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Rasip1 is dynamically involved in the establishment of apical compartments. A Schematic depicting the anastomosis of two tip cells in the formation of the DLAV. A stable cell-cell contact site is formed with the deposition of junctional proteins, including Cdh5 (I). The cell-cell contact site opens into a ring structure while the apical membrane is inserted into the luminal pocket within the junctional ring (II). Cell rearrangement leads to the elongation of the junctional ring (III). A transverse view of the luminal pocket located within the junctional ring (IV). B Time-lapse imaging of Cdh5-Venus and ZO1-tdTomato, captured from 30 hpf throughout the patch-to-ring transition. (B’) Intensities of Cdh5-Venus and ZO1-tdTomato along the dashed lines in (B). C Antibody staining for Cdh5 and Rasip1 at junctional patches and rings of varying sizes, from initial cell-cell contacts to the formation of stable junctional rings. The Dice-Sørensen coefficient was calculated as twice the joint area of the two signals divided by the sum of the area of each signal after automatic thresholding. D The Dice coefficient and the corresponding area of various junctional patches and rings, with data points color-coded by area (n = 42 cells from 12 embryos). E Time-lapse images showing the recruitment of Rasip1-Scarlet-I to the junctional patches and nascent junctional rings. Arrowheads mark Rasip1 clusters within the junctional patches. Arrows mark Rasip1 clusters localized at the periphery of junctional patches or within the nascent apical compartments. (E’) Automatic thresholding of Cdh5-Venus and Rasip1-Scarlet-I in (E). Black corresponds to the colocalized signals after thresholding. (E”) Graphic diagrams showing the localizations of signals in (E’). Consistent observations were made in 6 samples from 4 independent experiments. F Time-lapse images showing the recruitment of GFP-Podxl1 to the apical compartments. (F’) Intensities of Cdh5-Venus and GFP-Podxl1 along the dashed lines in (F). Consistent observations were made in 8 samples from 4 independent experiments. All scale bars: 5 μm. Source data are provided as a Source Data file.
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