摘要
C型凝集素(C-type lectins,CTLs)是广泛存在于昆虫体内一类含糖识别结构域的蛋白超家族,通常作为模式识别受体,在识别外来病原微生物并启动昆虫先天免疫过程中发挥重要功能。本研究克隆并鉴定了一条褐飞虱CTL编码基因NlCTL4(GenBank登录号:OQ032531),其c DNA序列全长936 bp,编码311个氨基酸。序列分析显示,Nl CTL4蛋白N端存在一段由21个氨基酸残基组成的信号肽,C端仅具有一个典型的糖识别结构域,属于S型CTLs。系统发育分析表明,Nl CTL4与半翅目其他昆虫的CTLs聚为一支,其中与玻璃翅叶蝉CTL亲缘关系最近。q RT-PCR分析显示,Nl CTL4基因在褐飞虱不同发育阶段和不同组织中均有表达。Nl CTL4在5龄若虫期的表达量最高,且在高龄若虫期(4~5龄)的表达量显著高于低龄若虫(1~3龄),但在雌、雄成虫中其表达量无显著差异。Nl CTL4在褐飞虱雌成虫肠道中表达量最高,脂肪体次之,在血淋巴中表达量最低。大肠杆菌(革兰氏阴性细菌)、金黄色葡萄球菌(革兰氏阳性细菌)和金龟子绿僵菌(丝状病原真菌)均能显著诱导Nl CTL4的表达水平,其中大肠杆菌和金黄色葡萄球菌在诱导36 h时Nl CTL4的表达量达到顶峰,金龟子绿僵菌接种24 h时的诱导水平最高。RNAi分析结果显示,抑制Nl CTL4表达后褐飞虱5龄若虫存活率及对金龟子绿僵菌侵染的抵御能力显著降低。本研究结果初步证实了褐飞虱Nl CTL4在宿主生长发育和免疫防御过程中发挥重要调控作用,且在开发褐飞虱RNAi抗虫和病原真菌防虫技术中具有重要的潜在应用价值。
C-type lectins(CTLs)are a protein superfamily containing one or more carbohydrate-recognition domains that widely exist in insects and often serve as pattern recognition receptors(PRRs),which play important roles in recognition of invading pathogenic microorganisms and the initiation of insect innate immunity.In this study,a CTL-encoding gene NlCTL4(GenBank accession no.:OQ032531)was successfully cloned and identified from the brown planthopper(BPH),Nilaparvata lugens.The full length cDNA of NlCTL4 is 936 bp,encoding 311 amino acids.Sequence analysis revealed that NlCTL4 protein contains a predicted signal peptide consisting of 21 amino acid residues at the N-terminus and belongs to CTL-S group as only one typical carbohydrate-recognition domain is included at the C-terminal region.Phylogenetic analysis showed that NlCTL4 clustered together with other hemipteran CTLs,with the highest homology with the CTL of Homalodisca vitripennis.The qRT-PCR results showed that NlCTL4 was expressed in all tested developmental stages and tissues.The highest expression level was observed in the 5th instar nymphs.The transcript level of NlCSP4 in older nymphs(4th and 5th instars)was significantly higher than those in young nymphs(1st,2nd and 3rd instars),but no significant difference in expression was observed between male and female adults.Among the different tissues of female adults,the highest transcript level of NlCTL4 was observed in the gut,followed by the fat body,and the lowest in the hemolymph.The expression of NlCTL4 in BPH was significantly up-regulated after the insects were infected with each of the Gram-negative bacteria Escherichia coli,Gram-positive bacteria Staphylococcus aureus,and the entomopathogenic fungus Metarhizium anisopliae.The expression of NlCTL4 reached the peak at 36 h post injection with E.coli or S.aureus,while the highest induction level was observed at 24 h after infection with M.anisopliae.RNAi results showed that suppression of NlCTL4 resulted in significant decreases in both the survival rate a
作者
王彦丹
邵珠龙
王正亮
俞晓平
WANG Yandan;SHAO Zhulong;WANG Zhengliang;YU Xiaoping(Zhejiang Provincial Key Laboratory of Biometrology and Inspection and Quarantine/College of Life Sciences,China Jiliang University,Hangzhou 310018,China)
出处
《中国生物防治学报》
CSCD
北大核心
2024年第2期337-346,共10页
Chinese Journal of Biological Control
基金
国家自然科学基金(31972347,U21A20223)
浙江省属高校基本科研业务费专项(2022YW82)。
关键词
褐飞虱
C型凝集素
基因克隆
表达模式
RNA干扰
Nilaparvata lugens
C-type lectins
gene cloning
expression profile
RNA interference
