Nuclear receptors potentially regulate phytochemical detoxification in Spodoptera litura.

Phytochemicals are a class of potential pesticides for pest control. Our previous studies have demonstrated that the development of Spodoptera litura is suppressed by two phytochemicals, flavone and xanthotoxin. Generally, phytochemical is metabolized by insect detoxification enzyme systems. Nuclear receptor (NR) is the ligand-activated transcription factor that involved in the regulation of detoxification gene expressions. To explore how NR responds to phytochemical to mediate detoxification gene expression, in the present study, 19 NRs were firstly identified in S. litura genome. The transcriptional levels of most NRs were significantly induced in the midgut of S. litura larvae after exposure to flavone and xanthotoxin. RNAi-mediated knockdown of FTZF1, EcR, Dsf, and HR3 remarkably reduced the larval tolerance to flavone or xanthotoxin. In addition, many crucial detoxification genes were downregulated by dsNR administrations, which might be responsible for the high sensitivity of S. litura to phytochemicals. Molecular docking indicated that phytochemicals as the potential ligands had high affinity to bind to NRs. This study suggested that NR potentially regulated the transcriptional expression of detoxification genes in response to phytochemical stresses, which partially elucidated the mechanism of extensive host adaptation in S. litura and provided the theoretical evidences for the development of NR-targeted insecticides.

Role of the epsilon glutathione S-transferases in xanthotoxin tolerance in Spodoptera litura.

Spodoptera litura, a polyphagous lepidopteran pest, demonstrates a remarkable capacity to adapt to varying host plants by efficiently detoxifying phytochemicals. However, the underlying mechanism for this adaptation is not well understood. Herein, twenty eplison glutathione S-transferase genes (GSTes) were characterized and their roles in phytochemical tolerance were analyzed in S. litura. Most of the GSTe genes were mainly expressed in the larval midgut and fat body. Exposure to the phytochemicals, especially xanthotoxin, induced the expression of most GSTe genes. Molecular docking analysis revealed that xanthotoxin could form stable bonds with six xanthotoxin-responsive GSTes, with binding free energies ranging from -36.44 to -68.83 kcal mol-1. Knockdown of these six GSTe genes increased the larval susceptibility to xanthotoxin. Furthermore, xanthotoxin exposure significantly upregulated the expression of two transcription factor genes CncC and MafK. Silencing of either CncC or MafK reduced the expression of GSTe16, which exhibited the largest change in response to xanthotoxin. Additionally, analysis of the promoter sequence of GSTe16 revealed the presence of seven CncC/Maf binding sites. Luciferase reporter assays showed that CncC and MafK enhanced the expression of GSTe16, leading to the increased xanthotoxin tolerance in S. litura. These findings provide insight into the functions and transcriptional regulatory mechanisms of GSTes, thereby enhancing our understanding of the role of GSTs in the adaptation of lepidopteran pests to phytochemicals.

Overexpression of peptidase inhibitor 16 attenuates angiotensin II-induced cardiac fibrosis via regulating HDAC1 of cardiac fibroblasts.

Cardiac hypertrophy and fibrosis are the major causes of heart failure due to non-ischaemia heart disease. To date, no specific therapy exists for cardiac fibrosis due to the largely unknown mechanisms of disease and lack of applicable therapeutic targets. In this study, we aimed to explore the role and associated mechanism of peptidase inhibitor 16 (PI16) in cardiac fibrosis induced by angiotensin II. In cardiac fibroblasts (CFs), overexpressed PI16 significantly inhibited CF proliferation and the levels of fibrosis-associated proteins. Further analysis of epigenetic changes in CF revealed that overexpressed PI16 decreases the nuclear level of histone deacetylase 1 (HDAC1) after angiotensin II treatment, resulting in increased histone 3 acetylation in K18 and K27 lysine. However, overexpression of HDAC1 by an adenovirus vector in CFs reversed these changes. Echocardiography showed that PI16 transgenic (Tg) mice have smaller left ventricle mass than wild-type mice. Histological analysis data showed that PI16 Tg mice demonstrated smaller cardiomyocyte size and less collagen deposition than wild-type mice. The effects of PI16 on HDAC1 and histone 3 were also confirmed in PI16 Tg mice using immunostaining. Generally, PI16 is a HDAC1 regulator specifically in CFs, and PI16 overexpression prevents cardiac hypertrophy and fibrosis by inhibiting stress-induced CF activation.

Nicotinamide deficiency promotes imidacloprid resistance via activation of ROS/CncC signaling pathway-mediated UGT detoxification in Nilaparvata lugens.

