Downregulation of a Chitin Deacetylase-Like Protein in Response to Baculovirus Infection and Its Application for Improving Baculovirus Infectivity
Downregulation of a Chitin Deacetylase-Like Protein in Response to Baculovirus Infection and Its Application for Improving Baculovirus Infectivity
复制标题
DOI：
10.1128/jvi.01860-09
复制
发表时间：
2010-03-01
影响因子：
5.4
通讯作者：
Herrero, Salvador
中科院分区：
文献类型：
--
作者：
Jakubowska, Agata K.;Caccia, Silvia;Herrero, Salvador
从棉铃虫感染棉铃虫单核多角体病毒后表达变化的一组基因中筛选出几个与甲壳素脱乙酰酶样蛋白(CDA)同源的表达序列标签(EST)。其中一些EST编码含有甲壳素脱乙酰基酶结构域(CDAD)的中肠蛋白。表达的蛋白HaCDA5a不具有甲壳素脱乙酰酶活性，但与甲壳素结合有很强的亲和力。序列分析表明，目前所有已知的营养周膜(PM)蛋白都缺乏甲壳素结合区。此前在棉铃虫PM中检测到HaCDA5a这种定位，加上病原体感染后它的下调，使我们假设该蛋白可能与PM结构的动态平衡有关，并且通过减少其表达，昆虫可能会降低PM的通透性，从而减少杆状病毒的进入。为了验证这一假设，我们构建了在昆虫细胞中表达HaCDA5a的重组核型多角体病毒，并测试了其对质膜通透性的影响以及HaCDA5a表达对杆状病毒性能的影响。实验表明，HaCDA5a以浓度依赖的方式增加质膜的通透性。对油菜夜蛾和甜菜夜蛾幼虫的生物测定表明，表达HaCDA5a的NPV比其亲本病毒具有更强的侵染性。然而，当病毒被注射到血腔内时，没有观察到毒力的差异。这些发现支持下调一种中肠特异的CDA样蛋白，这是棉铃虫通过降低PM通透性来降低对杆状病毒敏感性的可能机制。
Several expressed sequence tags (ESTs) with homology to chitin deacetylase-like protein (CDA) were selected from a group of Helicoverpa armigera genes whose expression changed after infection with H. armigera single nucleopolyhedrovirus (HearNPV). Some of these ESTs coded for a midgut protein containing a chitin deacetylase domain (CDAD). The expressed protein, HaCDA5a, did not show chitin deacetylase activity, but it showed a strong affinity for binding to chitin. Sequence analysis showed the lack of any chitin binding domain, described for all currently known peritrophic membrane (PM) proteins. HaCDA5a has previously been detected in the H. armigera PM. Such localization, together with its downregulation after pathogen infection, led us to hypothesize that this protein might be responsible for the homeostasis of the PM structure and that, by reduction of its expression, the insect may reduce PM permeability, decreasing the entrance of baculovirus. To test this hypothesis, we constructed a recombinant nucleopolyhedrovirus to express HaCDA5a in insect cells and tested its influence on PM permeability as well as the influence of HaCDA5a expression on the performance of the baculovirus. The experiments showed that HaCDA5a increased PM permeability, in a concentration-dependent manner. Bioassays on Spodoptera frugiperda and Spodoptera exigua larvae revealed that NPV expressing HaCDA5a was more infective than its parental virus. However, no difference in virulence was observed when the viruses were injected intrahemocoelically. These findings support the downregulation of a midgut-specific CDA-like protein as a possible mechanism used by H. armigera to reduce susceptibility to baculovirus by decreasing PM permeability.