Inducible Expression of dsRNA in Escherichia coli.

Double-stranded RNA (dsRNA) is a valuable tool for reverse genetics research and gene silencing applications. It is also an important management method for pests and diseases in agriculture. It can be synthesized both in vivo and in vitro. The latter presents the drawback of high production cost, the former is less expensive and suitable for scalable production. In general, dsRNAs are obtained in vivo from Escherichia coli heterologous systems that require the IPTG-inducible T7 RNA polymerase. In this report, we describe the construction of an RNAi system for the expression of dsRNA using the HT115 bacterial strain and the L4440 plasmid, and the extraction and identification of dsRNA.

LmFKBP24 interacts with LmEaster to inhibit the antifungal immunity of Locusta migratoria.

Locusta migratoria is one of the most destructive pests that threaten crop growth and food production security in China. Metarhizium anisopliae has been widely used to control locusts around the world. Previous laboratory studies have revealed that LmFKBP24 is significantly upregulated after M. anisopliae infection, suggesting that it may play a role in immune regulation, yet the mechanism remains largely unknown. To gain further insight, we conducted an RNA interference (RNAi) study to investigate the function of LmFKBP24 in the regulation of antifungal immunity and analyzed the expression patterns of immune-induced genes. Our research revealed that LmFKBP24 is activated and upregulated when locusts are infected by M. anisopliae, and it inhibits the expression of antimicrobial peptide (AMP) defensin in the downstream of Toll pathway by combining with LmEaster rather than LmCyPA, thus exerting an immunosuppressive effect. To further investigate this, we conducted yeast two-hybrid (Y2H) and pull down assays to identify the proteins interacting with LmFKBP24. Our results provided compelling evidence for revealing the immune mechanism of L. migratoria and uncovered an innovative target for the development of new biological pesticides. Furthermore, our research indicates that LmFKBP24 interacts with LmEaster through its intact structure, providing a strong foundation for further exploration.

Identification, characterization and spatiotemporal expression analysis of the FKBP family genes in Locusta migratoria.

FK506 binding proteins (FKBPs) are a highly-conserved group of proteins known to bind to FK506, an immunosuppressive drug. They play different physiological roles, including transcription regulation, protein folding, signal transduction and immunosuppression. A number of FKBP genes have been identified in eukaryotes; however, very little information about these genes has been reported in Locusta migratoria. Here, we identified and characterized 10 FKBP genes from L. migratoria. Phylogenetic analysis and comparison of domain architectures indicated that the LmFKBP family can be divided into two subfamilies and five subclasses. Developmental and tissue expression pattern analysis revealed that all LmFKBPs transcripts, including LmFKBP46, LmFKBP12, LmFKBP47, LmFKBP79, LmFKBP16, LmFKBP24, LmFKBP44b, LmFKBP53, were periodically expressed during different developmental stages and mainly expressed in the fat body, hemolymph, testis, and ovary. In brief, our work depicts a outline but panoramic picture of LmFKBP family in L. migratoria, and provides a solid foundation to further investigate the molecular functions of LmFKBPs.