CHARACTERIZATION OF OCT2 FROM THE CHANNEL CATFISH - FUNCTIONAL PREFERENCE FOR A VARIANT OCTAMER MOTIF

The Ig heavy chain enhancer of the channel catfish (Ictalurus punctatu s) has an unusual position and structure, being found in the 3' region of the mu gene and containing eight functional octamer motifs of cons ensus (ATGCAAAT) and variant sequences. The presence of multiple octam er motifs suggests that an Oct2 homologue may play an important role i n driving expression of the Ig heavy chain locus in a teleost fish, To test this hypothesis, two catfish Oct2 cDNAs (alpha and beta) were cl oned by screening a catfish B fell cDNA library, Catfish Oct2 alpha an d beta isoforms are derived by alternative RNA splicing; as determined by Southern analysis, Oct2 is a single copy gene, In comparisons with mammalian Oct2, the catfish Oct2 isoforms show high sequence conserva tion in their N-terminal regions and POU domains, but extensive diverg ence in their C-terminal regions, Catfish Oct2 alpha and beta are tiss ue restricted, bind both consensus and variant octamer motifs, and act ivate transcription in both catfish and murine cells, In contrast, mou se Oct2 activated transcription in mouse but not catfish cells, Catfis h Oct2 beta is a more potent transcriptional activator than Oct2 alpha . In transient expression assays, catfish Oct2 beta showed a marked pr eference for the octamer variant, ATGtAAAT, which occurs twice in the catfish enhancer, Mouse Oct2 also showed increased activity with the v ariant octamer when tested in mouse B cells, Gel-shift analysis compet ition assays indicated that catfish Oct2 binds the consensus octamer m otif with an apparently higher affinity than it does the variant motif .