Proton-driven sodium secretion in a saline water animal.

Aquatic animals residing in saline habitats either allow extracellular sodium concentration to conform to environmental values or regulate sodium to lower levels. The latter strategy requires an energy-driven process to move sodium against a large concentration gradient to eliminate excess sodium that diffuses into the animal. Previous studies of invertebrate and vertebrate species indicate a sodium pump, Na+/K+ ATPase, powers sodium secretion. We provide the first functional evidence of a saline-water animal, Aedes taeniorhynchus mosquito larva, utilizing a proton pump to power this process. Vacuolar-type H+ ATPase (VHA) protein is highly expressed on the apical membrane of the posterior rectal cells, and in situ sodium flux across this epithelium increases significantly in larvae held in higher salinity and is sensitive to Bafilomycin A1, an inhibitor of VHA. We also report the first evidence of splice variants of the sodium/proton exchanger, NHE3, with both high and low molecular weight variants highly expressed on the apical membrane of the posterior rectal cells. Evidence of NHE3 function was indicated with in situ sodium transport significantly inhibited by a NHE3 antagonist, S3226. We propose that the outward proton pumping by VHA establishes a favourable electromotive gradient to drive sodium secretion via NHE3 thus producing a hyperosmotic, sodium-rich urine. This H+- driven Na+ secretion process is the primary mechanism of ion regulation in salt-tolerant culicine mosquito species and was first investigated over 80 years ago.

Vitamin B6 deficiency cooperates with oncogenic Ras to induce malignant tumors in Drosophila.

Vitamin B6 is a water-soluble vitamin which possesses antioxidant properties. Its catalytically active form, pyridoxal 5'-phosphate (PLP), is a crucial cofactor for DNA and amino acid metabolism. The inverse correlation between vitamin B6 and cancer risk has been observed in several studies, although dietary vitamin B6 intake sometimes failed to confirm this association. However, the molecular link between vitamin B6 and cancer remains elusive. Previous work has shown that vitamin B6 deficiency causes chromosome aberrations (CABs) in Drosophila and human cells, suggesting that genome instability may correlate the lack of this vitamin to cancer. Here we provide evidence in support of this hypothesis. Firstly, we show that PLP deficiency, induced by the PLP antagonists 4-deoxypyridoxine (4DP) or ginkgotoxin (GT), promoted tumorigenesis in eye larval discs transforming benign RasV12 tumors into aggressive forms. In contrast, PLP supplementation reduced the development of tumors. We also show that low PLP levels, induced by 4DP or by silencing the sgllPNPO gene involved in PLP biosynthesis, worsened the tumor phenotype in another Drosophila cancer model generated by concomitantly activating RasV12 and downregulating Discs-large (Dlg) gene. Moreover, we found that RasV12 eye discs from larvae reared on 4DP displayed CABs, reactive oxygen species (ROS) and low catalytic activity of serine hydroxymethyltransferase (SHMT), a PLP-dependent enzyme involved in thymidylate (dTMP) biosynthesis, in turn required for DNA replication and repair. Feeding RasV12 4DP-fed larvae with PLP or ascorbic acid (AA) plus dTMP, rescued both CABs and tumors. The same effect was produced by overexpressing catalase in RasV12 DlgRNAi 4DP-fed larvae, thus allowing to establish a relationship between PLP deficiency, CABs, and cancer. Overall, our data provide the first in vivo demonstration that PLP deficiency can impact on cancer by increasing genome instability, which is in turn mediated by ROS and reduced dTMP levels.

Behavioral Assays Dissecting NMDA Receptor Function in Zebrafish.

Zebrafish are a powerful system to study brain development and to dissect the activity of complex circuits. One advantage is that they display complex behaviors, including prey capture, learning, responses to photic and acoustic stimuli, and social interaction (Dreosti et al., Front Neural Circuits 9:39, 2015; Bruckner et al., PLoS Biol 20:e3001838, 2022; Zoodsma et al., Mol Autism 13:38, 2022) that can be probed to assess brain function. Many of these behaviors are easily assayed at early larval stages, offering a noninvasive and high-throughput readout of nervous system function. Additionally, larval zebrafish readily uptake small molecules dissolved in water making them ideal for behavioral-based drug screens. Together, larval zebrafish and their behavioral repertoire offer a means to rapidly dissect brain circuitry and can serve as a template for high-throughput small molecule screens.NMDA receptor subunits are highly conserved in zebrafish compared to mammals (Zoodsma et al., Mol Autism 13:38, 2022; Cox et al., Dev Dyn 234:756-766, 2005; Zoodsma et al., J Neurosci 40:3631-3645, 2020). High amino acid and domain structure homology between humans and zebrafish underlie conserved functional similarities. Here we describe a set of behavioral assays that are useful to study the NMDA receptor activity in brain function.

