摘要：
Entomopathogenic fungi have high potential for controlling insect pests, although the slow killing speed has blocked their widespread application. To increase the virulence of entomopathogenic fungi, genetic modification can be employed. Egf1.0 is an immunosuppressive protein encoded by polydnavirus, carried by parasitoid wasp Microplitis demolitor, which blocks the prophenoloxidase (PPO) activation response of host insects. In this study, we explored the feasibility of genetically modifying entomopathogenic fungi with increased virulence by expressing Egf1.0. In comparison with the wild-type parents, the median lethal concentration (LC50) of Beauveria bassiana expressing Egf1.0 against Helicoverpa armigera was reduced by 2.7-fold, and the median lethal time (LT50) was reduced by 22.8%. In vitro assay showed that recombinant Egf1.0 was able to inhibit the PPO activation response of H. armigera. In vivo assay revealed that the expression of Egf1.0 in B. bassiana caused a higher degree of suppression to PPO activation response of H. armigera. These assays suggested that the increased virulence of the transgenic fungi is due to the increased ability to suppress the host insect's immune response. Moreover, colony growth, conidia yield, and germination assays revealed that the expression of Egf1.0 in B. bassiana had no effect on its growth and development. In conclusion, the expression of Egf1.0 can significantly enhance the pathogenicity of B. bassiana against host insects.
摘要：
Upon entry into the hemocoel of host insects, entomopathogenic fungi switch to yeast-like hyphal bodies that are not recognized by host hemocytes and replicate extensively in the hemolymph. The mechanism by which hyphal bodies evade host cellular immunity is not well understood. This study compares Metarhizium rileyi conidia and hyphal bodies with respect to elicitation of the immune response of Helicoverpa armigera and recognition by host pattern recognition receptors (PRRs). We found that the ability of host hemocytes to phagocytize and nodulate hyphal bodies was weaker than those responses against conidia, suggesting that hyphal bodies are more able to evade host cellular immunity. Additionally, we found that the binding affinity of H. armigera β-1,3-glucan recognition proteins was much lower for hyphal bodies than for conidia. We observed no agglutination response of H. armigera C-type lectin 3 (HaCTL3) against hyphal bodies, and HaCTL3 bound significantly less to hyphal bodies than to conidia, indicating that host PRRs have a lower affinity for hyphal bodies than for conidia. This study provides direct evidence that the mechanism whereby entomopathogenic fungi escape host cellular immunity involves the inability of host PRRs to sufficiently recognize hyphal bodies to elicit the cellular immune response.
Hyphal bodies had a greater ability to evade the host cellular immunity.
Hyphal bodies evade PRR recognition.
摘要：
An important innate immune response in Drosophila melanogaster is the production of antimicrobial peptides (AMPs). Expression of AMP genes is mediated by the Toll and immune deficiency (IMD) pathways via NF-kappaB transcription factors Dorsal, DIF and Relish. Dorsal and DIF act downstream of the Toll pathway, whereas Relish acts in the IMD pathway. Dorsal and DIF are held inactive in the cytoplasm by the IkappaB protein Cactus, while Relish contains an IkappaB-like inhibitory domain at the C-terminus. NF-kappaB factors normally form homodimers and heterodimers to regulate gene expression, but formation of heterodimers between Relish and DIF or Dorsal and the specificity and activity of the three NF-kappaB homodimers and heterodimers are not well understood. In this study, we compared the activity of Rel homology domains (RHDs) of Dorsal, DIF and Relish in activation of Drosophila AMP gene promoters, demonstrated that Relish-RHD (Rel-RHD) interacted with both Dorsal-RHD and DIF-RHD, Relish-N interacted with DIF and Dorsal, and overexpression of individual RHD and co-expression of any two RHDs activated the activity of AMP gene promoters to various levels, suggesting formation of homodimers and heterodimers among Dorsal, DIF and Relish. Rel-RHD homodimers were stronger activators than heterodimers of Rel-RHD with either DIF-RHD or Dorsal-RHD, while DIF-RHD-Dorsal-RHD heterodimers were stronger activators than either DIF-RHD or Dorsal-RHD homodimers in activation of AMP gene promoters. We also identified the nucleotides at the 6th and 8th positions of the 3' half-sites of the kappaB motifs that are important for the specificity and activity of NF-kappaB transcription factors.
期刊：
Journal of Biological Chemistry,2019年294(26):10172-10181 ISSN：0021-9258
通讯作者：
Yu, Xiao-Qiang;Strand, Michael R.
