Functional motifs in the IgH enhancer of the channel catfish

A

bstract The transcriptional enhancer (Eμ3′) within the Ig heavy chain (IgH) locus of the channel catfish differs from those found in mammalian IgH loci in both its location and structure. However, upon transfection into fish or mouse lymphocytes, it activates transcription to an extent equivalent to that of the mouse IgH intronic enhancer (Eμ). Potential transcription factor binding motifs in Eμ3′ are more numerous than in mammalian IgH enhancers, and are dispersed over 1.6 kilobases. We transfected catfish and mouse lymphoid cells with reporters under the control of artificial promoters containing motifs from the catfish enhancer. We demonstrate that 9 of 11 octamer motifs identified in the catfish enhancer, representing five variations of the consensus octamer (ATGCAAAT), are functional in both a catfish B-cell line (1B10) and the mouse plasmacytoma J558L. Only those octamer variants in which one of the first four bases is altered are active. Clear species differences in the strengths of the variant octamer motifs were evident, and in catfish B cells the ATGtAAAT motif was over threefold more active than the consensus octamer. The one μA and two μB motifs in Eμ3′ do not contribute to transcriptional activation. These results suggest that the relative functional contributions of IgH enhancer motifs has changed significantly during vertebrate evolution.