Search

Latrophilin (LPH) is an adhesion G protein-coupled receptor (aGPCR) that participates in multiple essential physiological processes. Our previous studies have shown that lph is not only indispensable for the development and reproduction of red flour beetles (Tribolium castaneum), but also for their resistance against dichlorvos or carbofuran insecticides. However, the regulatory mechanism of lph -mediated insecticide susceptibility remains unclear. Here, we revealed that knockdown of lph in beetles resulted in opposing changes in two chemoreception genes, chemosensory protein 10 (CSP10) and odorant - binding protein C01 (OBPC01), in which the expression of TcCSP10 was downregulated, whereas the expression of TcOBPC01 was upregulated. TcCSP10 and TcOBPC01 were expressed at the highest levels in early pupal and late larval stages, respectively. High levels of expression of both these genes were observed in the heads (without antennae) of adults. TcCSP10 and TcOBPC01 were significantly induced by dichlorvos or carbofuran between 12 and 72 h (hrs) after exposure, suggesting that they are likely associated with increasing the binding affinity of insecticides, leading to a decrease in sensitivity to the insecticides. Moreover, once these two genes were knocked down, the susceptibility of the beetles to dichlorvos or carbofuran was enhanced. Additionally, RNA interference (RNAi) targeting of lph followed by exposure to dichlorvos or carbofuran also caused the opposing expression levels of TcCSP10 and TcOBPC01 compared to the expression levels of wild-type larvae treated with insecticides alone. All these results indicate that lph is involved in insecticide susceptibility through positively regulating TcCSP10 ; and the susceptibility could also further partially compensated for through the negative regulation of TcOBPC01 when lph was knockdown in the red flour beetle. Our studies shed new light on the molecular regulatory mechanisms of lph related to insecticide susceptibility. lph was involved in insecticide susceptibility through positively regulating TcCSP10 , and it could further partially compensate for insecticide susceptibility by negatively regulating TcOBPC01 when lph was knockdown in Tribolium castaneum. Unlabelled Image • TcCSP10 was down-regulated and TcOBPC01 was up-regulated after knockdown of Tclph. • TcCSP10 and TcOBPC01 were significantly induced by insecticides of dichlorvos or carbofuran. • RNAi of TcCSP10 and TcOBPC01 increased the susceptibilities of larval to insecticides. • Tclph related to insecticide susceptibility via regulating TcCSP10 and partially compensated by TcOBPC01. [ABSTRACT FROM AUTHOR]

The zona pellucida domain protein Dusky (Dy) plays a vital role in wing morphogenesis in insects, but little information on its function has been reported. In this study, we found that dy regulated wing cell size, larval and pupal duration, and the metabolism of amino acid and 20‐hydroxyecdysone in Tribolium castaneum. Using RNA‐seq, 413 differentially expressed genes were identified between physiological buffer‐injected and dy‐double‐stranded RNA‐treated larvae, including 88 downregulated genes and 325 upregulated genes. Among these genes, dy knockdown increased CYP18A1 expression to elevate the 26‐hydroxylation of 20‐hydroxyecdysone, which ultimately led to growth defects in wing cells. Silencing of dy upregulated the transcription of genes encoding tyrosine aminotransferase, 4‐hydroxyphenylpyruvate dioxygenase, homogentisate 1, 2‐dioxygenase, and Pale to promote the catabolism of tyrosine and phenylalanine, which eventually reduced amino acid content. Furthermore, dy knockdown upregulated 4E‐BP expression, and 4E‐BP silencing partially phenocopied dy RNA interference‐mediated wing morphogenesis. These results suggest that Dy controls 20‐hydroxyecdysone and amino acid metabolism to regulate wing morphogenesis in the insect. [ABSTRACT FROM AUTHOR]

• Chat knockdown severely affected larval development and pupal eclosion, but vacht RNAi only disrupted pupal eclosion. • Parental RNAi of chat or vacht would reduce egg production and completely inhibite egg hatchability in T. castaneum. • Reduction of chat or vacht decreased the resistance to carbofuran and dichlorvos. Choline acetyltransferase (ChAT) and vesicular acetylcholine transporter (VAChT) are essential enzymes for synthesizing and transporting acetylcholine (ACh). But their functions in metamorphosis, reproduction, and the insecticide susceptibility were poorly understood in the insects. To address these issues, we identified the orthologues of chat and vacht in Tribolium castaneum. Spatiotemporal expression profiling showed Chat has the highest expression at the early adult stage, while vacht shows peak expression at the early larval stage. Both of them were highly expressed at the head of late adult. RNA interference (RNAi) of chat and vacht both led to a decrease in ACh content at the late larval stage. It is observed that chat knockdown severely affected larval development and pupal eclosion, but vacht RNAi only disrupted pupal eclosion. Further, parental RNAi of chat or vacht led to 35 % or 30 % reduction in fecundity, respectively, and knockdown of them completely inhibited egg hatchability. Further analysis has confirmed that both the reduction in fecundity and hatchability caused through the maternal specificity in T. castaneum. Moreover, the transcript levels of chat and vacht were elevated after carbofuran or dichlorvos treatment. Reduction of chat or vacht decreased the resistance to carbofuran and dichlorvos. This study provides the evidence for chat and vacht not only involved in development and reproduction of insects but also could as the potential targets of insecticides. [ABSTRACT FROM AUTHOR]

Myosin Myo20 plays vital roles in the morphogenesis of wings and legs among insects, but the function and signalling of Myo20 remain unclear. We show that Myo20 regulates wing cell division, ecdysteroid and amino acid metabolism, and gene expression in Tribolium castaneum. By RNA-seq, we identified 582 differentially expressed genes (DEGs) between control and ds- Myo20 larvae of T. castaneum. Of these DEGs, silencing Myo20 significantly decreased the mRNA and protein levels of lowfat. During development, lowfat has the highest expression in early pupae and the lowest level in 1-day embryos. Tissue-specific analysis indicated that lowfat was abundantly expressed in the head, fat body and epidermis of late-stage larvae and in wings and legs of 1, 2 and 5-day pupae. Likewise, knockdown of lowfat affected wing and leg morphogenesis, ecdysteroid and amino acid metabolism, and gene expression in T. castaneum. Silencing Myo20 or lowfat activated CYP18A1 to degrade ecdysteroids, stimulated amino acids catabolism to increase the transcription of 4E-BP but reduce S6K and cycE expression. These results suggest that Lowfat works downstream of Myo20 to employ target of rapamycin (TOR) signalling for wing and leg morphogenesis in insects. [Display omitted] • Myo20 regulates wing cell proliferation, ecdysteroid and amino acid metabolism, and gene expression. • Lowfat functions downstream of Myo20. • Myo20 and lowfat work upstream of TOR signalling to regulate wing morphogenesis. [ABSTRACT FROM AUTHOR]