Dynamic Development

CONTENTS

Main Page Dynamic Development

The Foundations of Developmental Biology

Gametogenesis

From Sperm and Egg to Embryo

Genetic Regulation of Development

Organizing the Multicellular Embryo

Generating Cell Diversity


Dynamic Development at a Glance


Learning Resources

Research Resources

The Developmental Biology Journal Club

Developmental Biology Tutorial

The Origins of Polarity and Embryo Patterning in Drosophila

Which came first, the fly or the egg?

Extensive genetic analysis in Drosophila has identified maternal effect genes, which are transcribed during oogenesis and exert their phenotypic effects during development. They are typically involved in organization of the body plan of the embryo. How can expression of genes during oogenesis leave a molecular imprint on the egg? We shall briefly examine the establishment of the body axes in Drosophila and will return to the important topic of body plan organization later.

Anterior-posterior asymmetry is evident before dorsal-ventral asymmetry (Gavis, 1995). Polarization is evident by the localization of the bicoid and oskar transcripts to opposite ends of the oocyte. (For localization of oskar transcripts, see Fig. 1, below.) The localization of these transcripts is dependent upon the oocyte cytoskeleton (Micklem, 1995). Each egg chamber contains a single interconnecting microtubule complex. During early oogenesis, the microtubule organizing center (minus end) is located at the oocyte's posterior end with its plus end extending into the nurse cells. That complex is then dismantled, and a new one is assembled, with the microtubule-nucleating activity at the anterior end and the plus ends of the microtubules extending toward the posterior pole. Both the transport and anchoring of bicoid and oskar transcripts are MT-dependent.


Figure 1. In situ hybridization to whole mount Drosophila egg chamber. Oskar mRNA is synthesized in the polyploid nurse cells (left), transported into the oocyte (right) and localized to the posterior pole of the oocyte. (Reproduced with permission of Dr. Ruth Lehmann. Visit Dr. Lehmann's Web page.)

Organization of the microtubule network depends upon localization of the oocyte to the posterior end of the egg chamber early in oogenesis. A signal produced by the oocyte reaches only the follicle cells at that end. A subset of them (the polar cells) respond to the signal and differentiate with posterior fates. The posterior polar cells, in turn, signal back to the oocyte to set up the anterior-posterior axis of the oocyte by reorganizing the microtubule cytoskeleton (González-Reyes and St Johnston, 1994).

The signal from the oocyte to the follicle cells depends upon gurken. gurken mRNA localizes to the posterior end of the oocyte during early oogenesis. gurken encodes a TGF alpha-like signaling molecule that has an epidermal growth factor (EGF) repeat element. The receptor in the follicle cells is an EGF receptor-like molecule called top/DER. Any disruption of this signaling due to mutation prevents determination of the posterior follicle cells. The oocyte microtubule cytoskeleton fails to become polarized, bcd mRNA becomes distributed to both the anterior and posterior poles, and osk mRNA is not localized to the posterior pole (Gonzalés-Reyes and St. Johnston, 1994).

Remember that oocyte trancripts originate in the nurse cells. They are transported into the oocyte, apparently directed by a microtubule minus-end-directed motor. The reorganization of the microtubule network redirects the minus ends to the anterior end of the oocyte and moves the transcripts to the anterior end of the oocyte. Most subsequently become uniformly distributed, with the exception of those that are to become localized (i.e., bicoid, oskar, gurken and cyclin-B). gurken signaling occurs early while the microtubule network has its minus ends in the oocyte. That signaling subsequently reverses the polarity of the network.

Dorsal-ventral patterning has been shown to be dependent upon a similar communication mechanism, and the key signaling molecules involved in that process have been identified. Dorsalization is first detectable in the oocyte when the nucleus moves to the anterior/dorsal corner of the oocyte as a consequence of microtubular function. The nucleus is accompanied by gurken transcripts. As a consequence, Gurken protein is synthesized in this localized region of the oocyte. The localization of Gurken protein is shown below.


