Evolution of transcriptional enhancers in the immunoglobulin heavy-chain gene: functional characteristics of the zebrafish Eμ3′ enhancer

Abstract

Past studies of the channel catfish immunoglobulin heavy-chain (IgH) locus indicates that it lacks an Eμ enhancer in the J _H –Cμ1 intron but does have an enhancer, termed Eμ3′, in the μ–δ intergenic region. The positioning of the catfish enhancer downstream of the μ-chain exons is predicted to be unfavorable for antibody-affinity maturation in catfish, and would also have been an impediment to the evolution of class switch recombination, had it existed in early tetrapods. To determine if this downstream enhancer is a general feature of teleost fish, we have identified the location of the transcriptional enhancer in the zebrafish IgH locus. We find that zebrafish, like catfish, only have an Eμ3′-like enhancer that has cross-species activity, but which is B-cell-specific in its activity. A 300-bp region of the zebrafish enhancer shares sequence homology with the core of the catfish Eμ3′, although there has been loss and gain of specific octamer enhancer motifs. Mutagenesis studies demonstrate that the zebrafish IgH enhancer depends on a pair of E-box motifs that are found in the enhancer core. Similarly spaced E-box motifs appear to exist in the Eμ3′ enhancer regions of other teleost fish, suggesting this is a common feature among fish IgH enhancers. We discuss how this distal positioning of the enhancer may influence affinity maturation in extant teleosts as well as the evolution of this process in the early tetrapods.