scale bars, 10 m.?TJ, tight junction; SC, stratum corneum. DOI: http://dx.doi.org/10.7554/eLife.19593.003 Figure 1source data 1.Percentage of double-edged polygons in ZO-1-positive honeycomb. The number of single- and double-edged polygons in 20 square images of 15376 m2 from five independent assays. A represented image is shown in Figure 1B. DOI: http://dx.doi.org/10.7554/eLife.19593.004 Click here to view.(36K, xlsx) Figure 1source data 2.Size of the ZO-1-positive polygons. The size of single- and double-edged polygons in ZO-1-positive honeycomb statistically analyzed in Number 1C. How to define areas of polygons are demonstrated in Number 1figure product 1. DOI: http://dx.doi.org/10.7554/eLife.19593.005 Click here to view.(40K, xlsx) Number 1source data 3.Z-axis position of the ZO-1-positive polygons. The Z-axis position of solitary- and double-edged polygons in ZO-1-positive honeycomb statistically analyzed in Number 1E. DOI: http://dx.doi.org/10.7554/eLife.19593.006 Click here to view.(59K, xlsx) Number 1figure product 1. Open in a separate window Areas of outside, interior and single-edged polygons.(ACC) Drawings of the ZO-1-positive edges shown in Number 1B. The areas of polygons evaluated in Number 1C are depicted for the exterior [pink in (A)] and interior [yellow in (B)] polygons of double-edged polygons (*), and for the single-edged polygon [green in (C)] adjacent to the double-edged polygons. The area of the single-edged polygon includes the overlapping area with the adjacent outside polygons (reddish arrows). DOI: http://dx.doi.org/10.7554/eLife.19593.007 Figure Radiprodil 1figure supplement 2. Open in a separate window Relative Z-axis position of TJ polygons in TJ honeycomb GRK4 evaluated in vivo.A representative image of a ZO-1-positive double-edged polygon surrounded by six single-edged polygons evaluated in Number 1E. The Z-axis position of each polygon was defined by an average of the Z-axis positions (figures) of its vertices, analyzed by Imaris software (purple arrowheads, external polygon; yellow arrowheads, internal polygon; green arrowheads, adjacent single-edged polygons). Level pub, 10 m.?TJ,?limited?junction. DOI: http://dx.doi.org/10.7554/eLife.19593.008 Figure 1figure supplement 3. Open in a separate windowpane Epidermal TJ in ZO-1-Venus transgenic mice.(A) Colocalization of occludin, a transmembrane protein located in the TJs, with Venus in whole-mounted epidermal sheet from your ear skin of a ZO-1-Venus transgenic mouse. No morphological changes were observed in the TJ honeycomb.