HAX1 was originally described as HS1‐associated protein with a suggested function in receptor‐mediated apoptotic and proliferative responses of lymphoid cells. Recent publications refer to a complex and multifunctional role of this protein. To investigate the in vivo function of HAX1 (HS1‐associated protein X1) in B cells, we generated a Hax1‐deficient mouse strain. Targeted deletion of Hax1 resulted in premature death around the age of 12 wk accompanied by a severe reduction of lymphocytes in spleen, thymus and bone marrow. In the bone marrow, all B‐cell populations were lost comparably. In the spleen, B220+ cells were reduced by almost 70%. However, as investigated by adoptive transfer experiments, this impairment is not exclusively B‐cell intrinsic and we hypothesize that a HAX1‐deficient environment cannot sufficiently provide the essential factors for proper lymphocyte development, trafficking and survival. Hax1−/− B cells show a significantly reduced expression of CXCR4, which might have an influence on the observed defects in B‐cell development.