Detection of transcription factors associated with cell-type specific transcription of the secretory leukoprotease inhibitor gene in epithelial cells.

分泌型白蛋白水解酶抑制物(SLPI)是一种丝氨酸蛋白酶抑制物，在呼吸道和生殖腺中局部产生，但在肝脏中不产生。在本研究中，我们分析了该基因的启动子区域，以更好地了解参与转录调控的分子机制。在表达SLpi mRNA的II型肺泡细胞系A549和宫颈癌细胞系HeLa的染色质结构中，均在外显子I上游1kbp处检测到DNase-I超敏部位。以荧光素酶基因作为瞬时表达载体，通过缺失分析研究了SLPI启动子围绕这些DNase-I超敏感位点的功能。在这项分析中，我们发现了三个转录控制区，它们在A549细胞中起作用，但在非肺细胞系中不起作用，如HeLa和肝癌Hep G2。在三个顺式调控区中，近端的41个碱基对(相对于转录起始点-132到-92个碱基)是最显著的转录活性区域。该区域对应于在另一个肺细胞系HS-24中检测到的转录激活序列，表明这个41个碱基的序列是肺细胞特异性表达所必需的。电泳迁移率改变分析表明，这一41个碱基的启动子区域包含一个11个碱基的核结合蛋白识别序列，其中一个在肺细胞系中丰富，另一个在非肺细胞系中丰富。这些结果表明，这两种核糖体结合蛋白的比例在SLPI基因的表达模式上具有细胞类型特异性。

Secretory leukoprotease inhibitor (SLPI) is a serine protease inhibitor, produced locally in respiratory and genital glands, but not in the liver. In the present study the promoter region of this gene was analyzed to better understand the molecular mechanisms involved in transcriptional regulation. DNase-I hypersensitive sites were detected within 1 kbp upstream of exon I in chromatin structures of type II pneumocyte cell line A549 and uterocervical cell line HeLa, both of which express SLPI mRNA transcripts. The function of the SLPI promoter encompassing these DNase-I hypersensitive sites has been studied by deletion analysis with the luciferase gene as a transient expression vector. In this analysis, we foud three transcription control regions that function in A549 cells but not in nonlung cell lines, such as HeLa and hepatoma Hep G2. Among three cis-regulatory regions, a proximal 41-bp region (-132 to -92 bp relative to the transcription start site) is responsible for the most striking magnitude of transcriptional activity. This region corresponds to the transcriptional activating sequence detected in another lung cell line, HS-24, indicating that this 41-bp sequence is required for lung cell-specific expression. An electrophoretic mobility shift assay demonstrated that this 41-bp promoter region contains an 11-bp recognition sequence for two nuclear binding proteins, one of which is abundant in lung cell lines, and the other in nonlung cell lines. These results suggest that the ratio of these two nuclesr binding proteins confers the cell type specificity on the expression pattern of the SLPI gene.