Transcription factor Sp1 regulates SERCA2 gene expression in pressure-overloaded hearts: a study using in vivo direct gene transfer into living myocardium.
Transcription factor Sp1 regulates SERCA2 gene expression in pressure-overloaded hearts: a study using in vivo direct gene transfer into living myocardium.
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DOI：
10.1016/s0022-2828(03)00122-6
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发表时间：
2003-07
影响因子：
5
通讯作者：
T. Takizawa;M. Arai;Koichi Tomaru;N. Koitabashi;D. L. Baker;M. Periasamy;M. Kurabayashi
中科院分区：
文献类型：
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作者：
T. Takizawa;M. Arai;Koichi Tomaru;N. Koitabashi;D. L. Baker;M. Periasamy;M. Kurabayashi
压力过载肥厚导致肌浆网Ca2+- atp酶泵的下调，编码SERCA2基因，调节Ca2+摄取和心肌松弛。我们之前描述了一个近端启动子区域包含四个Sp1元件一致序列（-284到-72碱基对（bp）），负责压力过载诱导的转录调控。本研究的目的是确定哪一个Sp1位点负责压力过载下SERCA2基因转录的下调。使用体内直接基因转移法，在压力过载下测量SERCA2基因转录活性。对SERCA2基因启动子（-284至-72 bp）的四个Sp1位点（I-IV）进行定点突变。将野生型和Sp1突变型荧光素酶报告基因构建体注射到压力过载或假手术大鼠的左心室尖，并在手术后3至14 d连续测量Sp1 mRNA和SERCA2基因荧光素酶活性。在第5天，压力过载大鼠的Sp1 mRNA水平增加到假手术组的124±7%，当所有四个Sp1位点保持完整时，压力过载诱导的SERCA2转录活性为假手术组的15±4%。Sp1突变位点I （-196 ~ -191 bp）和III （-118 ~ -113 bp）的突变阻断了压力过载的抑制作用，导致sham组SERCA2基因转录活性分别为54±15%和56±7%。我们得出结论，压力过载诱导的SERCA2 mRNA减少是由Sp1位点I和III介导的。
Pressure-overload hypertrophy results in downregulation of the sarcoplasmic reticulum Ca2+-ATPase pump encoding SERCA2 gene that regulates Ca2+uptake and myocardial relaxation. We previously characterized a proximal promoter region containing four Sp1 element consensus sequences (–284 to –72 base pairs (bp)) that was responsible for pressure-overload-induced transcriptional regulation. The purpose of the present study was to determine which of the Sp1 sites was responsible for the downregulation of SERCA2 gene transcription under pressure overload. Using an in vivo direct gene transfer assay, SERCA2 gene transcriptional activity was measured under pressure overload. Site-directed mutagenesis of the four Sp1 sites (I-IV) in the SERCA2 gene promoter (–284 to –72 bp) was performed. Wild-type and Sp1 mutant-luciferase reporter constructs were injected into the left-ventricular apices of pressure overload or sham-operated rats, and Sp1 mRNA and SERCA2 gene-luciferase activity was measured sequentially from 3 to 14 d after surgery. At 5 d, Sp1 mRNA in the pressure-overload rats increased to 124 ± 7% of sham group levels, and pressure-overload-induced SERCA2 transcriptional activity was 15 ± 4% of sham group when all four Sp1 sites remained intact. Mutation of the Sp1 mutant sites I (–196 to –191 bp) and III (–118 to –113 bp) blocked the inhibitory effect of pressure overload and resulted in SERCA2 gene transcriptional activity of 54 ± 15% and 56 ± 7% of sham group, respectively. We conclude that the pressure-overload-induced decrease in SERCA2 mRNA is mediated by Sp1 sites I and III.