A novel RNA helicase-like protein during early embryonic development in silkworm Bombyx mori:: Molecular characterization and intracellular localization
A novel RNA helicase-like protein during early embryonic development in silkworm Bombyx mori:: Molecular characterization and intracellular localization
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DOI：
10.1016/j.ibmb.2006.09.002
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发表时间：
2006-12-01
影响因子：
3.8
通讯作者：
Iino, T.
中科院分区：
文献类型：
--
作者：
Sawada, H.;Yamahama, Y.;Iino, T.
为了解家蚕滞育卵和非滞育卵早期胚胎发育的分子机制，采用差异显示技术比较了滞育卵和非滞育卵的mRNA。我们用RACE方法克隆了编码一种新的RNA解旋酶(RHL)蛋白的全长cDNA，该片段是非滞育卵中的一种特异的cDNA。用预测的RHL的氨基酸序列进行BLAST搜索，发现它与DEAD-box RNA解旋酶的同源性很低(部分长度的同源性为21-25%)。在产卵后60h，用RT-PCR方法研究了滞育卵和非滞育卵中RHL基因的表达。在非滞育卵的各个时期均可观察到扩增的RHL基因，而在滞育卵中，在产卵后0-12h可检测到该基因，但在产卵后24-60h已消失。当滞育卵在产卵后48h(人工终止滞育)后，在4℃下冷却6d后，用盐酸处理激活滞育卵，从12h开始观察到cDNAs。我们还用抗重组His-tag RHL抗血清研究了RHL在非滞育卵中的免疫组织化学分布和定位。RHL分布于胚层细胞和卵黄细胞，定位于卵黄细胞的胞核和胞浆。这些数据表明，RHL在家蚕早期胚胎中起着重要的作用。(C)2006爱思唯尔有限公司。保留所有权利。
In order to understand the molecular mechanism of development during early embryogenesis in diapause and non-diapause of the silkworm, mRNA from diapause and non-diapause eggs was compared using the differential display technique. We cloned the full length of a cDNA encoding a novel RNA helicase-like (RHL) protein by the RACE method using a cDNA fragment which was one of the specific cDNAs in the non-diapause eggs. A BLAST search using the predicted amino acid sequence of RHL revealed a low homology (21-25% identity of its partial length) with that of the DEAD-box RNA helicase. Gene expression of the RHL gene of the diapause and non-diapause eggs was investigated by RT-PCR until 60 h after oviposition. Amplified RHL cDNA was observed through all the stages in the non-diapause eggs, while in the diapause eggs, cDNA was found in eggs 0-12h after oviposition but disappeared 24-60h after oviposition. When the diapause eggs were activated by HCl treatment after chilling at 4 degrees C for 6 days from 48h after oviposition (artificial diapause termination), cDNA was observed from 12h after HCl treatment. We also investigated the immunohistochemical distribution and localization of RHL in non-diapause eggs using anti-recombinant His-tag RHL antiserum. RHL was distributed in blastoderm cells and yolk cells and was localized in the nucleus and the cytosol of yolk cells. These data suggest that RHL has an important role in the early embryo of the silkworm. (c) 2006 Elsevier Ltd. All rights reserved.