EVOLUTION OF PAIRED DOMAINS - ISOLATION AND SEQUENCING OF JELLYFISH AND HYDRA PAX GENES RELATED TO PAX-5 AND PAX-6

Pax proteins are a family of transcription factors with a highly conse rved paired domain; many members also contain a paired-type homeodomai n and/or an octapeptide, Nine mammalian Pax genes are known and classi fied into four subgroups: Pax-1/9, Pax-2/5/8, Pax-3/7, and Pax-4/6. Mo st of these genes are involved in nervous system development, In parti cular, Pax-6 is a key regulator that controls eye development in verte brates and Drosophila. Although the Pax-4/6 subgroup seems to be more closely related to Pax-2/5/8 than to Pax-3/7 or Pax-1/9, its evolution ary origin is unknown. We therefore searched for a Pax-6 homolog and r elated genes in Cnidaria, which is the lowest phylum of animals that p ossess a nervous system and eyes, A sea nettle (a jellyfish) genomic l ibrary was constructed and two pax genes (Pax-A and -B) were isolated and partially sequenced. Surprisingly, unlike most known Par genes, th e paired box in these two genes contains no intron, In addition, the c omplete cDNA sequences of hydra Pax-A and -B were obtained, Hydra Pax- B contains both the homeodomain and the octapeptide, whereas hydra Pax -A contains neither. DNA binding assays showed that sea nettle Pax-A a nd -B and hydra Pax-A paired domains bound to a Pax-5/6 site and a Pax -5 site, although hydra Pax-B paired domain bound neither, An alignmen t of all available paired domain sequences revealed two highly conserv ed regions, which cover the DNA binding contact positions. Phylogeneti c analysis showed that Pax-A and especially Pax-B were more closely re lated to Pax-2/5/8 and Pax-4/6 than to Pax-1/9 or Pax-3/7 and that the Pax genes can be classified into two supergroups: Pax-A/Pax-B/ Pax-2/ 5/8/4/6 and Pax-19/3/7, From this analysis and the gene structure, we propose that modern Pax-4/6 and Pax-2/ 5/8 genes evolved from an ances tral gene similar to cnidarian Pax-B, having both the homeodomain and the octapeptide.