Walter J. Gehring

Pax 6 : mastering eye morphogenesis and eye evolution

Pax 6 genes from various animal phyla are capable of inducing ectopic eye development, indicating that Pax 6 is a master control gene for eye morphogenesis. It is proposed that the various eye-types found in metazoa are derived from a common prototype, monophyletically, by a mechanism called intercalary evolution.

twin of eyeless, a Second Pax-6 Gene of Drosophila, Acts Upstream of eyeless in the Control of Eye Development

The Drosophila Pax-6 gene eyeless (ey) plays a key role in eye development. Here we show tht Drosophila contains a second Pax-6 gene, twin of eyeless (toy), due to a duplication during insect evolution. Toy is more similar to vertebrate Pax-6 proteins than Ey with regard to overall sequence conservation, DNA-binding function, and early expression in the embryo, toy and ey share a similar expression pattern in the developing visual system, and targeted expression of Toy, like Ey, induces the formation of ectopic eyes. Genetic and biochemical evidence indicates, however, that Toy functions upstream of ey by directly regulating the eye-specific enhancer of ey. Toy is therefore required for initiation of ey expression in the embryo and acts through Ey to activate the eye developmental program.

Regulation and function of the Drosophila segmentation gene fushi tarazu

The Drosophila segmentation gene fushi tarazu (ftz) is expressed in a pattern of seven stripes at the blastoderm stage. Two cis-acting control elements are required for this expression: the zebra element, which confers the striped pattern by mediating the effects of a subset of segmentation genes; and the upstream element, an enhancer element requiring ftz+ activity for its action. Fusion of the upstream element to a basal promoter results in activation of the heterologous promoter in a ftz-dependent striped pattern, supporting the idea that ftz regulates itself by acting through its enhancer. The upstream element can also confer expression patterns similar to that of the homeotic gene Antennapedia, suggesting that a similar element may play a role in the activation of Antennapedia.

An improved in situ hybridization method for the detection of cellular RNAs in Drosophila tissue sections and its application for localizing transcripts of the homeotic Antennapedia gene complex.

An improved method for the detection of cellular RNAs in tissue sections has been developed. It involves in situ hybridization of tritium‐labeled cloned DNA probes to tissue sections and autoradiography. The method was calibrated by using a cloned DNA probe complementary to transcripts abundant in the midgut cells of Drosophila larvae. The improved method also permitted the detection of these transcripts in sectioned embryos where they are much less abundant. The sensitivity of the method can be approximated by quantifying the signal intensities over the hybridizing embryonic midgut cells relative to the larval midgut cells for which the number of transcripts has been estimated. Based on these calculations we estimate that the method is sensitive enough to detect ˜100 complementary RNA molecules per cell after 3 days of autoradiographic exposure with a signal‐to‐noise ratio of 10. The method has been successfully applied to detect transcripts of the homeotic gene Antennapedia. Serial sections allow us to study the spatial pattern of gene expression in the course of development.

Spatial distribution of transcripts from the segmentation gene fushi tarazu during Drosophila embryonic development

The locus fushi tarazu appears to be involved in the establishment of the segmentation pattern of the Drosophila embryo. The cuticle of ftz mutant embryos is missing structures in alternating segments such that only half the normal number of segments are present. We have localized ftz+ transcripts in tissue sections of wild-type Drosophila embryos by in situ hybridization. Transcripts from the ftz+ gene were first detected during nuclear cleavage prior to cell formation. During the last two nuclear divisions ftz+ transcription becomes gradually restricted such that at the cellular blastoderm stage the ftz+ transcripts are localized in seven evenly spaced bands of cells. The size of each band is similar to the size of the segment primordia at the blastoderm. By the time segmentation becomes morphologically distinct ftz+ transcripts are no longer detected. These results suggest that the ftz+ gene plays a key role in the determination of the segmentation pattern in the embryo.

