DNA clones representing the negative acute-phase gene coding for the plasma proteinase inhibitor alpha 1-inhibitor III were isolated from a rat genomic library. Structural analysis established the existence of at least four different members of the alpha 1-inhibitor III gene family. Partial DNA sequence analysis of the 5'-terminal regions was performed for the alpha 1-inhibitor III gene and the related alpha 1-inhibitor IV gene. The transcription start site of the alpha 1-inhibitor III gene was located by S1 mapping and primer extension. No stable alpha 1-inhibitor IV mRNA was detected in rat liver. In an experimentally induced acute-phase reaction, the transcription rate of the alpha 1-inhibitor III gene was reduced 12.7-fold at 6 h after stimulation. Four hours after injection of a high dose of dexamethasone into rats, the transcription rate of this gene was reduced 9-fold. Thus, glucocorticoids alone are capable of causing a strong transient down-regulation of the transcription of this gene in rats, independent of other inflammatory mediators. An inverted consensus glucocorticoid responsive element (5'GGACACAATAT3') shared with the glucocorticoid-regulated alpha 1-fetoprotein, alpha 2u-globulin, and alpha 1-acid glycoprotein genes was detected by computer-assisted sequence analysis in the promoter proximal 5'-flanking region of the alpha 1-inhibitor III gene.