13.6 From Flies to Fingers, Feathers, and Floor Plates: The Many Roles of Individual Toolkit Genes

We have seen that toolkit proteins and regulatory RNAs have multiple roles in development. For example, recall that the Ultrabithorax protein represses limb formation in the fly abdomen and promotes hind-wing development in the fly thorax. Similarly, Sloppy-paired and Engrailed participate in the generation of the basic segmental organization of the embryo and collaborate with Hox proteins to suppress limb formation. These roles are just a few of the many roles played by these toolkit genes in the entire course of fly development. Most toolkit genes function at more than one time and place, and most may influence the formation or patterning of many different structures that are formed in different parts of the larval or adult body. Those that regulate gene expression may directly regulate scores to hundreds of different genes. The function of an individual toolkit protein (or RNA) is almost always context dependent, which is why the toolkit analogy is perhaps so fitting. As with a carpenter’s toolkit, a common set of tools can be used to fashion many structures.

To illustrate this principle more vividly, we will look at the role of one toolkit protein in the development of many vertebrate features, including features present in humans. This toolkit protein is the vertebrate homolog of the Drosophila hedgehog gene. The hedgehog gene was first identified by Nüsslein-Volhard and Wieschaus as a segment-polarity gene. It has been characterized as encoding a signaling protein secreted from cells in Drosophila.

Figure 13-26: The Sonic hedgehog toolkit gene has multiple roles
Figure 13-26: The Shh gene is expressed in many different parts of the developing chick embryo (indicated by dark staining), including (a) the zone of polarizing activity in each of the two developing limb buds and the long neural tube and (b) the developing feather buds. Shh mRNA is visualized by in situ hybridization.
[Photomicrographs courtesy of (a) Cliff Tabin and (b) Dr. John Fallon, University of Wisconsin, and Matthew Harris, Harvard Medical School, Department of Genetics.]

As the evidence grew that toolkit genes are common to different animal phyla, the discovery and characterization of fly toolkit genes such as hedgehog became a common springboard to the characterization of genes in other taxa, particularly vertebrates. The cloning of homologous genes based on sequence similarity (see Chapter 14) was a fast track to the identification of vertebrate toolkit genes. The application of this strategy to the hedgehog gene illustrates the power and payoffs of using homology to discover important genes. Several distinct homologs of hedgehog were isolated from zebrafish, mice, chickens, and humans. In the whimsical spirit of the Drosophila gene nomenclature, the three vertebrate homologs were named Sonic hedgehog (after the video-game character), Indian hedgehog, and Desert hedgehog.

One of the first means of characterizing the potential roles of these genes in development was to examine where they are expressed. Sonic hedgehog (Shh) was found to be expressed in several parts of the developing chicken and other vertebrates. Most intriguing was its expression in the posterior part of the developing limb bud (Figure 13-26a). This part of the limb bud was known for decades to be the zone of polarizing activity (ZPA) because it is an organizer responsible for establishing the anteroposterior polarity of the limb and its digits (Figure 13-26b). To test whether Shh might play a role in ZPA function, Cliff Tabin and his colleagues at Harvard Medical School caused the Shh protein to be expressed in the anterior region of developing chick limb buds. They observed the same effect as transplantation of the ZPA—the induction of extra digits with reversed polarity. Their results were stunning evidence that Shh was the long-sought morphogen produced by the ZPA.

Shh is also expressed in other intriguing patterns in the chicken and other vertebrates. For example, Shh is expressed in developing feather buds, where it plays a role in establishing the pattern and polarity of feather formation (see Figure 13-26b). Shh is also expressed in the developing neural tube of vertebrate embryos, in a region called the floor plate (see Figure 13-26a). Subsequent experiments have shown that Shh signaling from these floor-plate cells is critical for the subdivision of the brain hemispheres and the subdivision of the developing eye into the left and right sides. When the function of the Shh gene is eliminated by mutation in the mouse, these hemispheres and eye regions do not separate, and the resulting embryo is cyclopic, with one central eye and a single forebrain (it also lacks limb structures).

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The dramatic and diverse roles of Shh are a striking example of the different roles played by toolkit genes at different places and times in development. The outcomes of Shh signaling are different in each case: the Shh signaling pathway will induce the expression of one set of genes in the developing limb, a different set in the feather bud, and yet another set in the floor plate. How are different cell types and tissues able to respond differently to the same signaling molecule? The outcome of Shh signaling depends on the context provided by other toolkit genes that are acting at the same time.

KEY CONCEPT

Most toolkit genes have multiple roles in different tissues and cell types. The specificity of their action is determined by the context provided by the other toolkit genes that act in combination with them.