Expression of the yeast ASP3 gene is strongly regulated by nitrogen catabolite repression (NCR)- NCR is directly related to the change in gene expression and can be controlled by the 5' upstream regulatory region. To determine the upstream regulatory sequences and protein factors which are involved in the regulatory expression of the ASP3 gene, we carried out nucleotide sequencing, Southwestern analysis and gel retardation assay. As a result, we identified a putative upstream activation sequence (UAS) element from the nucleotide sequence data. Then, we detected two protein factors specific for the upstream regulatory regions of the ASP3 gene. In the derepressed condition, when the ASP3 gene is highly expressed, a 70 kDa protein factor binds to the - 1 to - 290 region of the 5' upstream sequences, probably a putative UAS element. In addition, in the repressed condition, when the transcription of the ASP3 gene is suppressed, a 90 kDa protein factor binds to the - 291 to - 465 region. Taken together, we suggested a working model for the transcriptional regulatory mechanism of the yeast ASP3 gene.