Biased V(H) gene usage in early lineage human B cells: Evidence for preferential Ig gene rearrangement in the absence of selection

Certain V(H) genes are predominantly expressed in mature B cells. We hypothesized that several, mutually nonexclusive V(H)-dependent mechanisms operating at distinct stages during B cell development may be responsible for overrepresentation of these V(H) genes. In the present study, we have assessed whether one of the mechanisms involves preferential rearrangement at the pro-B cell stage. The frequency of individual V(H)4 and V(H)3 genes in rearrangement libraries from FACS-purified human CD34⁺/CD19⁺ pro-B and CD34^-/CD19⁺ pre-B cells was assessed. The in-frame and out-of-frame rearrangements from both cell populations were analyzed using a high resolution PAGE system. The frequencies of individual V(H) gene segments among out-of-frame rearrangements from pro-B cells were determined, because these frequencies should reflect only processes before the translation of the μ-heavy chain and should not be biased by selection mechanisms. Our results demonstrate that, at the pro-B cell stage, the V4-34, V4-39, and V4-59 gene segments are the most frequently rearranged V(H)4 family genes, and the V3-23 and V3-30 gene segments are the most frequently rearranged V(H)3 family genes. This finding suggests that the predominant expression of these V(H) genes in peripheral mature B cells is determined to a significant degree by their preferential rearrangement during V-DJ recombination.