Carty M D, Lillquist J S, Peshavaria M, Stein R, Soeller W C
Department of Molecular Sciences, Central Research Division, Pfizer, Inc., Groton, Connecticut 06340, USA.
J Biol Chem. 1997 May 2;272(18):11986-93. doi: 10.1074/jbc.272.18.11986.
Islet amyloid polypeptide is expressed almost exclusively in pancreatic beta- and delta-cells. Here we report that beta cell-specific expression of the human islet amyloid polypeptide gene is principally regulated by promoter proximal sequences. The sequences that control tissue-specific expression were mapped between nucleotides -2798 and +450 of the human islet amyloid polypeptide (IAPP) gene using transgenic mice. To localize the cis-acting elements involved in this response, we examined the effects of mutations within these sequences using transfected islet amyloid polypeptide promoter expression constructs in pancreatic beta cell lines. The sequences between -222 and +450 bp were found to be necessary for beta cell-specific expression. Linker-scanning mutations of the 5'-promoter proximal region defined several key distinct control elements, including a negative-acting element at -111/-102 base pairs (bp), positive-acting elements like the basic helix-loop-helix-like binding site at -138/-131 bp, and the three A/T-rich, homeobox-like sites at -172/-163, -154/-142, and -91/-84 bp. Mutations within any one of these elements eliminated transcriptional expression by the promoter. Gel mobility shift assays revealed that the PDX-1 homeobox factor, which is required for insulin gene transcription in beta cells, interacted specifically at the -154/-142- and -91/-84-bp sites. Since PDX-1 is highly enriched in beta and delta cells, these results suggest that this factor plays a principal role in defining islet beta cell- and delta cell-specific expression of the IAPP gene.
胰岛淀粉样多肽几乎仅在胰腺β细胞和δ细胞中表达。在此我们报告，人胰岛淀粉样多肽基因的β细胞特异性表达主要受启动子近端序列调控。利用转基因小鼠，将控制组织特异性表达的序列定位在人胰岛淀粉样多肽（IAPP）基因的核苷酸-2798至+450之间。为了定位参与此反应的顺式作用元件，我们在胰腺β细胞系中使用转染的胰岛淀粉样多肽启动子表达构建体，检测了这些序列内突变的影响。发现-222至+450 bp之间的序列对于β细胞特异性表达是必需的。5'-启动子近端区域的接头扫描突变定义了几个关键的不同控制元件，包括位于-111/-102碱基对（bp）处的负性作用元件、位于-138/-131 bp处类似碱性螺旋-环-螺旋的结合位点等正性作用元件，以及位于-172/-163、-154/-142和-91/-84 bp处的三个富含A/T的、类似同源框的位点。这些元件中任何一个的突变都会消除启动子的转录表达。凝胶迁移率变动分析显示，β细胞中胰岛素基因转录所需的PDX-1同源框因子在-154/-142和-91/-84 bp位点特异性相互作用。由于PDX-1在β细胞和δ细胞中高度富集，这些结果表明该因子在定义IAPP基因的胰岛β细胞和δ细胞特异性表达中起主要作用。
