Antigen presentation to the T cell receptor leads to sustained cytosolic Ca(2+) elevation, which is critical for T-cell activation. We previously showed that in activated T cells, Ca(2+) clearance is inhibited by the endoplasmic reticulum Ca(2+) sensor stromal interacting molecule 1 (STIM1) via association with the plasma membrane Ca(2+)/ATPase 4 (PMCA4) Ca(2+) pump. Having further observed that expression of both proteins is increased in activated T cells, the current study focused on mechanisms regulating both up-regulation of STIM1 and PMCA4 and assessing how this up-regulation contributes to control of Ca(2+) clearance. Using a STIM1 promoter luciferase vector, we found that the zinc finger transcription factors early growth response (EGR) 1 and EGR4, but not EGR2 or EGR3, drive luciferase activity. We further found that neither STIM1 nor PMCA4 is up-regulated when both EGR1 and EGR4 are knocked down using RNA interference. Further, under these conditions, activation-induced Ca(2+) clearance inhibition was eliminated with little effect on Ca(2+) entry. Finally, we found that nuclear factor of activated T cell (NFAT) activity is profoundly attenuated if Ca(2+) clearance is not inhibited by STIM1. These findings reveal a critical role for STIM1-mediated control of Ca(2+) clearance in NFAT induction during T-cell activation.-Samakai, E., Hooper, R., Martin, K. A., Shmurak, M., Zhang, Y., Kappes, D. J., Tempera, I., Soboloff, J. Novel STIM1-dependent control of Ca(2+) clearance regulates NFAT activity during T-cell activation.