Hemolymph metabolism of black soldier fly (Diptera: Stratiomyidae), response to different supplemental fungi.

The black soldier fly, Hermetia illucens L. (Diptera: Stratiomyidae), is commonly used for organic waste recycling and animal feed production. However, the often inadequate nutrients in organic waste necessitate nutritional enhancement of black soldier fly larvae, e.g., by fungal supplementation of its diet. We investigated the amino acid composition of two fungi, Candida tropicalis (Castell.) Berkhout (Saccharomycetales: Saccharomycetaceae) and Pichia kudriavzevii Boidin, Pignal & Besson (Saccharomycetales: Pichiaceae), from the black soldier fly gut, and commercial baker's yeast, Saccharomyces cerevisiae Meyen ex E.C. Hansen (Saccharomycetales: Saccharomycetaceae), and their effects on larval growth and hemolymph metabolites in fifth-instar black soldier fly larvae. Liquid chromatography-mass spectrometry was used to study the effect of fungal metabolites on black soldier fly larval metabolism. Amino acid analysis revealed significant variation among the fungi. Fungal supplementation led to increased larval body mass and differential metabolite accumulation. The three fungal species caused distinct metabolic changes, with each over-accumulating and down-accumulating various metabolites. We identified significant alteration of histidine metabolism, aminoacyl-tRNA biosynthesis, and glycerophospholipid metabolism in BSF larvae treated with C. tropicalis. Treatment with P. kudriavzevii affected histidine metabolism and citrate cycle metabolites, while both P. kudriavzevii and S. cerevisiae treatments impacted tyrosine metabolism. Treatment with S. cerevisiae resulted in down-accumulation of metabolites related to glycine, serine, and threonine metabolism. This study suggests that adding fungi to the larval diet significantly affects black soldier fly larval metabolomics. Further research is needed to understand how individual amino acids and their metabolites contributed by fungi affect black soldier fly larval physiology, growth, and development, to elucidate the interaction between fungal nutrients and black soldier fly physiology.

Glucose and trehalose metabolism through the cyclic pentose phosphate pathway shapes pathogen resistance and host protection in Drosophila.

Activation of immune cells requires the remodeling of cell metabolism in order to support immune function. We study these metabolic changes through the infection of Drosophila larvae by parasitoid wasp. The parasitoid egg is neutralized by differentiating lamellocytes, which encapsulate the egg. A melanization cascade is initiated, producing toxic molecules to destroy the egg while the capsule also protects the host from the toxic reaction. We combined transcriptomics and metabolomics, including 13C-labeled glucose and trehalose tracing, as well as genetic manipulation of sugar metabolism to study changes in metabolism, specifically in Drosophila hemocytes. We found that hemocytes increase the expression of several carbohydrate transporters and accordingly uptake more sugar during infection. These carbohydrates are metabolized by increased glycolysis, associated with lactate production, and cyclic pentose phosphate pathway (PPP), in which glucose-6-phosphate is re-oxidized to maximize NADPH yield. Oxidative PPP is required for lamellocyte differentiation and resistance, as is systemic trehalose metabolism. In addition, fully differentiated lamellocytes use a cytoplasmic form of trehalase to cleave trehalose to glucose and fuel cyclic PPP. Intracellular trehalose metabolism is not required for lamellocyte differentiation, but its down-regulation elevates levels of reactive oxygen species, associated with increased resistance and reduced fitness. Our results suggest that sugar metabolism, and specifically cyclic PPP, within immune cells is important not only to fight infection but also to protect the host from its own immune response and for ensuring fitness of the survivor.

The zona pellucida protein piopio regulates the metamorphosis and reproduction in Tribolium castaneum.

