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Abstract

Hox genes code for transcriptional regulators that play an important role in the evolution of animal morphology. In the phylogenetic lineage of nematodes, Hox gene number has been reduced, resulting in only four core members in Caenorhabditis elegans. Previous studies in C. elegans and Pristionchus pacificus indicated that Hox genes acquired new functions and that their sequences diverged substantially within the nematodes. For example, when MAB-5, LIN-39 and EGL-5 are compared between C. elegans and P. pacificus, sequence similarities are basically restricted to the homeodomain and the hexapeptide, whereas the rest of the protein differs tremendously. Are these sequence differences crucial for the acquisition of specific Hox functions? To address this question, we used the male rays as an in vivo test system and compare ortologous, paralogous and chimeric Hox genes. Using Cel-mab-5 mutants, we found that, under the endogenous Cel-mab-5 regulatory sequences, cDNA cassettes containing Cel-mab-5 and Ppa- mab-5, but not Cel-lin-39 and Cel-egl- 5 can rescue Cel-mab-5 function. By studying hybrid and deletion constructs, we show that the functional specificity resides within the homeodomain and the hexapeptide. Constructs, containing chimeric Cel-LIN-39 and Cel-MAB-5 proteins, support this observation. We are currently using in vitro mutagenesis experiments to fine map the functional specificity to single amino acids in helix I and II of the homeodomain. Together, our data indicate that the protein function and sequence of Hox transcription factors evolved differentially within nematodes. Furthermore, this and previous data on the evolution of the function of lin-39 indicates different patterns of Hox protein evolution in nematodes and insects.