Origin of the metazoan bodyplan: characterization and functional testing of the promoter of the homeobox gene EmH-3 from the freshwater sponge Ephydatia muelleri in mouse 3T3 cells.

Abstract

Porifera [sponges] represent the lowest metazoan phylum, probably already existing prior to the 'Cambrian explosion'. Based on amino acid sequences deduced from cDNAs that code for structural proteins, the monophyly of Metazoa was established. Now we analyzed for the first time a promoter of a sponge gene for its activity in a heterologous cell system from higher Metazoa. The promoter of the homeobox gene EmH-3 was cloned and sequenced from a genomic library of the freshwater sponge Ephydatia muelleri. For the determination of functional promoter activity, transient transfection experiments in mouse NIH 3T3 cells were performed; the promoter was fused with the luciferase reporter gene. The data revealed that a 401 nt long promoter fragment, comprising several binding elements for metazoan transcription factors, showed the highest activity, while the 175 bp long promoter segment, comprising solely the TATA- and Cap boxes, showed only 25% of that activity. This result demonstrates that the sponge promoter is activated by factors present in mammalian cells and supports the view that Porifera, together with the other metazoan phyla, are of monophyletic origin.