The phenotypes of 24 mutants that successively delete DNA sequences adjacent to the 5' end of the Saccharomyces cerevisiae (yeast) his3 structural gene are described. Deletions retaining greater than 155 base pairs before the mRNA coding sequences are phenotypically indistinguishable from the wild-type his3 allele. Deletions having end points between 113 and 65 base pairs before the transcription initiation site express his3 at reduced levels. Mutations retaining less than 45 base pairs are indistinguishable from null alleles of the his3 locus. These results indicate (i) that a sequence(s) located 113--155 base pairs upstream from the transcribed region is necessary for wild-type expression and (ii) that the T-A-T-A box (a sequence in front of most eukaryotic genes) is not sufficient for wild-type promoter function. Thus, the yeast his3 promoter region appears large when compared with prokaryotic promoters, suggesting that it may be more complex than a simple site of interaction between RNA polymerase and DNA.