Clonal analysis of primordial disc cells in the early embryo of Drosophila melanogaster

Abstract Single cells were marked by X-ray-induced somatic recombination at the blastoderm stage (3 hr) and at 7 and 10 hr of development, when the embryos are relatively insensitive to radiation damage. The resulting clones of genetically marked cells were analyzed on the adult cuticle. For the three stages examined there is a decrease in clone size with developmental age indicating proliferation of the primordial disc cells which give rise to the adult cuticular structures. The extent of the clones has been used to study the specificity of determination in the primordial disc cells at the time of induction. A significant fraction of the clones induced at all three stages extend from the antenna into the head. In the thorax, some of the clones induced at 3 hr extend from the second leg into the wing, thus indicating that these cells are not yet disc-specifically determined. No clones, however, were found extending from the first leg into the second or from the second into the third, and by 7 hr all thoracic clones were restricted to a single disc. The results provide evidence that determination at the blastoderm stage is not yet disc-specific.

spalt encodes an evolutionarily conserved zinc finger protein of novel structure which provides homeotic gene function in the head and tail region of the Drosophila embryo

The region specific homeotic gene spalt (sal) of Drosophila melanogaster promotes the specification of terminal pattern elements as opposed to segments in the trunk. Our results show that the previously reported sal transcription unit was misidentified. Based on P‐element mediated germ line transformation and DNA sequence analysis of sal mutant alleles, we identified the transcription unit that carries sal function. sal is located close to the misidentified transcription unit, and it is expressed in similar temporal and spatial patterns during embryogenesis. The sal gene encodes a zinc finger protein of novel structure composed of three widely spaced ‘double zinc finger’ motifs of internally conserved sequences and a single zinc finger motif of different sequence. Antibodies produced against the sal protein show that sal is first expressed at the blastoderm stage and later in restricted areas of the embryonic nervous system as well as in the developing trachea. The antibodies detect sal homologous proteins in corresponding spatial and temporal patterns in the embryos of related insect species. Sequence analysis of the sal gene of Drosophila virilis, a species which is phylogenetically separated by approximately 60 million years, suggests that the sal function is conserved during evolution, consistent with its proposed role in head formation during arthropod evolution.

Expression of the caudal gene in the germ line of Drosophila: Formation of an RNA and protein gradient during early embryogenesis

The caudal (cad) gene of Drosophila encodes a maternal and a zygotic transcript which have different promoters. Both mRNAs contain the same open reading frame, including a homeo box. In situ hybridization and antibody staining show that the maternal RNA and protein are localized in an anteroposterior gradient during the syncytial blastoderm stage. The protein is found mainly in the nuclei and is also present in the pole cells. Zygotic RNA and protein are localized in the primordia of the terminal abdominal segment, the hindgut, and in the posterior midgut rudiment. In third instar larvae, cad is expressed in the gut, the gonads, and parts of the genital discs. It is the first homeo box-containing gene expressed in the germ line of Drosophila.

Molecular cloning and chromosome mapping of a mouse DNA sequence homologous to homeotic genes of drosophila

Some of the homeotic genes of Drosophila, involved in the control of segmental development, form a diverged multigene family. A conserved DNA sequence common to these genes has been used to isolate a clone (Mo-10) from the mouse genome which contains a sequence coding for a protein domain that is homologous to the domain conserved in the Drosophila homeotic genes. By structural analogy, this sequence may be involved in the control of metameric pattern formation in the mouse. Mo-10 has been mapped to the proximal portion of mouse chromosome 6, and its position in relationship to genes known to influence mouse morphogenesis is discussed.

The master control gene for morphogenesis and evolution of the eye

The human Aniridia, the murine Small eye, and the eyeless mutations of Drosophila affect homologous (Pax‐6) genes that contain both a paired‐ and a homeobox. By ectopic expression of these genes, functional eyes can be induced on the legs, wings, and antennae of the fly, indicating that eyeless (Pax‐6) is the master control gene for eye morphogenesis. The finding of Pax‐6 from flatworms to humans suggests that eyeless is a universal master control gene and that the various types of eyes in the various animal phyla may have evolved from a single prototype.