V-H GENE REPLACEMENT OCCURS IN THE SPLEEN AND BONE-MARROW OF NON-AUTOIMMUNE QUASI-MONOCLONAL MICE

Genes encoding the heavy chain portion of immunoglobulin molecules ari se from the combinatorial association of V, D and J gene segments, whi ch occurs during discrete stages of B lineage development in the bone marrow. Recently, V-H replacement, a form of receptor editing, has bee n described, in which the variable region of an existing VDJ(H) rearra ngement is replaced by another V-H gene segment in a recombination eve nt believed to involve an embedded heptamer within the coding region o f the V-H. Studies of transgenic mice with ''knocked-in'' VDJ(H) genes encoding anti-DNA specificity have demonstrated that receptor editing of the heavy chain is one mechanism by which autoreactive B cell rece ptors can be modified. Another mouse, the ''quasi-monoclonal'', which encodes a ''knocked-in'' VDJH for the hapten NP also contains B lineag e cells that undergo V-H replacement. This suggests that V-H replaceme nt may play a role in the normal diversification of the antibody reper toire. Using a ligation-mediated PCR assay, we have identified V-QM do uble-stranded DNA breaks indicative of V-H replacement intermediates f rom bone marrow and splenic B lineage cells of quasi-monoclonal mice i n the absence of immunization. V-QM to J558 recombination deletion pro ducts consistent with V-H replacement were also detected in both the b one marrow and spleen of non-immunized quasi-monoclonal mice. Moreover , RAG-1 transcripts were detected in the spleen. These data suggest th at V-H replacement can be part of the mechanism(s) used by B lineage c ells to generate diversity throughout B lineage development, including later stages occurring in secondary lymphoid tissues.