INACTIVATION OF BTK BY INSERTION OF LACZ REVEALS DEFECTS IN B-CELL DEVELOPMENT ONLY PAST THE PRE-B-CELL STAGE

Bruton's tyrosine kinase (Btk) is a cytoplasmic protein kinase that is defective in X-linked agammaglobulinaemia in man and in X-linked immu nodeficiency in the mouse. There is controversy regarding the stages o f B cell development that are dependent on Btk function, To determine the point in B cell differentiation at which defects in Btk become app arent, we generated a mouse model by inactivating the Btk gene through an in-frame insertion of a lacZ reporter by homologous recombination in embryonic stem cells. The phenomenon of X-chromosome inactivation i n Btk(+/-) heterozygous female mice enabled us to evaluate the competi tion between B cell progenitors expressing wildtype Btk and those expr essing the Bfk(-)/lacZ allele in each successive step of development, Although Bfk was already expressed in pro-B cells, the first selective disadvantage only became apparent at the transition from small pre-B cells to immature B cells in the bone marrow, A second differentiation arrest was found during the maturation from IgD(low)IgM(high) to IgD( high)IgM(low) stages in the periphery, Our results show that Bfk expre ssion is essential at two distinct differentiation steps, both past th e pre-B cell stage.