The "eyes absent" (eya) gene in the eye-bearing hydrozoan jellyfish Cladonema radiatum: conservation of the retinal determination network.

Eyes absent (Eya) is a member of the Retinal Determination Gene Network (RDGN), a set of genes responsible for eye specification in Drosophila. Eya is a dual function protein, working as a transcription factor in the nucleus and as a tyrosine phosphatase in the cytoplasm. It had been shown that Pax and Six family genes, main components of the RDGN, are present in the hydrozoan Cladonema radiatum and that they are expressed in the eye. However, nothing had been known about the Eya family in hydrozoan jellyfish. Here we report the presence of an Eya homologue (CrEya) in Cladonema. Real-time PCR analysis and in situ hybridization showed that CrEya is expressed in the eye. Furthermore, the comprehensive survey of eukaryote genomes revealed that the acquisition of the N-terminal transactivation domain, including the EYA Domain 2 and its adjacent sequence shared by all eumetazoans, happened early in evolution, before the separation of Cnidaria and Bilateria. Our results uncover the evolution of the two domains and show a conservation of the expression pattern of the Eya gene between Cnidaria and Bilateria, which, together with previous data, supports the hypothesis of the monophyletic origin of metazoans eyes. We additionally show that CrEya is also expressed in the oocytes, where two other members of the RDGN, CrPaxB, and Six4/5-Cr, are known to be expressed. These data suggest that several members of the RDGN have begun to be localized also into the different context of egg development early in the course of metazoan evolution.

Spatial and temporal expression of an Antennapedia/lac Z gene construct integrated into the endogenous Antennapedia gene of Drosophila melanogaster.

In order to study the regulation of spatial and temporal expression of the homeotic gene Antennapedia (Antp) in Drosophila melanogaster, we have constructed fusion genes which contain Antp sequences linked to the reporter gene lac Z of Escherichia coli. In one case of P-element transformation, a fusion gene construct integrated into the endogenous Antp gene close to one of the two promoters (P1). The spatial expression from the reporter gene in this transformant line, as analysed by the detection of ß-galactosidase activity, was found to exactly mimic the normal expression from the P1 promoter of the Antp gene. We have used this unique transformant as a tool for studying the expression of the P1 promoter in embryonic, larval and adult development. Parallel lines transformed with the same fusion gene construct did not confer a correct P1 pattern of expression. The position in the genome was, therefore, crucial for the expression pattern of the reporter gene. Experiments aiming at the detection of autoregulatory control of Antp gene expression were designed. The results did not, however, support models of positive or negative autoregulation of P1 expression by Amp protein.

In situ localization of the transcripts of a homeobox gene in the honeybee Apis mellifera L. (Hymenoptera).

We have isolated and characterized a homeoboxcontaining gene from the honeybee Apis mellifera. Its homeobox region shows a high degree of sequence similarity to the homeobox of the Drosophila gene Deformed (Dfd). At the DNA level 82% of the basepairs are the same, whereas the putative amino acid sequences are identical between the bee and the fruitfly genes. Similarity is also present 5' and 3' to the homeobox. Using this isolate as a probe we have performed in situ hybridization on sections from blastoderm-stage embryos of the honeybee Apis mellifera. In early blastoderm stages we found a rather irregular pattern of labelled nuclei. In middle stages we found silver grains over each nucleus and also over the cytoplasm in a belt of blastoderm cells in the prospective gnathal region. These results indicate that the Deformed genes from honeybee and fruitfly are homologous both with respect to their DNA sequence and their spatial and temporal pattern of expression during embryogenesis.