The Deb-A gene from Drosophila melanogaster encodes a small membrane-associated protein, regulated during development, with peak abundance at 12-15 h of embryogenesis. The cis-acting regulatory elements that control expression of Deb-A during embryogenesis were localized using a somatic transformation assay. The Adh gene of D. melanogaster was used as a 'reporter' gene. The promoterless ADH coding sequence was fused to the 5'-upstream control region of Deb-A. Deletions were introduced into the 5'-region using various restriction sites and Bal31 deletion mutagenesis. A negative regulatory element, or silencer, was localized to a segment 47 base pairs long, between -395 and -442. It is responsible for 80% of the repression of gene expression during late development and reduces levels of Deb-A RNA nearly 5-fold. This negative element is temporally functional. It becomes active after 15 h of embryogenesis and it has no effect on gene expression prior to that. Within this negative element of 47 base pairs, two footprint regions were protected from DNase I digestion by embryonic nuclear extracts: one region contains an AP-1 binding site, but the other footprint is due to unknown element(s). High molecular weight DNA-protein complexes on an oligonucleotide probe spanning the AP-1 binding site were identified in gel retardation assays using partially purified bacterially expressed Djun protein or nuclear extracts from Drosophila embryos. These data suggest that the AP-1 site may be partly responsible for decreasing Deb-A expression during the late embryonic developmental stages of D. melanogaster.