Figure 7–27 The seven stripes of the protein encoded by the Even-skipped (Eve) gene in a developing Drosophila embryo. two and one-half hours after fertilization, the egg was fixed and stained with antibodies that recognize the eve protein (green) and antibodies that recognize the Giant protein (red). where eve and Giant proteins are both present, the staining appears yellow. At this stage in development, the egg contains approximately 4000 nuclei. the eve and Giant proteins are both located in the nuclei, and the eve stripes are about four nuclei wide. the pattern for the Giant protein is also shown in figure 7–26. (Courtesy of Michael levine.)
Figure 7–26 The nonuniform distribution of transcription regulators in an early Drosophila embryo. At this stage, the embryo is a syncytium; that is, multiple nuclei are contained in a common cytoplasm. Although not shown in these drawings, all of these proteins are concentrated in the nuclei. how such differences are established is discussed in Chapter 21.
start of segment
INSERT start of transcription
Figure 7–28 Experiment demonstrating the modular construction of the Eve gene regulatory region. (A) A 480-nucleotidepair section of the Eve regulatory region was removed and (B) inserted upstream of a test promoter that directs the synthesis of the enzyme β-galactosidase (the product of the E. coli LacZ gene—see figure 7–15). (C, D) when this artificial construct was reintroduced into the genome of Drosophila embryos, the embryos (D) expressed β-galactosidase (detectable by histochemical staining) precisely in the position of the second of the seven Eve stripes (C). β-Galactosidase is simple to detect and thus provides a convenient way to monitor the expression specified by a gene control region. As used here, β-galactosidase is said to serve as a reporter, since it “reports” the activity of a gene control region. (C and D, courtesy of Stephen Small and Michael levine.) the Drosophila Eve Gene is regulated by Combinatorial Controls
A detailed study of the stripe 2 regulatory module has provided insights into how it reads and interprets positional information. The module contains recognition sequences for two transcription regulators (Bicoid and Hunchback) that activate Eve transcription and for two transcription regulators (Krüppel and Giant) that repress it (Figure 7–29). The relative concentrations of these four proteins determine whether the protein complexes that form at the stripe 2 module activate transcription of the Eve gene. Figure 7–30 shows the distributions of the four transcription regulators across the region of a Drosophila embryo where stripe 2 forms. It is thought that either of the two repressor proteins, when bound to the DNA, will turn off the stripe 2 module, whereas both Bicoid and Hunchback must bind for this module’s maximal activation. This simple regulatory scheme suffices to turn on the stripe 2 module (and therefore the expression of the Eve gene) only in those nuclei located where the levels of both Bicoid and Hunchback are high and both Krüppel and Giant are absent—a combination that occurs in only one region of the early embryo. It is not known exactly how these four transcription regulators interact with coactivators and co-repressors to specify the final level of transcription across the stripe, but the outcome very likely relies on competition between activators and repressors that act by the mechanisms outlined in Figures 7–17, 7–19, and 7–23.