The tobacco anionic peroxidase gene encodes the predominant peroxidase isoenzyme in the aerial portions of tobacco. Three kb of the peroxidase promoter was joined to the coding region of the Escherichia coli beta-glucuronidase gene (GUS), and transiently expressed in tobacco mesophyll protoplasts in the presence or absence of plant growth regulators. Benzyladenine, ethylene, and gibberellic acid did not affect peroxidase gene expression. Abscisic acid slightly inhibited expression at high concentrations. The auxins indole-3-acetic acid (IAA) and naphthaleneacetic acid strongly suppressed peroxidase expression. We observed half maximal suppression at 30 microM IAA. An anti-auxin, p-chlorophenoxyisobutyric acid (PCIB), enhanced expression from the peroxidase promoter above that of untreated controls or restored activity when used in combination with IAA. Sequencing 3 kb of the peroxidase promoter revealed many potential regulatory elements based on sequence homology to previously characterized genes. This includes several consensus transcription factor binding sites found in auxin-regulated promoters. 5' deletions of the peroxidase promoter/GUS fusion revealed several positive and negative regulatory elements. An upstream enhancer element was found between -3146 and -638 from the start of transcription. A strong silencer element was observed between -638 and -220. Removal of this silencer resulted in a truncated promoter (-220) with 100% activity of the full-length promoter (-3146). Inhibition by auxin was observed with all 5' deletions.