Confocal fluorescence micrograph showing the distribution of Gurken protein (green; yellow where it co-localizes with cortical actin) on the presumptive anterior/dorsal side of the developing Drosophila oocyte.
(Photo courtesy of Dr. Trudi Schupbach. Image copyright © 1997, Trudi Schupbach.)

Dorsalization occurs in response to a signal produced by gurken (Micklem, 1995). The grk signal, which is produced by the oocyte, is received by follicle cells via Top/DER. It has been proposed that the binding of Gurken protein to Top/DER protein in the adjacent follicle cells activates a receptor tyrosine kinase signal transduction cascade. Follicle cells that receive the grk signal acquire dorsal fates, whereas the remainder become ventrally-inclined. Dorsal-ventral differentiation of the follicle cells causes ventral activation of a second signaling pathway, by which a signal is transmitted to the fertilized embryo to produce the gradient of Dorsal protein in embryonic nuclei.

(See Browder et al., Fig. 6.13) Gilbert, 1997, Fig. 14.37; Kalthoff, 1996, Fig. 21.34; Wolpert et al., 1998, Fig. 5.12, 5.14)

Gonzalés-Reyes et al. (1995) demonstrated that grk mRNA is not localized when the GV fails to migrate (in top /DER mutant egg chambers). The oocyte microtubule cytoskeleton is not properly polarized in egg chambers of these mutants, which lack both A-P and D-V patterning. These observations indicate that the formation of the D-V axis depends upon the prior polarization of the A-P axis, which polarizes the microtubule cytoskeleton that translocates the oocyte nucleus and the grk mRNA.

The displacement of mRNA molecules by the cytoskeleton is an interesting phenomenon. Once they have been translocated, they must remain in place in order to produce localized protein product. Localized RNAs in Drosophila appear to be associated with the oocyte cortex.

The requirement of microtubules for RNA transport is shown by using the microtubule inhibitor colchicine. As shown by Pokrywka and Stephenson (1995), oskar mRNA localization is abolished by treatment with colchicine. Other transcripts, however, either are not localized or have different destinations. The differences between transcripts imply that individual RNA molecules may have "postal codes" (zip codes for the Americans) that direct them to particular addresses within the cell (and attendants that keep them there). The delivery system that reads and utilizes the postal code is not well understood, but it has the ability to select among transcripts. For example, exuperantia and swallow are required for localization of bicoid. However, oskar mRNA is localized normally in exuperantia and swallow mutant egg chambers. Presumably, proteins that anchor specific transcripts to the microtubules are involved. Glotzer et al. (1997) have also demonstrated that colchicine prevents localization of oskar mRNA if labeled transcripts were injected into oocytes at sites distal to the posterior pole. However, when transcripts were injected close to the posterior pole, the injected mRNA localized even in the presence of colchicine. This result suggests that long-range transport of the messenger is dependent upon microtubules but that microtubule-independent processes are involved in short-range transport and anchoring of osk mRNA to the posterior pole.

What do these proteins respond to on mRNA? What is the postal code comprised of? In the case of osk and bcd mRNAs, distinct elements in the 3' UTRs have been identified that direct their transport and localization.

The localization of oskar mRNA has important implications for formation of the pole plasm. The pole plasm contains elements that are necessary for translation of nanos mRNA. This establishes a gradient in Nanos protein that is necessary for patterning of the posterior of the embryo. The pole plasm also contains determinants of the germ line. Oskar protein induces pole plasm assembly (Ephrussi and Lehmann, 1992). If osk mRNA is mislocalized to the anterior pole, it induces polar plasm there. However, in mutant ovaries in which oskar mRNA is not localized, ectopic pole plasm does not form (Ephrussi and Lehmann, 1992). These results suggest that unlocalized osk mRNA fails to be translated and that localization is necessary for its translation.

