BTEB2, a Kruppel-like transcription factor, regulates expression of the SMemb/nonmuscle myosin heavy chain B (SMemb/NMHC-B) gene
BTEB2, a Kruppel-like transcription factor, regulates expression of the SMemb/nonmuscle myosin heavy chain B (SMemb/NMHC-B) gene
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DOI：
10.1161/01.res.85.2.182
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发表时间：
1999-07-23
影响因子：
20.1
通讯作者：
Nagai, R
中科院分区：
文献类型：
--
作者：
Watanabe, N;Kurabayashi, M;Nagai, R
我们最近鉴定了兔胚胎平滑肌肌球蛋白重链(SMemb/NMHC-B)基因的启动子区域，并从转录起始点-105处确定了15个碱基的序列，命名为SE1，这是一个重要的转录调控元件，在本研究中，我们试图通过与这个顺式调控元件结合来分离编码转录因子的cDNA克隆，这些转录因子通过与这个顺式调控元件结合来控制SMemb基因的表达。我们以含SE1的放射性标记的串联寡核苷酸为探针，从兔来源的SMC系C2/2细胞中筛选到了lambda gt11的cDNA文库。序列分析表明，其中一个克隆与兔碱性转录元件结合蛋白-2(BTEB2)的同源物相对应，后者此前已被鉴定为Kruppel样转录因子之一。C2/2细胞核提取物的凝胶迁移率分析和抗体超移位分析表明，BTEB2是核因子SE1复合体的主要成分。此外，谷胱甘肽S-转移酶-BTEB2融合蛋白以序列特异性的方式与SE1结合。为了支持BTEB2结合的功能，基础启动子活性和BTEB2诱导的转录激活显著地被SE1破坏而减弱。在成年兔组织中，BTEB2 mRNA在肠、膀胱和子宫中的表达最高，在正常发育过程中兔主动脉中BTEB2的表达水平下调。此外，免疫组织化学分析显示，大鼠主动脉球囊损伤后，新生内膜SMC中BTEB2蛋白的表达明显增强。这些结果表明，BTEB2介导了SMemb/NMHC-B基因的转录调控，并可能在血管SMC表型转换过程中调节基因表达。
We have recently characterized the promoter region of the rabbit embryonic smooth muscle myosin heavy chain (SMemb/NMHC-B) gene and identified the 15-bp sequence, designated SE1, located at -105 from the transcriptional start site as an important regulatory element for its transcriptional activity in a smooth muscle cell (SMC) line, In this study, we attempted to isolate cDNA clones encoding for the transcription factors that control the expression of the SMemb gene through binding to this cis-regulatory element. We screened a lambda gt11 cDNA library prepared from C2/2 cells, a rabbit-derived SMC line, by using a radiolabeled concatenated oligonucleotide containing SE1 as a probe. Sequence analysis revealed that one of the cDNA clones corresponds to the rabbit homologue of basic transcriptional element binding protein-2 (BTEB2), which has previously been identified as one of the Kruppel-like transcription factor. Gel mobility shift assays and antibody supershift analyses with nuclear extracts from C2/2 cells indicate that BTEB2 is a major component of nuclear factor:SE1 complexes. Furthermore, a glutathione S-transferase-BTEB2 fusion protein binds to the SE1 in a sequence-specific manner. In support of the functionality of BTEB2 binding, basal promoter activity and BTEB2-induced transcriptional activation were markedly attenuated by the disruption of the SE1. In adult rabbit tissues, BTEB2 mRNA was most highly expressed in intestine, urinary bladder, and uterus, BTEB2 mRNA levels were downregulated in rabbit aorta during normal development. Moreover, immunohistochemical analysis indicated a marked induction of BTEB2 protein in the neointimal SMC after balloon injury in rat aorta. These results suggest that BTEB2 mediates the transcriptional regulation of the SMemb/NMHC-B gene and possibly plays a role in regulating gene expression during phenotypic modulation of vascular SMC.