The zona pellucida domain protein piopio (Pio) was only reported to mediate the adhesion of the apical epithelial surface and the overlying apical extracellular matrix in Drosophila melanogaster, but the developmental roles of Pio were poorly understood in insects. To address this issue, we comprehensively analyzed the function of Pio in Tribolium castaneum. Phylogenetic analysis indicated that pio exhibited one-to-one orthologous relationship among insects. T. castaneum pio had a 1236-bp ORF and contained eight exons. During development pio was abundantly expressed from larva to adult and lowly expressed at the late stage of embryo and adult, while it had more transcripts in the head, epidermis, and gut but fewer in the fat body of late-stage larvae. Knockdown of pio inhibited the pupation, eclosion, and reproduction of T. castaneum. The expression of vitellogenin 1 (Vg1), Vg2, and Vg receptor (VgR) largely decreased in pio-silenced female adults. Silencing pio increased the 20-hydroxyecdysone titer by upregulating phm and spo expression but decreased the juvenile hormone (JH) titer through downregulating JHAMT3 and promoting JHE, JHEH-r4, and JHDK transcription. These results suggested that Pio might regulate the metamorphosis and reproduction via modulating the ecdysone and JH metabolism in T. castaneum. This study found the novel roles of pio in insect metamorphosis and reproduction, and provided the new insights for analyzing other zona pellucida proteins functions in insects.

Effects of chironomid larvae density and mosquito biocide on methane and carbon dioxide dynamics in freshwater sediments.

Small lentic water bodies are important emitters of methane (CH4) and carbon dioxide (CO2), but the processes regulating their dynamics and susceptibility to human-induced stressors are not fully understood. Bioturbation by chironomid larvae has been proposed as a potentially important factor controlling the dynamics of both gases in aquatic sediments. Chironomid abundance can be affected by the application of biocides for mosquito control, such as Bti (Bacillus thuringiensis var. israelensis). Previous research has attributed increases in CH4 and CO2 emissions after Bti application to reduced bioturbation by chironomids. In this study, we separately tested the effect of chironomid bioturbation and Bti addition on CH4 production and emission from natural sediments. In a set of 15 microcosms, we compared CH4 and CO2 emission and production rates with high and low densities of chironomid larvae at the bioturbating stage, and standard and five times (5x) standard Bti dose, with control sediments that contained neither chironomid larvae nor Bti. Regardless of larvae density, chironomid larvae did not affect CH4 nor CO2 emission and production of the sediment, although both rates were more variable in the treatments with organisms. 5xBti dosage, however, led to a more than three-fold increase in CH4 and CO2 production rates, likely stimulated by bioavailable dissolved carbon in the Bti excipient and priming effects. Our results suggest weak effects of bioturbating chironomid larvae on the CH4 and CO2 dynamics in aquatic ecosystems. Furthermore, our results point out towards potential functional implications of Bti for carbon cycling beyond those mediated by changes in the macroinvertebrate community.

Metabolism and cytotoxicity studies of the two hallucinogens 1cP-LSD and 4-AcO-DET in human liver and zebrafish larvae models using LC-HRMS/MS and a high-content screening assay.

The continuous emergence of new psychoactive substances (NPS) attracted a great deal of attention within recent years. Lately, the two hallucinogenic NPS 1cP-LSD and 4-AcO-DET have appeared on the global market. Knowledge about their metabolism to identify potential metabolic targets for analysis and their cytotoxic properties is lacking. The aim of this work was thus to study their in vitro and in vivo metabolism in pooled human liver S9 fraction (pHLS9) and in zebrafish larvae (ZL) by means of liquid chromatography-high-resolution tandem mass spectrometry. Monooxygenases involved in the initial metabolic steps were elucidated using recombinant human isozymes. Investigations on their cytotoxicity were performed on the human hepatoma cell line HepG2 using a multiparametric, fluorescence-based high-content screening assay. This included measurement of CYP-enzyme mediated effects by means of the unspecific CYP inhibitor 1-aminbenzotriazole (ABT). Several phase I metabolites of both compounds and two phase II metabolites of 4-AcO-DET were produced in vitro and in vivo. After microinjection of 1cP-LSD into the caudal vein of ZL, three out of seven metabolites formed in pHLS9 were also detected in ZL. Twelve 4-AcO-DET metabolites were identified in ZL after exposure via immersion bath and five of them were found in pHLS9 incubations. Notably, unique metabolites of 4-AcO-DET were only produced by ZL, whereas 1cP-LSD specific metabolites were found both in ZL and in pHLS9. No toxic effects were observed for 1cP-LSD and 4-AcO-DET in HepG2 cells, however, two parameters were altered in incubations containing 4-AcO-DET together with ABT compared with incubations without ABT but in concentrations far above expected in vivo concentration. Further investigations should be done with other hepatic cell lines expressing higher levels of CYP enzymes.

Deciphering the function of the fifth class of Gα proteins: regulation of ionic homeostasis as unifying hypothesis.

