7. How are mRNAs and proteins transported and localized?

29.6 Posterior development uses another localized regulator

Posterior development depends on the expression of a large group of genes. Embryos produced by females who are mutant for any one of these genes develop normal head and thoracic segments, but lack the entire abdomen. Some of these genes are concerned with exporting material from the nurse cells to the egg; others are required to transport or to localize the material within the egg.

Figure 29.8 The posterior pathway has two branches, responsible for abdominal development and germ cell formation.

The posterior pathway functions by a series of events in which one product is responsible for localizing the next. Figure 29.8 correlates the order of genes in the genetic pathway with the activities of their products in the embryo. The functions spir and capu are needed for Staufen protein to be localized at the pole. Staufen protein in turn localizes oskar RNA; possibly a complex of Staufen protein and oskar RNA is assembled. These functions are needed to localize Vasa, which is an RNA-binding protein. Its specificity and targets are not known.

If oskar is over-expressed or mislocalized in the embryo, it induces germ cell formation at ectopic sites. It requires only the products of vasa and tudor. This implies that all of the activities that precede oskar in the pathway are needed only to localize oskar RNA. The ability to form both pole cells and induce abdominal structures is possessed by oskar, in conjunction with vasa and tudor (and of course any components that are ubiquitously expressed in the egg). One effect of oskar function is to localize Vasa protein at the posterior end. The functions of valois and tudor are not known, but it is possible that valois is off the main pathway.

Figure 29.2 The early development of the Drosophila egg occurs in a common cytoplasm until the stage of cellular blastoderm.

Two types of pattern-determining event occur at the posterior pole, and the pathway branches at tudor. The polar plasm contains two morphogens: the posterior determinant (nanos) controls abdominal development; and an unidentified signal controls formation of the pole cells, which will give rise to the germline (see Figure 29.2). All of the posterior genes except nanos and pumilio are required for both processes, that is, they are defective in both abdominal development and pole cell formation. nanos and pumilio identify the abdominal branch. We do not know whether there are additional functions representing a separate branch for germ cell formation, or whether the pathway up to tudor is by itself sufficient (Sprenger and Nusslein-Volhard, 1992).

The posterior system resembles the anterior system in the basic nature of the morphogenetic event: a maternal mRNA is localized at the posterior pole. This is the product of nanos, and provides the morphogen. There are two important differences between the systems. Localization is more complex than in the case of the anterior system, because posterior determinants that originate in the nurse cells must be transported the full length of the oocyte to the far pole. And nanos protein acts to prevent translation of a transcription factor (hunchback). Its role is said to be permissive, since it functions to repress genes whose products would interfere with posterior development.

Figure 29.6 Mutant embryos that cannot develop can be rescued by injecting cytoplasm taken from a wild-type embryo. The donor can be tested for time of appearance and location of the rescuing activity; the recipient can be tested for time at which it is susceptible to rescue and the effects of injecting material at different locations.

How do we know that nanos is the morphogen at the end of the pathway? Rescue experiments (along the lines shown previously in Figure 29.6) with the mutants in the posterior group showed that in all but one case the cytoplasm of the nurse cell contained the posterior determinant (although it was absent from the posterior end of the oocyte itself). This indicates that these mutants all act in some subsidiary role, most probably concerned with transporting or localizing the morphogen in the egg. The exception was nanos, whose mutants did not contain any posterior-rescuing activity. Purified nanos RNA can rescue mutants in any of the other posterior genes, indicating that it is the last, or most downstream, component in the pathway. Indeed, injection of nanos RNA into ectopic locations in embryos can induce the formation of abdominal structures, showing that it provides the morphogen.

Figure 29.9 nanos products are localized at the posterior end of a Drosophila embryo. The upper photograph shows the tightly localized RNA inthe very early embryo (at the time of the 3rd nuclear division). The lower photograph shows the spreadingof nanos protein at the 8th nuclear division. Photographs kindly provided by Ruth Lehmann.

The upper part of Figure 29.9 shows the localization of nanos mRNA at the posterior end of an early embryo. But the localization poses a dilemma: nanos activity is required for development of abdominal segments, that is, for structures occupying approximately the posterior half of the embryo. How does nanos RNA at the pole control abdominal development? The lower part of Figure 29.8 shows that nanos protein diffuses from the site of translation to form a gradient that extends along the abdominal region.

Both bicoid and nanos act on the expression of the hunchback gene. hunchback codes for a repressor of transcription: its presence is needed for formation of anterior structures (in the region of the thorax), and its absence is required for development of posterior structures. It has a complex pattern of expression. It is transcribed during oogenesis to give an mRNA that is uniformly distributed in the egg. After fertilization, the hunchback pattern is changed in two ways. The bicoid gradient activates synthesis of hunchback RNA in the anterior region. And nanos prevents translation of hunchback mRNA in the posterior region; a result of this inhibition is that the mRNA is degraded. The anterior and posterior systems together therefore enhance hunchback levels in the anterior half of the egg, and remove it from the posterior half. We see later that the significance of this distribution lies with the genes that hunchback regulates. It represses the genes knirps and (probably) giant, which are needed to form abdominal structures. So the basic role of hunchback is to repress formation of abdominal structures by preventing the expression of knirps and giant in more anterior regions (Struhl et al., 1992).

Research

Sprenger, F. and Nusslein-Volhard, C. (1992). Cellular terminal regions of the Drosophila egg. Cell 71, 987-1001.

Struhl, G., Johnston, P., and Lawrence, P. A. (1992). Control of Drosophila body pattern by the hunchback morphogen gradient. Cell 69, 237-249.