Analysis of Biological Development (K. Kalthoff)

Updates to Topic 11: Cell Adhesion and Morphogenesis

Answers to Questions in Text

Sorting Out of Mixed Amphibian Neurula Cells (p. 253/254)

  1. Why are amphibian embryos the most suitable vertebrate embryos for cell dissociation and reaggregation experiments? (Consider how the cells of amphibian and other vertebrate embryos obtain their nutrients.) Answer: In amphibian embryos, each cell has its own yolk supply, whereas the cells of other vertebrate embryos obtain nutrients from a central yolk reservoir or from a placenta.
  2. Which of the results obtained by Townes and Holtfreterseems odd and not easiy explained? Answer: The sorting out of mixed endoderm and mesoderm cells so that the mesoderm cells are surrounded by endoderm cells. This does not seem to reflect the position of endoderm cells relative to mesoderm cells in the living embryo. However, the endoderm cells form the inner lining of the gut, and the gut is connected to the outside world by the mouth and anus. The lumen of the gut can be viewed as part of the embryos external environment. In this perspective, the mesoderm is internal to the endoderm in vivo.

Cell Adhesiveness during Sea Urchin Gastrulation (p. 268)

  1. The force required to dislodge the test cells increased with the length of time that the test cells were allowed to settle on their substratum (cultured cells or ECM layer). The investigators decided to keep the settle-down time constant at 10 min. Why would this seem to be an appropriate interval, given the different types of connections formed between cells? Answer: 10 min are long enough for cell adhesion molecules to engage but too short for gap junctions, anchoring junctions, or tight junctions to develop.
  2. The particular adhesiveness of wandering mesenchyme cells to fibronectin suggests that the latter may be necessary for normal mesenchyme cells migration. How could this hypothesis be tested? How could researchers find out whether fibronectin is associated with the basement membrane on which newly ingressed primary mesenchyme cells migrate? Answer: by immunostaining.
  3. If cells that move relative to one another are generally less adhesive than stationary cells, which other cells in sea urchin embryos should then lose adhesiveness during gastrulation? Answer: archenteron cells during elongation.

Oriented Migration of Newt Mesoderm Cells on ECM Coat from Blastocoel Roof (p. 270)

  1. Could the orientation of the migrating mesoderm cells in the experiment shown in Fig. 11.22 have been an artifact introduced by the way in which the ECM coat of the blastocoel roof was transferred to the glass cover slips? For example, flushing the ectoderm piece off the glass coverslip might have sheared the deposited ECM coat, causing orientation of its fibers.
  2. What controls can you think of to settle the artifact problem raised in question 1? Answer: Yes, by flushing the blastocoel roof off the glass cover slip from different directions and monitoring whether the direction has an impact on the direction of migrating cells later. Conversely, a true biological cause for orientation of ECM fibrils in the blastocoel roof could be the mediolateral cell intercalation that drives convergent extension of the dorsal animal cap and non-involuting marginal zone. This hypothesis could be tested by depositing ECM from embryos that have been completely ventralized by UV irradiation and therefore don't show the convergent extension that characterizes the dorsal blastocoel roof. ECM coat from such embryos should not orient mesodermal cell migration.

Effect of Ultrabithorax Gene Activity on Cell Adhesion (pp. 273-275)

  1. The investigators injected the reaggregated cells into adult flies before tansferring them into larvae. Could they have simplified their experiments by leaving the reaggragates in adults for a longer time or by injecting the reaggragates straight into larvae? Answer: No. A long growth period is best provided by an adult host, and the larval host is required to force the imaginal disc implants through metamorphosis.
  2. In the mutant Contrabithorax (Cbx), the Ubx gene is active in the posterior T2, and posterior wings are replaced with posterior halteres. Do you expect cells from Cbx mutant T2 discs to form integrated mosaics with cells from wild-type T2 and/or T3 discs? If so, which parts of wing and/or haltere should be present in these integrated mosaics? Answer: Yes. Anterior wing mosaics are expected in reaggregates of T2(+) and T2(cbx). Posterior haltere mosaics are expected reaggregates of T3(+) and T2(cbx).


Clarifications and Corrections

p. 274, legend to Fig. 11.26, line 1 should read: "Control of cell affinity by the selector gene Ultrabithorax (Ubx)."

p. 276, legend to Fig. 11.27, line 6 should read: "... by antibodies to the cell adhesion molecule (N-cad) present on the ..."

New Review Articles

Baeg G.H. and Perrimon N. (2000) Functional binding of secreted molecules to heparan sulfate proteoglycans in Drosophila. Curr. Opin. Cell Biol. 12: 575-580

Brown N.H., Gregory S.L., and Martin-Bermudo M.D. (2000) Integrins as mediators of morphogenesis in Drosophila. Devel. Biol. 223: 1-16

Lilien J., Balsamo J., Arregui C. and Xu G. (2002) Turn-off, drop-out: Functional state switching of cadherin. Devel. Dyn. 224: 18-29

Schmidt A. and Hall A. (2002) Guanine nucleotide exchange factors for Rho GTPases: turning on the switch. Genes & Devel. 16: 1587-1609

Yagi T. and Takeichi M. (2000) Cadherin superfamily genes: functions, genomic organization, and neurologic diversity. Genes & Devel. 14: 1169-1180

New Research Articles

Farge E. (2003) Mechanical induction of twist in the Drosophila foregut/stomodeal primordium. Curr Biol. 13: 1365-1377.

This study examines whether developmental gene expression can be mechanically regulated by morphogenetic movements. The investigator analyzed the effects of mechanical stress on the activation of Twist, a patterning gene normally expressed only in the most ventral cells of the cellular blastoderm embryo of Drosophila. Transient 10% lateral deformation caused the ectopic expression of Twist around the entire dorsal-ventral axis, resulting in the ventralization of the embryo. This effect depended on nuclear translocation of Armadillo (catenin) protein. In gastrulating wild-type embryos, Twist is also expressed in the anterior foregut and stomodeal primordia. This expression is blocked in mutants that suppress the morphogenetic movement of germ-band extension. The mutants can be rescued by gentle compression of these cells, indicating that the stomodeal-cell compression normally caused by germ-band extension may induce the expression of Twist. Indeed, laser ablation of dorsal cells in wild-type embryos relaxed stomodeal cell compression and reduced Twist expression in the stomodeal primordium. Again, the activation of Twist in these cells depended on the nuclear translocation of Armadillo protein. The author concludes that anterior-gut formation is mechanically induced by the movement of germ-band extension through the induction of Twist expression in stomodeal cells.

Vasioukhin V., Bauer C., Yin M. and Fuchs E. (2000) Directed actin polymerization is the driving force for epithelial cell-cell adhesion. Cell 100: 209-219

Using primary keratinocytes (epidermis cells) from mouse embryos, which do not crawl over one another in culture, the investigators have observed a novel mechanism of intercellular adhesion. When stimulated by 2mM calcium in their medium, these cells form filopodia that penetrate and embed into neighboring cells. At the filopodia tips, E-cadherin clusters called puncta (sing. punctum) form double rows of adhesion points. Actin then polymerizes to merge the puncta into a single row, thus sealing the opposing membranes. In keratinocytes from mice lacking alpha-catenin, filopodia embed but actin polymerization does not occur, and the membranes do not seal.

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