The p38/stress-activated protein kinase2 (p38/SAPK2) is activated by cellular stress and proinflammatory cytokines. Several transcription factors have been reported to be regulated by p38/SAPK2, and this kinase is involved in the control of expression of various genes. In human Jurkat T-cells, induction of the early growth response gene-1 (egr-1) by anisomycin is completely inhibited by SB203580, a specific inhibitor of p38/SAPK2a and -b. Northern blot and reporter gene experiments indicate that this block is at the level of mRNA biosynthesis. Using mutants of the egr-1 promoter, we demonstrate that a distal cAMP-responsive element (CRE; nucleotides -134 to -126) is necessary to control egr-1 induction by p38/SAPK2. Pull-down assays indicate that phospho-CRE binding protein (CREB) and phospho-activating transcription factor-1 (ATF1) bind to this element in a p38/SAPK2-dependent manner. In response to anisomycin, two known CREB kinases downstream to p38/SAPK2, MAPKAP kinase 2 (MK2) and mitogen- and stress-activated kinase 1 (MSK1), show increased activity. However, in MK2 -/- fibroblasts derived from mice carrying a disruption of the MK2 gene, the phosphorylation of CREB and ATF1 and the expression of egr-1 reach levels comparable with wild type cells. This finding excludes MK2 as an involved enzyme. We conclude that egr-1 induction by anisomycin is mediated by p38/SAPK2 and probably by MSK1. Phosphorylated CREB and ATF1 then bind to the CRE of the egr-1 promoter and cause a stress-dependent transcriptional activation of this gene.