In testicular Sertoli cells, IL-1β regulates steroid, lactate, and transferrin secretion; although each influences germ cell development and spermatogenesis, little is known about the signaling mechanisms involved. In other cell types, IL-1β potently induces reactive oxygen species and/or cyclooxygenase-2 (COX-2). In contrast, in Sertoli cells, IL-1β does not generate reactive oxygen species, but rapidly phosphorylates c-Jun-NH_2-terminal kinase (JNK), but not p44/42 or p38 MAPK. Phosphorylated JNK stimulates COX-2 activity, mediating the expression of ILs and steroidogenic acute regulatory (StAR)-related (StAR-related lipid transfer protein domain containing) proteins D1 and D5, but not D4. In a time- and dose-dependent manner, IL-1β rapidly increases levels of COX-2 mRNA (2-fold); induction of COX-2 protein (50-fold) requires de novo protein synthesis. Concomitantly, increases in IL-1a, IL-6, and IL-1β mRNAs (1-3 h) are observed. As StAR-related lipid transfer protein domain containing protein 1 (StARD1) mRNA decreases, StARD5 mRNA increases; substantial recovery phase induction of StARD1 mRNA above control is noted (24 h). Inhibition of JNK or COX-2 activities prevents IL-1β induction of IL and StARD5 mRNAs and subsequent increases in StARD1 mRNA (24 h), indicating that these effects depend on the activation of both enzymes. StARD1 and D5 protein levels are significantly altered, consistent with posttranscriptional and posttranslational regulation. IL-1β rapidly decreases levels of precursor and mature sterol regulatory element-binding protein-1, changes not altered by cycloheximide, suggesting coordinate regulation of StARD1 and -D5, but not StARD4, expression. These data demonstrate that JNK and COX-2 activities regulate Sertoli cytokines and particularly START domain-containing proteins, suggesting protective stress responses, including transcription and protein and lipid regulation, within this specialized epithelium.