NOVEL OCTAMER REGULATORY ELEMENT IN THE V(H)11 LEADER EXON OF B-1 CELLS

B-1 cells (CD5 B cells) represent an initial fetal wave of B cell lymp hopoiesis. B-1 cells have fundamental properties that are unique from conventional B cells, including a restricted Ab repertoire. We investi gated the mechanism for the overrepresentation of one such Ig H chain variable-region gene, V(H)11, by murine B-1 cells. We postulated that a cis-regulatory element contributed to the use of V(H)11. We observed that the DNA encoding the leader peptide of V(H)11 was atypically A/T rich and thus was a candidate for nuclear protein binding. By electro phoretic mobility shift analysis, we found that the V(H)11 leader DNA specifically bound to three protein complexes present in the nucleus o f the B-1 cell line AJ9. Of these bands, one was ubiquitous for all ce lls examined (lymphoid and nonlymphoid); another band was present only in B cells, and the third band was specific for B-1 cells that expres sed V(H)11 or V(H)12. In addition to its binding properties, the V(H)1 1 leader sequence also displayed modest tissue-specific enhancer activ ity. By DNA footprint analysis, all three protein complexes were found to bind to an octamer motif embedded within the V(H)11 leader DNA. To identify the octamer-binding proteins, a panel of octamer-specific Ab s was used. We found that the ubiquitous band was Oct-1, and the B cel l-specific band was Oct-2. The B-1 cell-specific nuclear binding prote in was neither Oct-1 nor Oct-2, but may be a novel POU domain protein. We hypothesize that the VH11 leader octamer site may target this gene for preferential rearrangement and/or expression and therefore would be a contributing factor in the increased use of this gene by B-1 cell s.