Long-Term Inositol Phosphate Release, but Not Tyrosine Kinase Activity, Correlates with IL-2 Secretion and NF-AT Induction in Anti-CD3-Activated Peripheral Human T Lymphocytes

The cascade of events within the first few minutes of T cell stimulation has been well characterized. Although many second messengers have been shown to be necessary and sufficient for T cell activation in a number of model systems, the rate-limiting step in peripheral T cells has not been demonstrated. To model effective versus ineffective CD3-mediated stimulation in peripheral T cells, we used two anti-CD3 mAbs that differ in their ability to stimulate purified T cells: OKT3, which causes early second messenger generation but is unable to activate T cells without a second signal, and 64.1, which stimulates T cell proliferation on its own. We found that tyrosine kinase activity was similar for both mAbs over a period of hours. However, the inositol phosphate response was stronger for 64.1 than for OKT3. To tie these events to gene activation, we measured NF-κB and NF-AT activity in the nucleus after anti-CD3 stimulation. Both stimuli induced the appearance of the NF-κB components (c-Rel, p65 (RelA), and p50 (NF-κB1)) and NF-κB DNA binding activity in the nucleus. However, only 64.1 induced NF-AT in the nucleus, correlating with its ability to activate T cells. Thus, NF-AT induction and IL-2 secretion were correlated with the levels of inositol phosphate release but not with gross levels of tyrosine kinase activity induced late following the response. On the other hand, NF-κB induction and IL-2 receptor expression occurred even with the smaller second messenger response generated by OKT3.