Skip to main content
. 2018 Jun 19;7:e37131. doi: 10.7554/eLife.37131

Figure 3. Lmx1bb is necessary for development of the ES’s ability to form breaks in its diffusion barrier and deflate.

(A) Lateral view of wild-type and lmx1bbjj410/jj410 mutant ears imaged by bright-field microscopy at 80 hpf, asterisk labels greatly enlarged mutant ES. Scale bar, 100 μm. (B) Slices from 3D confocal time course of an lmx1bb transcriptional reporter (cyan, Tg(lmx1bb:egfp)mw10/mw10; yellow, Tg(actb2:mem-mcherry2)hm29), n = 3. (C) Slices and select time points from 3D confocal time course of lmx1bbjj410/jj410 mutant embryos. Membrane (green) from ubiquitous membrane citrine transgenes. Perilymph (magenta) from 3 kDa dextran-Texas red, n = 4. (D) Quantification of segmented ES volumes (primary axis, green) and leak-in fluorescence (secondary axis, magenta) from lmx1bbjj410/jj410 time course in (C) (see also Figure 3—figure supplement 1 and Videos 56). (E) 3D transverse view (endolymph in yellow) from timelapse showing endolymph in dilated mutant ES, outlined with dashed blue line, n = 2. (F) Small regions with thin membranes (asterisks) form in the inflated ES of wild-type but not lmx1bb mutants. (G) Quantification of minimum epithelial thickness versus inflated ES volume in mutant (plotted in red, n = 9) and wild-type (plotted in black, n = 14). Compiled from 65 to 80 hpf embryos. (H) Uneven labeling from Tg(lmx1bb:egfp) reveals thin basal processes (white arrow). (I) Wild-type ES examples with sparsely labeled cells: membrane-labeled citrine (green) in a membrane-labeled cherry background (magenta), white arrows indicate lamellar projections, n = 15. (J) lmx1bbjj410/jj410 mutant ES examples with sparsely labeled cells: membrane cherry (magenta) in a membrane citrine background (green), n = 9. (K) Cartoon schematic of apico-basal organization of ES. (L) Supporting whole-mount immuno-stains for basal and apical markers (collagen and ZO-1, both magenta) in a membrane-labeled citrine background (green), n = 16, 15, 6, 32, 21, and 12 for (i-vi). Scale bars in (B-L), 10 μm.

Figure 3.

Figure 3—figure supplement 1. Inflation of additional mutant ES.

Figure 3—figure supplement 1.

Quantification of segmented ES volumes (primary axis, green) and leak in fluorescence (secondary axis, magenta) from an additional time-lapse of an lmx1bbjj410/jj410 mutant (see Video 6).