A juvenile hormone-repressible transferrin-like protein from the bean bug, Riptortus clavatus: cDNA sequence analysis and protein identification during diapause and vitellogenesis.
A juvenile hormone-repressible transferrin-like protein from the bean bug, Riptortus clavatus: cDNA sequence analysis and protein identification during diapause and vitellogenesis.
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DOI：
10.1002/(sici)1520-6327(200005)44:1
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发表时间：
2000-05
影响因子：
2.2
通讯作者：
Makoto Hirai;Daisuke Watanabe;Yasuo Chinzei
中科院分区：
文献类型：
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作者：
Makoto Hirai;Daisuke Watanabe;Yasuo Chinzei
我们通过使用 mRNA 差异显示，在豆虫 Riptortus clavatus 中发现了几种保幼激素反应性 cDNA（Hirai 等人，1998）。其中之一是保幼激素抑制性 cDNA JR-3，经过克隆、测序、表征并鉴定为转铁蛋白 (RcTf)。 RcTf cDNA 编码 652 个氨基酸，计算分子量为 71,453 Da。推导的氨基酸序列与几种昆虫的转铁蛋白基因显着同源，这些昆虫包括Manduca sexta（43%同一性）、Blaberus disoidalis（43%）、埃及伊蚊（43%）、果蝇（36%）、Sarcophaga peregrina（36%）和人类（25%）。以大肠杆菌表达的重组RcTf蛋白为抗原制备抗血清。抗血清与虫子的重组蛋白和天然蛋白发生特异性反应，大小分别为 70 kDa 和 75 kDa。从滞育雌虫的血淋巴中部分纯化了75 kDa蛋白，发现前10个氨基酸与RcTf cDNA相同，表明75 kDa蛋白是RcTf。通过蛋白质印迹分析检查 RcTf 在 bug 中的组织分布。在滞育动物中，在脂肪体、血淋巴和卵巢中检测到 RcTf，但在肠道中未检测到。在滞育后阶段，除了脂肪体和卵巢外，还在鸡蛋中检测到RcTf。这些结果表明，RcTf 在卵黄发生过程中被掺入卵母细胞中，并表明它可能为发育中的胚胎提供铁。
We found several juvenile hormone-responsive cDNAs in the bean bug, Riptortus clavatus, by using mRNA differential display (Hirai et al., 1998). One of them, a juvenile hormone-repressible cDNA, JR-3, was cloned, sequenced, characterized and identified as a transferrin (RcTf). RcTf cDNA encoded 652 amino acids with a calculated molecular weight of 71,453 Da. The deduced amino acid sequence showed significant homology with the transferin genes of several insects, Manduca sexta (43% identity), Blaberus discoidalis (43%), Aedes aegypti (43%), Drosophila melanogaster (36%), Sarcophaga peregrina (36%) and the human (25%). Antiserum was prepared by using recombinant RcTf protein expressed in Escherichia coli as an antigen. The antiserum reacted specifically with both the recombinant protein and the native protein from the bugs, with sizes of 70 and 75 kDa, respectively. The 75 kDa protein was partially purified from hemolymph of diapausing female bugs and the first ten amino acids were found to be identical to that of RcTf cDNA, indicating that the 75 kDa protein is RcTf. The tissue distribution of RcTf in the bug was examined by Western blot analysis. In diapausing animals, RcTf was detected in the fat body, hemolymph and ovary but not in the gut. In the post-diapause stage, RcTf was also detected in eggs, in addition to the fat body and ovary. These results indicate that RcTf is incorporated into the oocytes during vitellogenesis, and suggest that it may provide iron for the developing embryos.