T cell development within the thymus involves the ordered expression of a number of tissue-specific components such as the CD2 gene. Control of expression of this gene is regulated by a well characterized 3' enhancer together with a promoter and upstream elements. The CD2 promoter is typical of a group of T cell-specific promoters that lack a TATA box and use multiple sites for initiation of transcription. An "E box" motif CACGTG, located just upstream from the most 5' initiation start site, was found to contribute a major effect to the level of basal transcription of a reporter gene. Analysis of the proteins in T cell extracts that bound to this site revealed that the bHLH-LZ protein USF was the major component. A functional role for USF was established in transient transfection experiments. Thus, this protein restored full promoter activity following repression caused by cotransfection with the E box binding bHLH-LZ protein Max. Taken together, these results indicate that an E box motif is critical to expression of the CD2 gene during T cell development and that the HLH protein USF acts as a transcriptional activator of the CD2 promoter.