ACTIVATION OF THE IG GERM-LINE GAMMA-1 PROMOTER - INVOLVEMENT OF C ENHANCER-BINDING PROTEIN TRANSCRIPTION FACTORS AND THEIR POSSIBLE INTERACTION WITH AN NF-IL-4 SITE/

Ig isotype switching in B lymphocytes is preceded by transcription of the corresponding unrearranged, or germ-line (GL), C-H gene. The promo ter of mouse GL C gamma 1 transcripts has been shown to be located wit hin a 349-bp KpnI/BglII fragment, extending from -147 to +202 bp relat ive to the first transcription initiation site. By the electrophoretic mobility shift assay, we have analyzed nuclear extracts from three B cell lines and splenic B cells for the presence of proteins binding to this fragment. We show that they give different patterns of DNA bindi ng, implying significant complexity in the regulation of this locus. W e focused on the slgM(+) mouse B lymphoma line L10A6.2 that has been s hown able to confer responsiveness of the GL gamma 1 promoter to phorb ol ester plus IL-4. Activation of this cell line results in altered ex pression of several nuclear DNA-binding complexes involving two member s of the C/enhancer-binding protein (EBP) family of transcription fact ors, namely C/EBP beta (nuclear factor (NF)-IL-6/LAP) and Ig/EBP-1 (C/ EBP gamma). The complexes bind to two C/EBP elements, one at about -11 5 bp and one near the first RNA start site. In normal B cells stimulat ed by LPS or IL-4, new complexes appear that bind to C/EBP and NF-IL-4 elements, respectively, located within the -125/-101 region. The -125 /-101 segment previously has been shown to be required for transcripti onal activation. We discuss these findings in relation to the presence of consensus C/EBP binding sites in other IL-4-regulated promoters.