The alpha-1-antitrypsin gene promoter in human A549 lung derived cells, and a novel transcription initiation site

Abstract

Alpha-1-antitrypsin (AAT), also called serine proteinase inhibitor A1 (Serpin A1), is the most abundant serpin in human plasma. A major physiological role of AAT is to protect the lung from the destructive effects of excess uninhibited neutrophil elastase. During inflammation, circulating levels of AAT may increase twofold-to-threefold as part of the acute-phase response. The liver is the main contributor to this increase. However, local synthesis may provide an important mechanism for controlling neutrophil elastase activity at sites of inflammation, and previous studies have shown a marked increase in production after cytokine stimulation. In the current study we report a distinct transcription initiation site for AAT expression in the lung alveolar epithelial cell line A549, which is located nine bases upstream of the previously mapped full-length monocyte transcription start-site, and show using site-directed mutagenesis that two Sp1 sites and a putative TATA box are functional. EMSA experiments provide evidence for Sp1 and Sp3 binding to these two Sp1 sites. We have also mapped the minimal promoter region and a cell-specific element essential for expression in A549 cells, both of which reside in an 865bp fragment upstream of the transcription start-site. Understanding the mechanisms of AAT gene regulation in a lung-derived cell line has important implications for understanding the control of localised lung tissue damage which occurs as a result of excess proteolytic activity.