Xenbase Image ID: 131322

Fig. 8. Spacing of ciliated cells is imposed by intercalation. Notch signaling was blocked in the ectoderm of Xenopus embryos by injecting 2 ng of X-Su(H)-DBM RNA into two blastomeres of 4-cell-stage embryos, along with nLacZ RNA as a tracer. (A-C) At stage 17, a sample of embryos were fixed and the distribution and density of the a-tubulin-expressing cells was assessed by in situ hybridization with a-tubulin probe (dark blue) after detection of the tracer by X-gal staining (light blue). (C) A section through the embryo shown in A,B illustrates that blocking Notch signaling can result in several layers of a-tubulin-expressing cells. (B) The white dashed line indicates an area of the uninjected side of the embryo that has been populated by cells descending from the injected blastomeres as a result of cell mixing. (D-N) The remaining embryos were fixed at stage 25. In these embryos, the distribution of injected RNA was measured by staining with Magenta-gal (magenta), the density of a-tubulin-expressing cells was measured by in situ hybridization using just BCIP for detection (light blue), and the status of differentiated ciliated cells was measured by staining with the 6-11B1 antibody using HRP immunohistochemistry (brown). Note that BCIP detection gives rise to a higher level of background (diffuse sky blue staining) compared to NBT-BCIP detection. (D-H) Example of an embryo with a relatively mild increase in the number of a-tubulin-expressing cells. Lateral views are shown with anterior to the left. (F,G) Magnified views of the area framed in D and E, respectively, showing that all the a-tubulin-expressing cells have differentiated since they are also stained with 6- 11B1. The density of the differentiated ciliated cells approximately doubled on the injected side without affecting the morphology of the skin. (H) Transverse section through the embryo shown in D and E showing that the extra a-tubulin-expressing cells have reached the surface, resulting in a spacing pattern twice as dense on the injected side: seventeen ciliated cells can be counted on the injected side, versus nine on the uninjected side. nc, notochord; sm, somitic mesoderm; lm, lateral mesoderm; epi, epiderm. (I-N) An embryo with a severe overproduction of a- tubulin-expressing cells. (I,J,L,M) Lateral views are shown anterior to the left. (L,M) Magnified views of the area framed in I and J. Note the continuous pattern of a-tubulin staining on the injected side (compare I to J and L to M). The arrow points to a region where 6-11B1 Ab staining can be distinguished despite the high level of blue staining due to a-tubulin overexpression. (K,N) Sections through the embryo shown in I and J allow the detection of the 6-11B1 staining much better than the whole-mount view. (K) The injected area, marked by purple-stained nuclei, lies between the two arrowheads. Note that overproduction of a-tubulin-expressing cells in this area leads to a thickening and altered morphology of the epidermis. (N) A high-power view of an adjacent section shows that most of the a-tubulin-expressing cells have differentiated since they are also stained by 6-11B1 antibody whichever position they have in the ectoderm. The differentiated ciliated cells (blue and brown) that have reached the surface of the skin are surrounded by non-ciliated cells (asterisks). A fraction of the double-labeled cells were retained in the inner layer of the epidermis (arrowheads). Note that some of them are likely to be migrating to the surface (arrows) indicating that intercalation is an ongoing process. Image published in: Deblandre GA et al. (1999) Copyright © 1999. Image reproduced with permission of the publisher and the copyright holder. This is an Open Access article distributed under the terms of the Creative Commons Attribution License. Larger Image Printer Friendly View

Return to previous page