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The Drosophila fushi tarazu (ftz) upstream element is an enhancer-like element that is required for the correct expression of fez in developing embryos and that directs transcription from a minimal promoter in a ftz-like seven-striped pattern. Using a deletion analysis, we have identified several independent c/s-regulatory elements in the upstream element. A distal enhancer directs fusion gene expression in seven stripes primarily in the mesoderm. A more complex proximal enhancer contains a mesodermally active element and a second element with which it interacts to generate seven stripes in the ectoderm. Striped expression directed by each enhancer is ftz-dependent, and each contains binding sites for purified ftz homeo domain. We suggest that ftz protein acts in combination with germ layer-restricted transcription factors directly and positively to regulate the transcription of its own gene. The development of a complex organism from a fertilized egg requires the differential expression of genetic information in a temporally and spatially restricted fashion. This differential expression is frequently regulated at the level of transcription initiation (for reviews, see Maniatis et al. 1987;Mitchell and Tjian 1989). In such cases, cis-acting DNA sequences mediate interactions between trans-acting protein factors and RNA polymerase II or the "general transcription machinery" to activate transcription. Promoter elements, located close to the transcription start site, are required for efficient initiation and positioning of the start site, whereas upstream regulatory elements or enhancers act to increase the rate of transcription from a given promoter. Enhancers are characterized by their abilities to (1) stimulate transcription over large and varying distances, (2) act in an orientation-independent fashion, and (3) stimulate transcription of heterologous promoters (Serfling et al. 1985). For both viral and cellular genes, discrete cisacting regulatory elements have been identified that direct transcription in a cell type-restricted fashion. Although some cell type-specific enhancers have been studied in detail (Atchison 1988), only a small number have been examined in their native cellular environments in transgenic animals (Posakony et al. 1985;Garabedian et al. 1986;Hammer et al. 1987; Pinkert et al. 1987; Johnson et al. 1989;Logan et al. 1989). Much of what we know about enhancer function comes from studies of viral model systems. Analysis of the wellcharacterized viral SV40 enhancers suggests that the basic building blocks of enhancers are short "enhansons" that correspond to the binding sites for transacting factors (Ondek et al. 1988}. Different enhancer motifs may be active in different cell types, suggesting that the availability of cell type-specific transcription factors may control the specificity of enhancer activation {Schirm et al. 1987). Given the complexity of promoter and enhancer elements, it is possible to construct a scenario in which transcriptional specificity is determined by the combinatoria...

We have identified a sine oculis gene in the planarian Girardia tigrina (Platyhelminthes; Turbellaria; Tricladida). The planarian sine oculis gene (Gtso) encodes a protein with a sine oculis (Six) domain and a homeodomain that shares significant sequence similarity with so proteins assigned to the Six-2 gene family. Gtso is expressed as a single transcript in both regenerating and fully developed eyes. Whole-mount in situ hybridization studies show exclusive expression in photoreceptor cells. Loss of function of Gtso by RNA interference during planarian regeneration inhibits eye regeneration completely. Gtso is also essential for maintenance of the differentiated state of photoreceptor cells. These results, combined with the previously demonstrated expression of Pax-6 in planarian eyes, suggest that the same basic gene regulatory circuit required for eye development in Drosophila and mouse is used in the prototypic eye spots of platyhelminthes and, therefore, is truly conserved during evolution.homeobox ͉ eye morphogenesis ͉ platyhelmint ͉ eye evolution T he study of the genetic network that regulates the development of the Drosophila visual system has resulted in the identification of several transcription factors and other nuclear proteins that are required for the specification of early eye morphogenesis (1-4). These factors seem to act in a hierarchy in which sine oculis (so) is regulated directly by Pax-6 (5, 6), the master control function. In turn, so requires eyes absent (eya), encoding a nuclear protein (7), to induce ectopic eyes (4). This genetic pathway has been established in Drosophila (8), but homologous proteins also regulate eye development in vertebrates, suggesting that this regulatory network is old, is conserved in evolution, and has been adapted to the control of development of different visual systems found in both clades (9). Both the identification and functional characterization of homologous genes in more primitive organisms, such as the platyhelminthes, will help to clarify the age and extent of conservation of this genetic cascade.Sine oculis is a homeobox-containing gene that is required for the development of the visual system in Drosophila (10, 11). A murine homologue, Six3, is expressed in the developing eye (12). In both of these model systems, so and Six are expressed early in eye development as well as in other structures. Combined overexpression of so and eya in Drosophila induces ectopic eyes (4), whereas, in vertebrates, Six3 overexpression results in ectopic lens formation (13,14). Planarians (Platyhelminthes; Turbellaria; Tricladida) are located at the base of the Lophotrochozoa Protostomia clade (15, 16). The eye spots of planarians are one of the most ancestral and simple types of visual systems, close to the prototypic eye proposed by Charles Darwin (see ref.8). The planarian eye spots consist of two cell types: a bipolar nerve cell with a rhabdomere as a photoreceptive structure and a cup-shaped structure composed of pigment cells (17). During head regeneration, new e...