Adaptive evolution of gene expression in Antarctic fishes: Divergent transcription of the 5 '-to-5 ' linked adult alpha 1- and beta-globin genes of the Antarctic teleost Notothenia coriiceps is controlled by dual promoters and intergenic enhancers

Unlike temperate fishes, Antarctic fishes of the notothenioid suborder, who se body temperatures (-2 to +1 degreesC) conform to the Southern Ocean, mus t express their genomes In an extremely cold thermal regime. To determine w hether these fishes have evolved compensatory adjustments that maintain eff icient gene transcription at low temperatures, we have initiated studies of the cis-acting regulatory elements that control globin gene expression in the Antarctic rockcod Notothenia coriieeps and in its close relative, the t emperate New Zealand black cod N. angustata (habitat temperature = +6 to +1 5 degreesC). The genes encoding the major alpha1 and beta globins of these fishes are tightly linked in head-to-head (5' to 5') orientation. The inter genic regions separating the globin genes in the two fishes, similar to4.3 kb in N. coriiceps and similar to3.2 kb in N. angustata, are highly similar in sequence, the major difference being the absence of a 1.1-kb, repeat-co ntaining segment in the latter. To assess the promoter and enhancer activit ies of the intergenic regions, each was cloned into the luciferase-reporter vector pGL3-Basic, and the constructs were transfected into MEL cells. Upo n DMSO induction of MEL cell differentiation, each of the alpha/beta -inter genic regions functioned in both orientations as erythroid-responsive trans criptional regulators. However, expression of luciferase mediated by the N. coriieeps intergene was 6-fold greater in the a orientation than that for the N. angustata intergene and 2-fold greater for the beta. The greater tra nscription-stimulating activity of the N. coriiceps intergene can be attrib uted to two enhancers composed of combinations of CAC/Sp1 and GATA motifs a nd located in direct repeat elements. N. angustata, which lacked repetitive structure in its intergene, contained a single copy of the enhancer. We pr opose that cold adaptation of globin gene expression in N. coriiceps evolve d in part through duplication and refinement of critical cis-acting regulat ory elements as the Southern Ocean cooled during the past 25 million years.