Interleukin-3 (IL-3) is a hematopoietic growth factor that regulates the differentiation of multilineage and committed progenitor cells and the functions of some mature blood cells. The expression of human IL-3 appears to be restricted to stimulated T lymphocytes. We have investigated the kinetics and mechanisms involved in the induction of IL-3 expression in the human T lymphocytic tumor cell line Jurkat. We show that accumulation of IL-3 mRNA is controlled at both the transcriptional and posttranscriptional level. Transcription of the IL-3 gene in these cells appears to be constitutive but no IL-3 mRNA was detected in unstimulated cells, indicating that in resting cells IL-3 mRNA is highly unstable. Treatment with phytohemagglutinin (PHA) induced a small and transient increase in the IL-3 gene transcription rate and led to the production of detectable levels of IL-3 mRNA and protein. Optimal induction of IL-3 expression required a second stimulus. Costimulation of Jurkat cells with both phorbol myristate acetate and PHA caused both a transient increase in IL-3 gene transcription, which is dependent on new protein synthesis, and also a transient increase in mRNA stability.