Alone among piscine taxa, the antarctic icefishes (family Channichthyidae, suborder Notothenioidei) have evolved compensatory adaptations that maintain normal metabolic functions in the absence of erythrocytes and the respiratory oxygen transporter hemoglobin. Although the uniquely "colorless" or "white" condition of the blood of icefishes has been recognized since the early 20th century, the status of globin genes in the icefish genomes has, surprisingly, remained unexplored. Using alpha- and beta-globin cDNAs from the antarctic rockcod Notothenia coriiceps (family Nototheniidae, suborder Notothenioidei), we have probed the genomes of three white-blooded icefishes and four red-blooded notothenioid relatives (three antarctic, one temperate) for globin-related DNA sequences. We detect specific, high-stringency hybridization of the alpha-globin probe to genomic DNAs of both white- and red-blooded species, whereas the beta-globin cDNA hybridizes only to the genomes of the red-blooded fishes. Our results suggest that icefishes retain inactive genomic remnants of alpha-globin genes but have lost, either through deletion or through rapid mutation, the gene that encodes beta-globin. We propose that the hemoglobinless phenotype of extant icefishes is the result of deletion of the single adult beta-globin locus prior to the diversification of the clade.

The icefishes of the Southern Ocean (family Channichthyidae, suborder Notothenioidei) are unique among vertebrates in their inability to synthesize hemoglobin. We have shown previously (Cocca, E., Ratnayake-Lecamwasam, M., Parker, S. K., Camardella, L., Ciaramella, M., di Prisco, G., and Detrich, H. W., III (1995)Proc. Natl. Acad. Sci. U. S. A. 92, 1817–1821) that icefishes retain inactive genomic remnants of adult notothenioid α-globin genes but have lost the gene that encodes adult β-globin. Here we demonstrate that loss of expression of the major adult α-globin, α1, in two species of icefish (Chaenocephalus aceratus and Chionodraco rastrospinosus) results from truncation of the 5′ end of the notothenioid α1-globin gene. The wild-type, functional α1-globin gene of the Antarctic yellowbelly rockcod, Notothenia coriiceps, contains three exons and two A + T-rich introns, and its expression may be controlled by two or three distinct promoters. Retained in both icefish genomes are a portion of intron 2, exon 3, and the 3′-untranslated region of the notothenioid α1-globin gene. The residual, nonfunctional α-globin gene, no longer under positive selection pressure for expression, has apparently undergone random mutational drift at an estimated rate of 0.12–0.33%/million years. We propose that abrogation of hemoglobin synthesis in icefishes most likely resulted from a single mutational event in the ancestral channichthyid that deleted the entire β-globin gene and the 5′ end of the linked α1-globin gene.