Identification and analysis of the promoter region of the human HAS3 gene.
Identification and analysis of the promoter region of the human HAS3 gene.
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人HAS3基因启动子区的鉴定和分析。
DOI：
10.1016/j.bbrc.2015.03.142
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发表时间：
2015-05
期刊：
影响因子：
--
通讯作者：
Bu, Youquan
中科院分区：
文献类型：
--
作者：
Ji, Ying;Du, Gang;Wang, Yitao;Bu, Youquan
透明质酸（HA）是脊椎动物细胞外基质的关键组分，其由透明质酸脱氢酶（包括HAS 1、HAS 2和HAS 3）在质膜处合成。HAS 1 -3的表达和调节涉及许多生理和病理过程。人HAS 1和HAS 2基因的启动子已被鉴定，而HAS 3基因的启动子尚不清楚。在本研究中，我们首次鉴定和表征了人HAS 3基因启动子区。5′ RACE分析发现HAS 3基因存在两个新的转录起始位点。HAS 3基因5′侧翼区的渐进缺失分析表明，HAS 3近端启动子主要局限于450 bp区域（即主要HAS 3转录起始位点上游的−761至−305 bp），而其核心启动子位于最小的129 bp区域（即主要HAS 3转录起始位点上游的−433至−305 bp）。转录因子结合分析表明，HAS 3基因启动子缺少经典的TATA盒，但含有经典的GC盒以及C/EBP和NFκB等转录因子的结合位点。此外，定点突变实验表明，近端Sp1结合位点是HAS 3基因稳定的近端启动子活性所必需的，而核心启动子基序（core promotermotiftenelements，MTE）基序是HAS 3基因基本的核心启动子活性所必需的。本研究为进一步研究该基因的表达调控机制奠定了基础。
Hyaluronan (HA) is a key component of the vertebrate extracellular matrix that is synthesized at the plasma membrane by the hyaluronan synthases including HAS1, HAS2 and HAS3. The expression and regulation of HAS1-3 are implicated in numerous physiological and pathological processes. The promoters of human HAS1 and HAS2 genes have been identified previously whereas HAS3 promoter remains unclear. In the present study, we have for the first time identified and characterized the human HAS3 gene promoter region. 5′ RACE assay revealed two novel transcriptional variants of HAS3 gene with distinct transcription start sites. Progressive deletion analysis of the 5′-flanking region of HAS3 gene demonstrated that HAS3 proximal promoter is mainly restricted to a 450-bp region (i.e. −761 to −305 bp upstream of the major HAS3 transcription start site), whereas its core promoter is located to a minimal 129-bp region (i.e. −433 to −305 bp upstream of the major HAS3 transcription start site). Transcriptional factor binding analysis indicated that HAS3 gene promoter lacks of canonical TATA box, but contains classical GC box as well as other putative binding sites for transcriptional factors such as C/EBP and NFκB. In addition, site-directed mutagenesis assay demonstrated that the proximal Sp1 binding site is essential for the robust proximal promoter activity of HAS3 gene whereas the core MTE (core promotermotiftenelements) motif is required for the basic core promoter activity of HAS3 gene. Our present study should facilitate further studies on the mechanism regulating the expression of this important gene.
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