Isolation of complementary DNAs for heat shock protein (HSP) 70 and heat shock cognate protein (HSC) 70 genes and the expressions in post-ischaemic gerbil brain.
Isolation of complementary DNAs for heat shock protein (HSP) 70 and heat shock cognate protein (HSC) 70 genes and the expressions in post-ischaemic gerbil brain.
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DOI：
10.1080/01616412.1992.11740088
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发表时间：
1992
影响因子：
1.9
通讯作者：
Shigeru Sato;Koji Abe;J. Kawagoe;M. Aoki;K. Kogure
中科院分区：
文献类型：
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作者：
Shigeru Sato;Koji Abe;J. Kawagoe;M. Aoki;K. Kogure
以沙土鼠双侧颈总动脉结扎10 min后再灌注8 h的大脑皮层为材料，构建了沙土鼠大脑皮层cDNA文库。用人HSP 70基因组DNA探针（pH2.3）对该cDNA文库进行第三次筛选，得到4个cDNA克隆（分别命名为pGA 3、pGB 1、pGD 3和pGE 4）。Southern和北方杂交分析及部分核苷酸序列分析表明，pGA 3和pGE 4为HSP 70 cDNA克隆，pGB 1和pGD 3为HSC 70 cDNA克隆，它们分别选择性识别HSP 70和HSC 70 mRNA。HSP70 mRNA在正常对照中以非常少量存在，并且在短暂缺血后被大量诱导。HSC 70 mRNA在正常沙鼠脑中组成型表达，但仍是可诱导的。原位杂交结果显示，假手术组海马锥体细胞和齿状颗粒细胞中HSP70mRNA表达量很低，再灌注8h后，海马、齿状回、内侧缰核、丘脑腹侧核、尾壳核、下丘脑腹内侧核、弓状核、杏仁核和大脑皮质细胞中HSP70mRNA表达量明显增加。HSC 70 mRNA在假手术对照的几乎相同的区域中存在，并且在再灌注8h后轻微诱导。我们的研究结果表明，HSP 70和HSC 70的cDNA克隆首次从缺血后的沙土鼠脑中分离，并选择性地识别HSP 70或HSC 70的mRNA，分别。用原位杂交技术观察了沙土鼠脑缺血后HSP 70和HSC 70 mRNA表达的区域差异。
A complementary DNA (cDNA) library was constructed with a plasmid vector from cerebral cortices of gerbils at 8 h of reperfusion after 10 min of bilateral common carotid ligation. After the 3rd screening of this cDNA library with a human genomic DNA probe for HSP70 (pH2.3), 4 cDNA clones were isolated (named pGA3' pGB1' pGD3 and pGE4' respectively). Southern and Northern blot analysis, and partial nucleotide sequence analysis indicated that pGA3 and pGE4 were the HSP70 cDNA clones, and that pGB1 and pGD3 were the HSC70 cDNA clones, which selectively recognized HSP70 or HSC70 mRNA, respectively. HSP70 mRNA is present in a very small amount in normal controls, and is greatly induced after the transient ischaemia. HSC70 mRNA is constitutively expressed in a normal gerbil brain, but is still inducible. In situ hybridization study demonstrated that the HSP70 mRNA was present in a very small amount in the hippocampal pyramidal and dentate granule cells in the sham control, and that the mRNA was greatly induced in the cells of hippocampus, dentate gyrus, medial habenula, ventral thalamic nuclei, caudate putamen, ventromedial and arcuate hypothalamic nuclei, amygdaloid nuclei and cerebral cortex after 8 h of reperfusion. HSC70 mRNA was present in almost the same areas of sham control, and was slightly induced after 8 h of reperfusion. Our results indicate that HSP70 and HSC70 cDNA clones were first isolated from post-ischaemic gerbil brain, and selectively recognize HSP70 or HSC70 mRNA, respectively. A regional difference in the induction of the HSP70 and HSC70 mRNA in post-ischaemic gerbil brain was observed by in situ hybridization.