CD8 is a cell surface glycoprotein on major histocompatibility complex class I-restricted T cells. Thymocytes and most peripheral T cells express CD8 as heterodimers of CD8 alpha and CD8 beta. The intestinal intraepithelial lymphocytes (IEL), which have been suggested to be generated extrathymically, express CD8 predominantly as homodimers of CD8 alpha. We have generated CD8 beta gene-targeted mice. CD8 alpha+ T cell population in the thymus and in most peripheral lymphoid organs was reduced to 20-30% of that in wild-type littermates. CD8 alpha expression on thymocytes and peripheral T cells also decreased to 44 and 53% of the normal levels, respectively. In contrast, neither the population size nor the CD8 alpha expression level of CD8 alpha+ IEL was reduced. This finding indicates that CD8 beta is important only for thymic-derived CD8+ T cells. The lack of CD8 beta reduces but does not completely abolish thymic maturation of CD8+ T cells. Our result also reveals the role of CD8 beta in regulating CD8 alpha expression on thymic derived T cells. Peripheral T cells in these mice were efficient in cytotoxic activity against lymphocytic choriomeningitis virus and vesicular stomatitis virus, suggesting that CD8 beta is not essential for the effector function of CD8+ T cells.