Why are unlocalized oskar transcripts inactive? The evidence suggests that their translation is repressed outside the posterior domain. This repression is dependent upon the binding of a protein called Bruno to elements in the 3' UTR of oskar mRNA. In the absence of a functional bruno gene, oskar is translated prematurely. The Oskar protein that is synthesized accumulates throughout the oocyte, causing posteriorization of the embryo (Kim-Ha et al., 1995).


Learning Objectives

  • Summarize the roles of the cytoskeleton in establishment of the asymmetry of the Drosophila oocyte. Document the experimental evidence to support these roles.
  • What is the role of the posterior polar cells in polarization?
  • What is gurken? Describe the odyssey of gurken mRNA during oogenesis.
  • What is the first overt sign of dorsalization of the oocyte?
  • What is top/Der?
  • What happens to the distribution of gurken mRNA in top/Der mutants?
  • What are the developmental implications of the localization of oskar mRNA?
  • Why are unlocalized oskar transcripts inactive?


Digging Deeper:

Additional Reading

Markussen, F.H., Michon, A.M., Breitwieser, W. and Ephrussi, A. 1995. Tranlsational control of oskar generates short OSK, the isoform that induces pole plasm assembly. Development 121: 3723-3732.

Neuman-Silberberg, F.S. and Schupbach, T. 1996. The Drosophila TGF-alpha-like protein Gurken: expression and cellular localization during Drosophila oogenesis. Mech. Dev. 59: 105-113.

Newmark, P.A., Mohr, S.E., Gong, L. and Boswell, R.E. 1997. mago nashi mediates the posterior follicle cell-to-oocyte signal to organize axis formation in Drosophila. Development 124: 3197-3207.

Ray, R.P. and Schupbach, T. 1996. Intercellular signaling and the polarization of body axes during Drosophila oogenesis. Genes & Dev. 10: 1711-1723.

Wilson, J.E., Connell, J.E. and Macdonald, P.M. 1996. aubergine enhances oskar translation in the Drosophila ovary. Development 122: 1631-1639.


References

Browder, L.W., Erickson, C.A. and Jeffery, W.R. 1991. Developmental Biology. Third edition. Saunders College Pub. Philadelphia.

Ephrussi, A. and Lehmann, R. 1992. Induction of germ cell formation by oskar. Nature 358: 387-392.

Gavis, E. R. 1995. Gurken meets torpedo for the first time. Current Biology 5: 1252-1254.

Gilbert, S.F. 1997. Developmental Biology. Fifth Edition. Sinauer. Sunderland, MA.

Glotzer, J.B., Saffrich, R., Glotzer, M. and Ephrussi, A. 1997. Cytoplasmic flows localize injected oskar RNA in Drosophila oocytes. Curr. Biol. 7: 326-337.

Gonzáles-Reyes and St Johnson, D. 1994. Role of oocyte position in establishment of anterior- posterior polarity in Drosophila. Science 266: 639-642.

Gonzáles-Reyes, A., Elliott, H. and St Johnson, D. 1995. Polarization of both major body axes in Drosophila by gurken-torpedo signalling. Nature 375: 654-658.

Kalthoff, K. 1996. Analysis of Biological Development. McGraw-Hill. New York.

Kim-Ha, J., Kerr, K. and Macdonald, P.M. 1995. Translational regulation of oskar mRNA by bruno, an ovarian RNA-binding protein, is essential. Cell 81: 403-412.

Micklem, D.R. 1995. mRNA localisation during development. Dev. Biol. 172: 377-395.

Pokrywka, N. J. and Stephenson, E. C. 1995. Microtubules are a general component of mRNA localization systems in Drosophila oocytes. Developmental Biology 167: 363-370.

Wolpert, L., Beddington, R., Brockes, J., Jessell, T., Lawrence, P. and Meyerowitz, E. 1998. Principles of Development. Current Biology. London.


Dynamic Development at a Glance
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Leon Browder & Laurie Iten (Ed.) Dynamic Development
Last revised Wednesday, June 17, 1998