Metabolic alternation is a typical characteristic of insecticide resistance in insects. However, mechanisms underlying metabolic alternation and how altered metabolism in turn affects insecticide resistance are largely unknown. Here, we report that nicotinamide levels are decreased in the imidacloprid-resistant strain of Nilaparvata lugens, may due to reduced abundance of the symbiotic bacteria Arsenophonus. Importantly, the low levels of nicotinamide promote imidacloprid resistance via metabolic detoxification alternation, including elevations in UDP-glycosyltransferase enzymatic activity and enhancements in UGT386B2-mediated metabolism capability. Mechanistically, nicotinamide suppresses transcriptional regulatory activities of cap 'n' collar isoform C (CncC) and its partner small muscle aponeurosis fibromatosis isoform K (MafK) by scavenging the reactive oxygen species (ROS) and blocking the DNA binding domain of MafK. In imidacloprid-resistant N. lugens, nicotinamide deficiency re-activates the ROS/CncC signaling pathway to provoke UGT386B2 overexpression, thereby promoting imidacloprid detoxification. Thus, nicotinamide metabolism represents a promising target to counteract imidacloprid resistance in N. lugens.

Characterization and functional analysis of UDP-glycosyltransferases reveal their contribution to phytochemical flavone tolerance in Spodoptera litura.

Efficient detoxification is the key factor for phytophagous insect to adapt to phytochemicals. However, the role of uridine diphosphate (UDP)-glycosyltransferases (UGTs) in insect anti-defense to phytochemical flavone is largely unknown. In this study, 52 UGT genes were identified in Spodoptera litura and they presented evident gene duplication. UGT played a crucial part in larval tolerance to flavone because the enzyme activity and transcriptional level of 77 % UGT members were remarkably upregulated by flavone administration and suppression of UGT enzyme activity and gene expressions significantly increased larval susceptibility to flavone. Bacteria coexpressing UGTs had high survival rates under flavone treatment and flavone was dramatically metabolized by UGT recombinant cells, which indicated the involvement of UGTs in flavone detoxification. What's more, ecdysone pathway was activated by flavone. Topical application of 20-hydroxyecdysone highly upregulated UGT enzyme activity and more than half of UGT expressions. The effects were opposite when ecdysone receptor (EcR) and ultraspiracle (USP)-mediated ecdysone signaling pathway was inhibited. Furtherly, promoter reporter assays of 5 UGT genes showed that their transcription activities were notably increased by cotransfection with EcR and USP. In consequence, this study suggested that UGTs were involved in flavone detoxification and their transcriptional expressions were regulated by ecdysone pathway.

Characterization of Neuropeptides from Spodoptera litura and Functional Analysis of NPF in Diet Intake.

Neuropeptides are involved in many biological processes in insects. However, it is unclear what role neuropeptides play in Spodoptera litura adaptation to phytochemical flavone. In this study, 63 neuropeptide precursors from 48 gene families were identified in S. litura, including two neuropeptide F genes (NPFs). NPFs played a positive role in feeding regulation in S. litura because knockdown of NPFs decreased larval diet intake. S. litura larvae reduced flavone intake by downregulating NPFs. Conversely, the flavone intake was increased if the larvae were treated with NPF mature peptides. The NPF receptor (NPFR) was susceptible to the fluctuation of NPFs. NPFR mediated NPF signaling by interacting with NPFs to regulate the larval diet intake. In conclusion, this study suggested that NPF signaling regulated diet intake to promote S. litura adaptation to flavone, which contributed to understanding insect adaptation mechanisms to host plants and provide more potential pesticidal targets for pest control.

Whole-genome resequencing reveals genetic diversity and adaptive evolution in Chinese honeybee (Apis cerana cerana) in Guizhou, China.

Introduction: Guizhou Province, characterized by complex and diverse geographic and climatic environments, has rich genetic resources for the Chinese honeybee (Apis cerana cerana) and is one of the main bee-producing areas in China. However, research on the genetic diversity of Chinese honeybee in the Guizhou region is very limited, despite implications for conservation of biodiversity. Methods: In this study, we analyzed the genetic diversity, differentiation, and selection signals based on 116 Chinese honeybees from 12 regions in Guizhou Province using whole-genome sequencing. Results: We identified 1,400,430 high-quality SNPs across all samples. A population structure analysis revealed two independent genetic subgroups of Chinese honeybees in Guizhou, a Yunnan-Guizhou Plateau population in western Guizhou and a hilly-mountainous population in eastern Guizhou. The average nucleotide diversity (Pi) ranged from 0.00138 to 0.00161 and average expected heterozygosity (He) ranged from 0.2592 to 0.2604. The average genetic differentiation index (F ST) for Chinese honeybees in pairwise comparisons of 12 regions ranged from 0.0094 to 0.0293. There was clear genetic differentiation between the western plateau and the eastern hilly mountainous areas of Guizhou; however, F ST values between the eastern and western populations ranged from 0.0170 to 0.0293, indicating a low degree of differentiation. A genome-wide scan revealed a number of genes under selection in the Yunnan-Guizhou Plateau environment. These genes were related to growth and development, reproduction, and cold resistance, and several candidate genes involved in environmental adaptation were identified, including CTR, MAPK, MAST, HSF, and MKKK. Discussion: The results of the present study provide important theoretical bases for the conservation, evaluation, development, and utilization of genetic resources for Chinese honeybees in the Guizhou region and for further investigations of environmental adaptation and underlying mechanisms in the species.

