Ribosome biogenesis has been associated with regulation of cell growth and cell division, but the molecular mechanisms that integrate the effect of ribosome biogenesis on these processes in mammalian cells remain unknown. To study the effect of impaired ribosome functions in vivo, we conditionally deleted one or two alleles of the 40S ribosomal protein S6 gene in T cells in the mouse. While complete deletion of S6 abrogated T-cell development, hemizygous expression did not have any effect on T-cell maturation in the thymus, but inhibited the accumulation of T cells in the spleen and lymph nodes, as a result of their decreased survival in the peripheral lymphoid organs. Additionally, TCR-mediated stimulation of S6-heterozygous T cells induced a normal increase in their size, but cell cycle progression was impaired. Genetic inactivation of p53 tumor suppressor rescued development of S6-homozygous null thymocytes and proliferative defect of S6-heterozygous T cells. These results demonstrate the existence of a p53-dependent checkpoint mechanism that senses changes in the fidelity of the translational machinery to prevent aberrant cell division or eliminate defective T cells in vivo. Failure to activate this checkpoint response could potentially lead to a development of pathological processes such as tumors and autoimmune diseases.[Keywords: S6 ribosomal protein; ribosome biogenesis; cell growth; cell proliferation; checkpoint] Supplemental material is available at http://www.genesdev.org.