Ascidian and amphioxus adh genes correlate functional and molecular features of the ADH family expansion during vertebrate evolution

The alcohol dehydrogenase (ADH) family has evolved into at least eight ADH classes during vertebrate evolution. We have characterized three prevertebr ate forms of the parent enzyme of this family, including one from an urocho rdate (Ciona intestinalis) and two from cephalochordates (Branchiostoma flo ridae and Branchiostoma lanceolatum). An evolutionary analysis of the famil y was performed gathering data from protein and gene structures, exon-intro n distribution, and functional features through chordate lines. Our data st rongly support that the ADH family expansion occurred 500 million years ago , after the cephalochordate/vertebrate split, probably in the gnathostome s ubphylum line of the vertebrates. Evolutionary rates differ between the anc estral, ADH3 (glutathione-dependent formaldehyde dehydrogenase), and the em erging forms, including the classical alcohol dehydrogenase, ADH1, which ha s an evolutionary rate 3.6-fold that of the ADH3 form. Phylogenetic analysi s and chromosomal mapping of the vertebrate Adh gene cluster suggest that f amily expansion took place by tandem duplications, probably concurrent with the extensive isoform burst observed before the fish/tetrapode split, rath er than through the large-scale genome duplications also Postulated in earl y vertebrate evolution. The absence of multifunctionality in lower chordate ADHs and the structures compared argue in favor of the acquisition of new functions in vertebrate ADH classes. Finally, comparison between B. florida e and B. lanceolatum Adhs provides the first estimate for a cephalochordate speciation, 190 million years ago, probably concomitant with the beginning of the drifting of major land masses from the Pangea.