家蚕变态过程中血细胞组织蛋白酶B cDNA的分离与表达
Isolation and expression of cathepsin B cDNA in hemocytes during metamorphosis of Bombyx mori.
作者信息
Xu Y, Kawasaki H
机构信息
College of Animal Sciences, Zhejiang University, 310029, Hangzhou, PR China
出版信息
Comp Biochem Physiol B Biochem Mol Biol. 2001 Oct;130(3):393-9. doi: 10.1016/s1096-4959(01)00448-1.
By using RT-PCR and 5'- and 3'-RACE methods, we isolated cathepsin B cDNA from hemocytes of Bombyx mori. The predicted open reading frame encoded 337 amino acids with 54% identity with the human cathepsin B, and 55% identity with the 29-kDa proteinase of the blowfly. Three active sites characteristic for cathepsin B were conserved in the deduced amino acid sequences of BmCtB cDNA at positions Cys-111, His-280 and Asn-300. Northern analysis identified a 1.3-kb mRNA. Expression of BmCtB was observed in the hemocytes from the day of wandering and was strongest at day 1 pupa. With in situ hybridization, strong signals were observed in the granular cells and plasmatocytes of day 0 pupa, but not in those of day 5 of the fifth larval instar. The results indicate another function of cathepsin B and insect hemocyte during metamorphosis.
通过使用逆转录聚合酶链反应（RT-PCR）以及5'和3'端快速扩增cDNA末端（5'-和3'-RACE）方法，我们从家蚕血细胞中分离出组织蛋白酶B的cDNA。预测的开放阅读框编码337个氨基酸，与人类组织蛋白酶B的同源性为54%，与绿蝇29 kDa蛋白酶的同源性为55%。组织蛋白酶B特有的三个活性位点在家蚕组织蛋白酶B（BmCtB）cDNA推导的氨基酸序列中，在半胱氨酸-111、组氨酸-280和天冬酰胺-300位置保守。Northern印迹分析鉴定出一个1.3 kb的mRNA。在化蛹当天的血细胞中观察到BmCtB的表达，在蛹期第1天最强。通过原位杂交，在化蛹第0天的颗粒细胞和浆细胞中观察到强信号，但在五龄幼虫第5天的细胞中未观察到。结果表明组织蛋白酶B和昆虫血细胞在变态过程中的另一种功能。
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