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Fig. 8
Planar polarized distribution of Crumbs2b-mediated adhesion organizes differentiating cones into linear columns, and column fragments are retained in tbx2b mutants.
A, C, E) Confocal images of retinal flat-mounts immunostained for ZO-1 (white or cyan) and focused at the level of the OLM. B, D, F) Confocal images of retinal flat-mounts immunostained for Crb2b and focused at the level of the inner segments of cone photoreceptors. A) Retinal margin from a wild-type adult double transgenic (UV and blue cone reporters); proliferative germinal zone at the right edge. Cohorts of cones generated nearly synchronously at the germinal zone form straight columns; rows of blue cones and UV cones alternate with rows of red/green double cones (ZO-1 profiles). Expression of the blue cone reporter (blue) is initiated in differentiating cones (blue arrows) as the column emerges from the germinal zone. Cone profiles in the predicted position of UV cones (magenta arrows) emerge from the germinal zone prior to onset of the UV cone reporter. B) Retinal margin from a wild-type adult with the blue cone reporter; some UV cones, identified by their position in the mosaic lattice, are indicated by magenta dots. Crb2b immunostaining (yellow) is localized to the flattened boundaries between inner segments of five adjacent cones (green-red-blue-red-green) along a column, but not between columns; UV cones (magenta dots) are in the gaps separating the pentamers. Polarized Crb2b is in linear segments oriented orthogonal to the retinal margin in the earliest cone columns (arrows). C, D) Retinal margin from a tbx2b mutant double immunolabeled for ZO-1 (C) in white and Crb2b (D) in yellow; inset shows overlay of ZO-1 and Crb2b. Note that Crb2b is already polarized approximately orthogonal to the margin in the earliest cones. E) Dorsal retinal margin from a double transgenic (UV and blue cone reporters) tbx2b mutant adult with ZO-1 (white). F) Double immunolabeled tbx2b mutant. Left panel: ZO-1 (cyan) at the level of the OLM superimposed on Crb2b (yellow) at the level of the inner segments (3–6 μm apical to the OLM). Right panel: Level of Crb2b only. Dashes outline column fragments. Scale bars: 10 μm (A) and (B–F).