作者机构：
[He, Zhen; Yu, Xiao-Qiang; Li, Chun-Feng; Chowdhury, Munmun] Univ Missouri, Sch Biol Sci, Div Cell Biol & Biophys, Kansas City, MO 64110 USA.;[Liu, Xu-Sheng; He, Zhen; Yu, Xiao-Qiang; Wang, Yu-Feng] Cent China Normal Univ, Sch Life Sci, Wuhan 430079, Hubei, Peoples R China.;[Yu, Xiao-Qiang; Lu, Yuzhen] South China Normal Univ, Guangzhou Key Lab Insect Dev Regulat & Applicat R, Inst Insect Sci & Technol, Guangzhou 510631, Guangdong, Peoples R China.;[Yu, Xiao-Qiang; Lu, Yuzhen] South China Normal Univ, Sch Life Sci, Guangzhou 510631, Guangdong, Peoples R China.;[Li, Chun-Feng] Southwest Univ, State Key Lab Silkworm Genome Biol, Chongqing 400716, Peoples R China.
通讯机构：
[Yu, Xiao-Qiang; Strand, Michael R.] U;Univ Missouri, Sch Biol Sci, Div Cell Biol & Biophys, Kansas City, MO 64110 USA.;Univ Georgia, Dept Entomol, Athens, GA 30602 USA.
摘要：
[目的]前期从棉铃虫血细胞转录组中鉴定到一个免疫相关基因，命名为defense protein 1(DFP1)。本文研究了DFP1基因在棉铃虫先天免疫中的具体功能。[方法]利用qRT-PCR、重组蛋白体外功能分析、Western blotting、RNAi等方法研究了DFP1基因在棉铃虫血细胞包囊作用中的作用方式。
作者机构：
[Liu, Xu-Sheng; Zhou, Jing; Zhang, Ning-Zhao; Liu, Kai-Yu; Ai, Hui; Xia, Yu-Qian; Chen, Zu-Wen; Lu, Dandan] Cent China Normal Univ, Sch Life Sci, Hubei Key Lab Genet Regulat & Integrat Biol, Wuhan 430079, Peoples R China.;[Zhou, Li-Lin] Wuhan Vegetable Res Inst, Dept Plant Protect, Wuhan, Peoples R China.
通讯机构：
[Ai, Hui] C;Cent China Normal Univ, Sch Life Sci, Hubei Key Lab Genet Regulat & Integrat Biol, Wuhan 430079, Peoples R China.
关键词：
Spodoptera exigua;ATG5;autophagy;qPCR;Western blot
摘要：
Autophagy is not only involved in development, but also has been proved to attend immune response against invading pathogens. Autophagy protein 5 (ATG5) is an important autophagic protein, which plays a crucial role in autophagosome elongation. Although ATG5 has been well studied in mammal, yeast, and Drosophila, little is known about ATG5 in lepidopteran insects. We cloned putative SeAtg5 gene from Spodoptera exigua larvae by the rapid amplification of cDNA ends method, and its characteristics and the influences of multiple exogenous factors on its expression levels were then investigated. The results showed that the putative S. exigua SeATG5 protein is highly homologous to other insect ATG5 proteins, which has a conserved Pfm domain and multiple phosphorylation sites. Next, fluorescence microscope observation showed that mCherry-SeATG5 was distributed in both nucleus and cytoplasm of Spodoptera litura Sl-HP cells and partially co-localized with BmATG6-GFP, but it almost has no significant co-localization with GFP-HaATG8. Then, the Western blot analysis demonstrated that GFP-SeATG5 conjugated with ATG12. Moreover, real-time PCR revealed that its expression levels significantly increased at the initiation of pupation and the stage of adult. In addition, the expression levels of SeAtg5 can be enhanced by the starvation, UV radiation, and infection of baculovirus and bacterium. However, the expression levels of SeAtg5 decreased at 24 h post treatments in all these treatments except in starvation. These results suggested that SeATG5 might be involved in response of S. exigua under various stress conditions.
摘要：
Rab3, a member of the Rab GTPase family, has been found. to be involved in Innate immunity. However, the precise function of this GTPase in innate immunity remains unknown. In this study, we identified a Rab3 gene (Ha-Rab3) from the cotton bollworm, Helicoverpa armigera and studied its roles in innate immune responses. Expression of Ha-Rab3 was upregulated in the hemocytes of H. armigera larvae after the injection of Escherichia coil or chromatography beads. The dsRNA-mediated knockdown of Ha-Rab3 gene in H. armigera larval hemocytes led to significant reduction in the phagocytosis and nodulation activities of hemocytes against E. coil, significant increase in the bacterial load in larval hemolymph, and significant reduction in the encapsulation activities of hemocytes toward invading chromatography beads. Further-more, Ha-Rab3 knockdown significantly suppressed spreading of plasmatocytes. These results' suggest that Ha-Rab3 plays important roles in H. arrnigera cellular immune responses, possibly by mediating spreading of hemocytes. (C) 2015 Elsevier Ltd. All rights reserved.
