QUICK SEARCH:   [advanced] Author: Keyword(s):
Home     Help     Feedback     Subscriptions     Archive     Search     Table of Contents

doi: 10.1242/10.1242/dev.00490

This Article Summary Full Text Full Text (PDF) Movie Alert me when this article is cited Alert me if a correction is posted Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Chalmers, A. D. Articles by Papalopulu, N. Search for Related Content PubMed PubMed Citation Articles by Chalmers, A. D. Articles by Papalopulu, N.

Oriented cell divisions asymmetrically segregate aPKC and generate cell fate diversity in the early Xenopus embryo

Andrew D. Chalmers^*, Bernhard Strauss^* and Nancy Papalopulu

Wellcome Trust/Cancer Research UK Institute, Tennis Court Road, Cambridge CB2 1QR, UK
Department of Anatomy, Downing Street, University of Cambridge, Cambridge, UK

View larger version (81K): [in a new window]   Fig. 1. Generation of the deep cells during development. Histological sections show when the deep cells are generated from the initially single layered embryo. Arrow indicates first deep cell.

View larger version (59K): [in a new window]   Fig. 2. Three orientations of cell division occur in early Xenopus development. (A-D) Stained mitotic spindles (red) of whole-mount embryos showed three orientations of division. The division plane was extrapolated based on the observation that the spindle orientates at 90° to future division plane (Strome, 1993). Chromosomes are shown in yellow. (A) top view of parallel and perpendicular divisions; (B) side view showing parallel, oblique and perpendicular divisions. (C) side view showing parallel and perpendicular divisions in anaphase. (D) Side view showing oblique division in anaphase. Blue arrows, parallel spindles. Red arrows, perpendicular spindles. Green arrows, oblique spindles. (E) Schematic showing the three orientations of division. Parallel divisions generate two superficial daughter cells. Oblique divisions generate one superficial cell with a large and one superficial cell with a small external surface. Perpendicular divisions generate a superficial and a deep cell. (F) The three orientations of division can be seen in isolated blastomeres judged by the distribution of the pigmented, originally external, surface.

View larger version (62K): [in a new window]   Fig. 3. Perpendicular divisions start in the 32-cell embryo and continue up to stage 9. For each stage, a whole-mount embryo (A,D,G,J,M,P), and a top (B,E,H,K,N,Q) and side (C,F,I,L,O,R) view of whole-mount embryos with stained mitotic spindles (red) and DNA (yellow) are shown. Red arrows indicate perpendicular divisions. Preparations in F, O and R still have their vitelline membrane.

View larger version (64K): [in a new window]   Fig. 4. Three orientations of divisions can be seen in timelapse analysis of cleavage stage embryos. (A-C) Examples of each kind of division are shown. The colour code depicts the type of division that a cell is about to undergo (blue indicates parallel, green indicates oblique and red indicates perpendicular). (D) Corresponding still images from the timelapse movie. All panels are views from the animal pole and dorsal is towards the top.

View larger version (63K): [in a new window]   Fig. 5. Distribution and numbers of the three orientations of division shown by timelapse analysis. (A) Still images from one movie are labelled to show the orientation each cell is about to undergo (see Movie at http://dev.biologists.org/supplemental/) (blue indicates parallel, green indicates oblique and red indicates perpendicular). (B) Percent of each orientation of division from the analysis of 10 timelapse movies. Number of cells refers to the cell-stage of the embryos. (C) Variable distribution of different orientations of division in 4x 128-cell embryos. Dorsal is towards the top. (D) Percent of each type of division that occurs in all superficial daughters of the left/dorsal (LD), right dorsal (RD), left ventral (LV) and right ventral (RV) blastomere (left), from eight-cell up to the 1024-cell stage.

View larger version (68K): [in a new window]   Fig. 6. Orientation of division correlates with cell shape. (A) Percentage of each type of oriented division that the daughters of an oblique division, one with a large (`oblique big') and one with a small (`oblique small') external surface, will undergo in the next division is shown. A high percentage (88%) of oblique small cells divided perpendicularly in the next division. The percentages are based on n=184 `oblique big' cells and n=180 `oblique small' cells from five embryos (eight-cell to 1024-cell stage). (B) Wholemount and sections from the same embryo showing that cells with a small external surface (labelled 1 and 2) have a long internal axis. (C) Unstained section showing that cells with small external surface (arrow) have an elongated apicobasal axis. (D,E) Antibody staining of spindles (red) and DNA (yellow/green) showing that elongated cells with small external surface (arrows) divide in a perpendicular orientation.

View larger version (121K): [in a new window]   Fig. 7. aPKC is apically localised and asymmetrically inherited during the perpendicular divisions. (A,B,E) Stage 8 aPKC localisation. (C,F) Stage 8 occludin localisation. (D) No primary antibody negative control. (G,H) Stage 8 aPKC and a tubulin double staining. (I) Transmitted light image of fertilised egg. (J,K) aPKC localisation in fertilised egg. In this case, a pigmented embryo was used to allow identification of animal hemisphere. (L,M) aPKC staining at the four-cell stage. (N) No primary antibody negative control at the four-cell stage. (A-H) No vitelline membrane. In I-M, embryos with vitelline membrane are shown (as they were less damaged), but the staining was the same in those without vitelline membrane. All images are from antibody stained sections, except G,H, which are stained as whole-mount preparations.

View larger version (46K): [in a new window]   Fig. 8. Perpendicular divisions generate molecularly distinct daughter cells. (A-C) Perpendicular divisions in isolated blastomeres generate one daughter cell with localised aPKC and one daughter cell without. Blastomeres were stained with aPKC (red) and ß1 integrin (green). A transmitted light image is shown below the fluorescent image. (A) An isolated blastomere. (B) An isolated blastomere during a perpendicular division. (C) Superficial (top) and deep (bottom) daughters generated after an isolated blastomere completes a perpendicular division. (D,E) The divisions generate differences that lead to later differences in gene expression. (D) An isolated 64-cell stage blastomere was left to divide, separated into a deep and a superficial cell, and cultured until control embryos reach stage 10. (E) Gene expression of the cultured blastomeres was analysed by quantitative real time RT-PCR. The y-axis shows expression in arbitrary units, with 100 being the same level as a stage 10 animal cap. Odc and XSox3 are equally expressed, while ESR6e is preferentially expressed in outer cell derived clones. The mean of three independent experiments is presented.