Pei L, Melmed S
Division of Endocrinology, Cedars-Sinai Medical Center-UCLA School of Medicine 90048, USA.
Biochem J. 1995 Jun 15;308 ( Pt 3)(Pt 3):719-23. doi: 10.1042/bj3080719.
The broad spectrum of vasoactive intestinal polypeptide (VIP) cellular functions are mediated by high-affinity binding sites. To determine regulation of the VIP receptor gene expression, we have isolated and characterized two genomic clones that contain the first three exons and the 5' flanking region of the VIP receptor gene. Using RNase protection assays, receptor gene expression was detected in adult rat lung, liver and intestine, but not in fetal lung, indicating that VIP receptor is expressed in diverse tissues, and its expression is differentially regulated during lung development. The transcription start site of the gene was mapped to a cytosine residue, 76 bp upstream from the ATG initiation codon. Transfection into rat lung cell lines shows that although 126 bp of the VIP receptor 5' DNA sequences are capable of activating VIP receptor gene basal transcription 30-fold over a promoterless control, 488 bp of the 5' sequences further induce this activation to 97-fold over control. However, inclusion of up to 859 bp 5' sequences results in a decrease in basal promoter activity (31-fold over control), indicating that while sequences between -126 and -488 bp contain potential enhancer sequences, sequences between -488 and -859 bp may include a transcriptional repressor sequence. Deletion analysis shows that transcription factor Sp1 plays an important role in activating basal promoter of the VIP receptor gene. DNase I footprinting and gel-mobility-shift assays show that Sp1 binds to its consensus binding sites in the VIP receptor promoter, suggesting that interaction of Sp1 with VIP receptor promoter transactivates this gene.
血管活性肠肽（VIP）广泛的细胞功能是由高亲和力结合位点介导的。为了确定VIP受体基因表达的调控机制，我们分离并鉴定了两个基因组克隆，它们包含VIP受体基因的前三个外显子和5'侧翼区域。使用核糖核酸酶保护分析，在成年大鼠的肺、肝和肠中检测到受体基因表达，但在胎儿肺中未检测到，这表明VIP受体在多种组织中表达，并且其表达在肺发育过程中受到差异调节。该基因的转录起始位点被定位到一个胞嘧啶残基，位于ATG起始密码子上游76 bp处。转染到大鼠肺细胞系中显示，尽管VIP受体5' DNA序列的126 bp能够在无启动子对照的基础上激活VIP受体基因基础转录30倍，但5'序列的488 bp能将这种激活进一步诱导至对照的97倍。然而，包含多达859 bp的5'序列会导致基础启动子活性降低（对照的31倍），这表明虽然-126至-488 bp之间的序列包含潜在的增强子序列，但-488至-859 bp之间的序列可能包含转录抑制序列。缺失分析表明转录因子Sp1在激活VIP受体基因的基础启动子中起重要作用。DNase I足迹分析和凝胶迁移率变动分析表明Sp1与其在VIP受体启动子中的共有结合位点结合，这表明Sp1与VIP受体启动子的相互作用可反式激活该基因。
