pupal case. For example, juvenile hormone, an isoprenoid, readily passes through the cuticle. However, water-soluble test chemicals would have to be included in the food fed to 3- and 4-day-old larvae (third instar) or injected into pupae or flies (see later discussion of successful cases).

Assays with Sensitized Pathways

Because Drosophila has already shown the utility of sensitized screens for detecting new components in developmental pathways, there is little doubt that appropriate genetic stocks can be developed that would be useful for identifying potential toxic compounds. Such compounds would formally act like secondary mutations in reducing or increasing the activity of some other pathway component. In cases in which the sensitized strains are vigorous, as was the case for the sevenless mutants in the Simon et al. (1991) screen, mutant flies can be used without further genetic manipulations. In many instances, however, the sensitized strains are weak due to the fact that the signaling pathway is used for multiple functions, resulting in reduced viability. In these cases, the FRT-FRP system for producing clones of cells (see Figure 7-4), homozygous for sensitizing mutations, will be extremely useful. Careful construction of strains will also provide for “twin clones,” one for the homozygous mutation and the other for clones homozygous for the wild-type genes. This twin clone will provide a record of the loss of the mutant clone in these instances where the homozygous mutant cells become inviable due to chemical treatment.

The compound eye and the wing are two excellent organs for the analysis of potential toxic chemicals. Both are nonvital organs under laboratory conditions, and indeed eyeless and wingless flies are viable and fertile. Both organs are easily scored for developmental effects. Both have a relatively large precursor population so that the frequency of clones is relatively high, if the FRT-FRP system is used. Finally, many of the major signaling pathways function in establishing the regular pattern. A few examples will illustrate these features.

Ras Signaling in Eye Development. Earlier in this chapter, the committee described an example of a screen that uses a sensitized genetic background to identify genes in the receptor tyrosine kinase pathway involved in Drosophilia eye development (Simon et al. 1991). The use of sensitized systems to reveal chemical modifiers has in fact been well demonstrated for the Drosophila eye. Peptidomimetics that block isoprenylation interfere with RAS membrane binding and activity and, as shown in Figure 7-5, when injected into flies, can abrogate the abnormal phenotype of the activating RAS val12 mutation (Kauffmann et al. 1995). The same agents block activated Ras-induced tumors in mice, confirming cross-species relevance of the assay used as a chemical and genetic indicator.



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