Characterization of a midgut-specific chitin synthase gene (LmCHS2) responsible for biosynthesis of chitin of peritrophic matrix in Locusta migratoria
Characterization of a midgut-specific chitin synthase gene (LmCHS2) responsible for biosynthesis of chitin of peritrophic matrix in Locusta migratoria
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负责飞蝗围食基质几丁质生物合成的中肠特异性几丁质合酶基因 (LmCHS2) 的表征
DOI：
10.1016/j.ibmb.2012.09.002
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发表时间：
2012-12-01
影响因子：
3.8
通讯作者：
Zhang, Jianzhen
中科院分区：
文献类型：
--
作者：
Liu, Xiaojian;Zhang, Huanhuan;Zhang, Jianzhen
甲壳素是围食基质的重要组成部分，它是由一系列生物化学反应产生的。几丁质合成酶在几丁质生物合成途径中起着重要的作用。本研究从东亚飞蝗中克隆了几丁质合成酶2基因（LmCHS2）的全长cDNA。该cDNA的开放阅读框为4569个核苷酸，编码1523个氨基酸残基，5 '和3'非编码区分别为76和373个核苷酸。利用实时定量PCR分析LmCHS2在蝗虫不同组织中的转录本表明，LmCHS2仅在中肠和胃盲囊（中肠的一部分）中表达。在中肠前部的表达量最高，转录水平从前部到后部逐渐下降。在蝗虫的生长发育过程中，该基因在卵中只有微弱的表达，但从幼虫到成虫表达量逐渐增加。原位杂交结果显示LmCHS2主要表达于胃盲囊刷状缘柱状细胞的顶端。将LmCHS2双链RNA注射到5龄家蚕体内，进一步研究LmCHS2的生物学功能。显著下调LmCHS2的转录导致昆虫停止进食和高死亡率。但在成虫蜕皮前，蝗虫的形态没有明显的异常变化。解剖后，我们发现注射LmCHS2 dsRNA的蝗虫中肠的平均长度比注射dsGFP的对照昆虫短。此外，中肠的显微切片显示，与对照组相比，注射LmCHS2 dsRNA的大鼠的PM是无定形的和薄的。我们的研究结果表明，LmCHS2负责与PM相关的几丁质的生物合成，并在蝗虫的生长和发育中起着至关重要的作用。(C)2012爱思唯尔有限公司保留所有权利。
Chitin, an essential component of peritrophic matrix (PM), is produced by a series of biochemical reactions. Chitin synthase plays a crucial role in chitin polymerization in chitin biosynthetic pathway. In this study, we identified and characterized a full-length cDNA of chitin synthase 2 gene (LmCHS2) from Locusta migratoria. The cDNA contains an open reading frame of 4569 nucleotides that encode 1523 amino acid residues, and 76- and 373-nucleotides for 5'- and 3'-noncoding regions, respectively. Analysis of LmCHS2 transcript in different tissues of the locust by using real-time quantitative PCR indicated that LmCHS2 was exclusively expressed in midgut and gastric caeca (a part of the midgut). The highest expression was found in the anterior midgut with a decline of the transcript level from the anterior to posterior regions. During growth and development of locusts, there was only a slight expression in eggs, but the expression gradually increased from nymphs to adults. In situ hybridization further revealed that LmCHS2 transcript mainly presented in the apical regions of brush border forming columnar cells of gastric caeca. LmCHS2 dsRNA was injected to fifth-instar nymphs to further explore biological functions of LmCHS2. Significantly down-regulated transcript of LmCHS2 resulted in a cessation of feeding and a high mortality of the insect. However, no visible abnormal morphological change of locusts was observed until insects molted to adults. After dissection, we found that the average length of midguts from the LmCHS2 dsRNA-injected locusts was shorter than that of the control insects that were injected with dsGFP. Furthermore, microsection of midguts showed that the PM of the LmCHS2 dsRNA-injected nymphs was amorphous and thin as compared with the controls. Our results demonstrate that LmCHS2 is responsible for the biosynthesis of chitin associated with PM and plays an essential role in locust growth and development. (C) 2012 Elsevier Ltd. All rights reserved.