SignificanceThe RNA interference (RNAi) discovered in nematodes has contributed to major advances in basic and applied sciences. RNAi-based methods are being developed for controlling pests and disease vectors. RNAi is highly efficient and systemic in coleopteran insects, but not in other insects. The lower efficiency of RNAi in economically important insects and concerns about resistance development are hindering the widespread use of this technology. To address these problems, a RNAi-sensitive Colorado potato beetle, Leptinotarsa decemlineata, and a cell line derived from this insect were used to identify a dsRNA-binding protein, StaufenC, as a major contributor to RNAi and its resistance. Interestingly, StaufenC homologs are present in only coleopteran insects and are essential for efficient RNAi response and its resistance in these insects.

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We identify and characterize 14 small heat-shock protein (sHSP) genes from the diamondback moth (DBM), Plutella xylostella (L.), a destructive pest. Phylogenetic analyses indicate that, except for sHSP18.8 and sHSP19.22, the other 12 DBM sHSPs belong to five known insect sHSP groups. Developmental expression analysis revealed that most sHSPs peaked in the pupal and adult stages. The transcripts of sHSPs display tissue specificity with two exhibiting constitutive expression in four tested tissues. Expression of sHSP18.8 in fourth instar larvae is not induced by the tested abiotic stressors, and unless sHSP21.8 is not sensitive to thermal stress, 12 sHSPs are significantly up-regulated. The messenger RNA (mRNA) levels of all sHSPs are reduced under oxidative stress. Food deprivation leads to significant down-regulation of three sHSPs. The majority of sHSPs show expression variation to various heavy metals, whereas mRNA abundances of sHSP22.1 and sHSP 28.9 are reduced by four heavy metals. The responses of sHSPs to indoxacarb and cantharidin are varied. Beta-cypermethrin and chlorfenapyr exposure results in an increase of 13 sHSP transcripts and a reduction of 12 sHSP transcripts, respectively. These results show that different sHSPs might play distinct roles in the development and regulation of physiological activities, as well as in response to various abiotic stresses of DBM.

Glutathione S-transferases (GSTs) are important detoxification enzymes involved in the development of metabolic resistance in Plutella xylostella. Uncovering the interactions between representative PxGSTs and the inhibitor S-hexyl glutathione (GTX), helps in the development of effective PxGST inhibitors for resistance management. As the PxGST most severely inhibited by GTX, PxGSTσ (sigma-class PxGST) adopts the canonical fold of insect GSTs. The formation of the PxGSTσ-GTX complex is mainly driven by H-bond and hydrophobic interactions derived from the side chains of favorable residues. Of the residues composing the active site of PxGSTσ, Lys43 and Arg99 are two hot spots, first reported in the binding of GSH derivatives to GSTs. Such differences indicate the metabolism discrimination of different insect GSTs. Unfavorable interactions between the PxGSTσ active site and GTX are depicted as well. The research guides the discovery and optimization of PxGSTσ inhibitors.

Sex determination has been studied in the model lepidopteran species Bombyx mori, but it remains poorly understood in lepidopteran pests. In the present study, we identified and characterized the Masculinizer (Masc) gene in a Noctuidae pest species, Agrotis ipsilon. Sequence analysis revealed that AiMasc encodes a protein of 658 amino acids that has two CCCH-type zinc finger domains and two conserved cysteine residues (Cys-277 and Cys-280). We assessed the masculinizing activity of AiMasc in BmN cells and found that AiMasc induced expression of the male-specific doublesex isoform. Disruption of Masc via clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated protein 9 (Cas9) in A. ipsilon caused abnormalities in abdominal segments and external genitalia, resulting in male-specific sterility. These results suggest that Masc participates in the process of sex determination in A. ipsilon. Successful identification of sex-determination gene in a pest species may enable the development of novel genetic approaches for pest control.

Serine/threonine protein phosphatase 5 (PP5) is a promising novel target for anticancer therapies. This work aims to uncover the key interactions at the atomic level between PP5 and three inhibitors (cantharidin, norcantharidin and endothall). We found that, unlike previous report, Arg 100 contributes less to PP5-inhibitor binding, and the residues His 69, Asn 128, His 129, Arg 225, His 252 and Arg 250 are of importance to PP5-inhibitor binding. The hydrophobic interactions established between the residues Val 254, Phe 271 and Tyr 276, especially Glu 253, are very important to enhance the inhibitive interaction. We suggested that, to increase the inhibitory activity, the interactions of inhibitor with three negatively charged unfavorable interaction residues, Asp 99, Glu 130 and Asp 213, should be avoided. However, the interactions of inhibitor with favorable interaction residue Arg 250 could enhance the inhibitory activity. The Manganese ion 2 (MN2) unfavorably contribute to the total interaction free energies. The coordination between MN2 and chemical group of inhibitor should be eliminated. This work provides insight into how cantharidin and its analogs bind to PP5c at the atomic level and will facilitate modification of cantharidin-like chemicals to rationally develop more specific and less cytotoxic anti-cancer drugs.

The changes in DBM GST gene expression levels exposed to different insecticides indicate that they may play individual roles in tolerance to insecticides and xenobiotics.