CYP321A Subfamily P450s Contribute to the Detoxification of Phytochemicals and Pyrethroids in Spodoptera litura.

Although the induction of cytochrome P450 monooxygenases involved in insect detoxification has been well documented, the underlying regulatory mechanisms remain obscure. In Spodoptera litura, CYP321A subfamily members were effectively induced by exposure to flavone, xanthotoxin, curcumin, and λ-cyhalothrin, while knockdown of the CYP321A genes increased larval susceptibility to these xenobiotics. Homology modeling and molecular docking analyses showed that these four xenobiotics could stably bind to the CYP321A enzymes. Furthermore, two transcription factor genes, CncC and MafK, were significantly induced by the xenobiotics. Knockdown of CncC or MafK reduced the expression of four CYP321A genes and increased larval susceptibility to the xenobiotics. Dual-luciferase reporter assays showed that cotransfection of reporter plasmids carrying the CYP321A promoter with CncC and/or MafK-expressing constructs significantly magnified the promoter activity. These results indicate that the induction of CYP321A subfamily members conferring larval detoxification capability to xenobiotics is mediated by the activation of CncC and MafK.

SPARC-related modular calcium binding 1 regulates aortic valve calcification by disrupting BMPR-II/p-p38 signalling.

AIMS: Aortic valve calcification is more prevalent in chronic kidney disease accompanied by hypercalcemia. Secreted protein acidic and rich in cysteine (SPARC)-related modular calcium binding 1 (SMOC1) is a regulator of BMP2 signalling, but the role of SMOC1 in aortic valve calcification under different conditions has not been studied. This study aimed to investigate the roles of SMOC1 in aortic valve calcification under normal and high calcium conditions, focusing on the effects on aortic valve interstitial cells (AVICs). METHODS AND RESULTS: SMOC1 was expressed by aortic valve endothelial cells and secreted into the extracellular matrix in non-calcific valves and downregulated in calcific aortic valves. In vitro studies demonstrated that HUVEC secreted SMOC1 could enter the cytoplasm of AVICs. Overexpression of SMOC1 attenuated warfarin-induced AVIC calcification but promoted high calcium/phosphate or vitamin D-induced AVIC and aortic valve calcification by regulating BMP2 signalling both in vitro and in vivo. Co-immunoprecipitation revealed that SMOC1 binds to BMP receptor II (BMPR-II) and inhibits BMP2-induced phosphorylation of p38 (p-p38) via amino acids 372-383 of its EF-hand calcium-binding domain. Inhibition of p-p38 by the p38 inhibitor SB203580 blocked the effects of SMOC1 on BMP2 signalling and AVIC calcification induced by high calcium/phosphate medium. In high-calcium-treated AVICs, SMOC1 lost its ability to bind to BMPR-II, but not to caveolin-1, promoting p-p38 and cell apoptosis due to increased expression of BMPR-II and enhanced endocytosis. CONCLUSIONS: These observations support that SMOC1 works as a dual-directional modulator of AVIC calcification by regulating p38-dependent BMP2 signalling transduction according to different extracellular calcium concentrations.

Perirenal adipose afferent nerves sustain pathological high blood pressure in rats.

Hypertension is a pathological condition of persistent high blood pressure (BP) of which the underlying neural mechanisms remain obscure. Here, we show that the afferent nerves in perirenal adipose tissue (PRAT) contribute to maintain pathological high BP, without affecting physiological BP. Bilateral PRAT ablation or denervation leads to a long-term reduction of high BP in spontaneous hypertensive rats (SHR), but has no effect on normal BP in control rats. Further, gain- and loss-of-function and neuron transcriptomics studies show that augmented activities and remodeling of L1-L2 dorsal root ganglia neurons are responsible for hypertension in SHR. Moreover, we went on to show that calcitonin gene-related peptide (CGRP) is a key endogenous suppressor of hypertension that is sequestered by pro-hypertensive PRAT in SHRs. Taken together, we identify PRAT afferent nerves as a pro-hypertensive node that sustains high BP via suppressing CGRP, thereby providing a therapeutic target to tackle primary hypertension.