Conserved non-coding sequence (CNS)-1 has been shown to coordinately regulate the expression of the Th2 cytokines genes IL4, IL13 and IL5. We have used the interaction between CNS-1 and the human IL13 and IL4 promoters as a model to pursue the molecular mechanisms underlying CNS-1-dependent regulation of Th2 cytokine gene transcription. CNS-1 potently enhanced the activity of IL13 and IL4 promoter reporter vectors upon full T cell activation. Analysis of CNS-1 deletion mutants mapped enhancer activity to a short core (CNS-1_270-337) that contains three closely spaced cyclic AMP responsive elements (CRE). CRE site 2 bound CRE binding protein (CREB) and activating transcription factor (ATF)-2 in vitro, and was essential for CNS-1-dependent upregulation of IL13 transcription. Cotransfection of an IL13 reporter construct with expression vectors for wild type or mutant CREB and ATF-2 showed CREB, but not ATF-2, regulates CNS-1 enhancer activity. Notably, chromatin immunoprecipitation analysis showed T cell activation recruits CREB and the coactivator CREB binding protein (CBP)/p300 to the endogenous CNS-1. Moreover, CBP/p300 activity was essential for CNS-1 mediated enhancement of IL13 transcription. Collectively, these data define the region within CNS-1 responsible for enhancement of IL13 and IL4 transcription, and suggest CREB/CBP-dependent mechanisms play an important role in facilitating Th2 cytokine gene expression in response to T cell receptor signaling.