REGULATION OF B-CELL DEVELOPMENT BY VARIABLE GENE COMPLEXITY IN MICE RECONSTITUTED WITH HUMAN-IMMUNOGLOBULIN YEAST ARTIFICIAL CHROMOSOMES

The relationship between variable (V) gene complexity and the efficien cy of B cell development was studied in strains of mice deficient in m ouse antibody production and engineered with yeast artificial chromoso mes (YACs) containing different sized fragments of the human heavy (H) chain and kappa light (L) chain loci. Each of the two H and the two k appa chain fragments encompasses, in germline configuration, the same core variable and constant regions but contains different numbers of u nique V-H (5 versus 66) or V kappa genes (3 versus 32). Although each of these YACs was able to substitute for its respective inactivated mu rine counterpart to induce B cell development and to support productio n of human immunoglobulins (Igs), major differences in the efficiency of B cell development were detected. Whereas the YACs with great V gen e complexity restored efficient development throughout all the differe nt recombination and expression stages, the YACs with limited V gene r epertoire exhibited inefficient differentiation with significant block s at critical stages of B cell development in the bone marrow and peri pheral lymphoid tissues. Our analysis identified four key checkpoints regulated by V-H and V kappa gene complexity: (a) production of functi onal mu chains at the transition from the pre B-I to the pre B-II stag e; (b) productive V kappa J kappa recombination at the small pre B-II stage; (c) formation of surface Ig molecules through pairing of mu cha ins with L chains; and (ri) maturation of B cells. These findings demo nstrate that V gene complexity is essential not only for production of a diverse repertoire of antigen-specific antibodies but also for effi cient development of the B cell lineage.