褐飞虱凝集素基因NlCTL4的克隆及功能分析OA北大核心CSTPCD

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 and the capability to resist M.anisopliae infection in the 5th instar nymphs of BPH.Our preliminary results confirmed that NlCTL4 plays key roles in host growth,development,and immune defense,and has great application potential for developing of RNAi-mediated and entomopathogenic fungi-based BPH biocontrol technology.