Abundant expression of Kallikrein 1 gene in human keratinocytes was mediated by GATA3
Abundant expression of Kallikrein 1 gene in human keratinocytes was mediated by GATA3
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DOI：
10.1016/j.gene.2009.02.002
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发表时间：
2009-05-01
期刊：
影响因子：
3.5
通讯作者：
Saijoh, Kiyofumi
中科院分区：
文献类型：
--
作者：
Son, Do Ngoc;Li, LiHua;Saijoh, Kiyofumi
在组织激肽释放酶基因（KLK）中，KLK 1在人类皮肤中大量表达。虽然已知其推定的启动子具有多种顺式元件，但尚未对其进行功能测试。在本研究中，研究了支持如此丰富表达的KLK 1启动子的调节机制。在NHEK人角质形成细胞上进行靶向KLK 1启动子（主要转录起始位点的核苷酸-1153/+40）的荧光素酶测定。-954/-855、-428/-236和-100/+40具有诱导活性。基序搜索程序未能在-428/-236中找到独特的结合基序，而-954/-855和-100/+40都具有独特的GATA结合基序。电泳迁移率变动分析（EMSA）和DNA足迹法证实了NHEK核蛋白与这些被抗GATA 3抗体超移位的基序的结合。在加塔各亚型中，GATA 3单独存在时，可在NHEK RNA中扩增。此外，将GATA 3导入成纤维细胞NIH 3 T3中，可增强含-954/+40的KLK 1启动子的活性，而GATA 3显性失活突变体对NHEK细胞的KLK 1启动子活性则有一定的影响。因此，发现GATA 3结合位于-954/-855的位点，并且是人角质形成细胞中丰富的KLK 1表达的关键调节剂。(C)2009爱思唯尔有限公司版权所有。
Among Tissue kallikrein genes (KLKs), KLK1 is abundantly expressed inhuman skin. Although its putative promoter is known to have various cis-elements, they have not been functionally tested. In the present study, the regulation mechanism of KLK1 promoter supporting such abundant expression was examined. Luciferase assay targeting the KLK1 promoter (nucleotide - 1153/+40 from the major transcriptional start site) was performed on NHEK human keratinocyte. -954/-855,-428/-236, and -100/+40 had the induction activity. The motif search program failed to find unique binding motifs in -428/-236, whereas both -954/-855 and -100/+40 had a unique GATAs binding motif. Electrophoretic mobility shift assay (EMSA) and DNA footprinting confirmed the binding of NHEK nuclear protein to these motifs that were supershifted by anti-GATA3 antibody. Among GATA isoforms, GATA3 alone could be amplified in RNA obtained from NHEK Moreover, introduction of GATA3 into fibroblastic NIH3T3 cells enhanced the activity of KLK1 promoter containing -954/+40, while that of GATA3 dominant negative mutant to NHEK cells impaired the same promoter's activity. Thus, GATA3 was found to bind the site located at -954/-855 and to be a key regulator of abundant KLK1 expression in human keratinocyte. (C) 2009 Elsevier B.V. All rights reserved.