Trimeric G proteins transduce signals from a superfamily of receptors and each G protein controls a wide range of cellular and systemic functions. Their highly conserved alpha subunits fall in five classes, four of which have been well investigated (Gs, Gi, G12, Gq). In contrast, the function of the fifth class, Gv is completely unknown, despite its broad occurrence and evolutionary ancient origin (older than metazoans). Here we show a dynamic presence of Gv mRNA in several organs during early development of zebrafish, including the hatching gland, the pronephros and several cartilage anlagen, employing in situ hybridisation. Next, we generated a Gv frameshift mutation in zebrafish and observed distinct phenotypes such as reduced oviposition, premature hatching and craniofacial abnormalities in bone and cartilage of larval zebrafish. These phenotypes could suggest a disturbance in ionic homeostasis as a common denominator. Indeed, we find reduced levels of calcium, magnesium and potassium in the larvae and changes in expression levels of the sodium potassium pump atp1a1a.5 and the sodium/calcium exchanger ncx1b in larvae and in the adult kidney, a major osmoregulatory organ. Additionally, expression of sodium chloride cotransporter slc12a3 and the anion exchanger slc26a4 is altered in complementary ways in adult kidney. It appears that Gv may modulate ionic homeostasis in zebrafish during development and in adults. Our results constitute the first insight into the function of the fifth class of G alpha proteins.

Biochemical characterization of Bombyx mori Dicer-2 that dices double-stranded RNAs into 20-nt small RNA.

We detected enzymatic activity that generates 20-nucleotide (nt) RNA from double-stranded RNAs (dsRNAs) in crude extracts prepared from various silkworm (Bombyx mori) organs. The result using knocked-down cultured cells indicated that this dicing activity originated from B. mori Dicer-2 (BmDcr2). Biochemical analyses revealed that BmDcr2 preferentially cleaves 5'-phosphorylated dsRNAs at the 20-nt site-counted from the 5'-phosphorylated end-and required ATP and magnesium ions for the dicing reaction. This is the first report of the biochemical characterization of Dicer-2 in lepidopteran insects. This enzymatic property of BmDcr2 in vitro is consistent with the in vivo small interfering RNA profile in virus-infected silkworm cells.

Phytomediated stress modulates antioxidant status, induces overexpression of CYP6M2, Hsp70, α-esterase, and suppresses the ABC transporter in Anopheles gambiae (sensu stricto) exposed to Ocimum tenuiflorum extracts.

The incorporation of phytoactive compounds in the management of malarial vectors holds promise for the development of innovative and efficient alternatives. Nevertheless, the molecular and physiological responses that these bioactive substances induce remain underexplored. This present study investigated the toxicity of different concentrations of aqueous and methanol extracts of Ocimum tenuiflorum against larvae of Anopheles gambiae (sensu stricto) and unraveled the possible underlying molecular pathways responsible for the observed physiological effects. FTIR and GCMS analyses of phytoactive compounds in aqueous and methanol crude extracts of O. tenuiflorum showed the presence of OH stretching vibration, C = C stretching modes of aromatics and methylene rocking vibration; ring deformation mode with high levels of trans-ß-ocimene, 3,7-dimethyl-1,3,6-octatriene in aqueous extract and 4-methoxy-benzaldehyde, 1,3,5-trimethyl-cyclohexane and o-cymene in methanol extract. The percentage mortality upon exposure to methanol and aqueous extracts of O. tenuiflorum were 21.1% and 26.1% at 24 h, 27.8% and 36.1% at 48 h and 36.1% and 45% at 72 h respectively. Using reverse transcription quantitative polymerase chain reaction (RT-qPCR), down-regulation of ABC transporter, overexpression of CYP6M2, Hsp70, and α-esterase, coupled with significantly increased levels of SOD, CAT, and GSH, were observed in An. gambiae (s.s.) exposed to aqueous and methanol extracts of O. tenuiflorum as compared to the control. Findings from this study have significant implications for our understanding of how An. gambiae (s.s.) larvae detoxify phytoactive compounds.

A novel adipose loss-of-function mutant in Drosophila.

To identify genes required for brain growth, we took an RNAi knockdown reverse genetic approach in Drosophila. One potential candidate isolated from this effort is the anti-lipogenic gene adipose (adp). Adp has an established role in the negative regulation of lipogenesis in the fat body of the fly and adipose tissue in mammals. While fat is key to proper development in general, adp has not been investigated during brain development. Here, we found that RNAi knockdown of adp in neuronal stem cells and neurons results in reduced brain lobe volume and sought to replicate this with a mutant fly. We generated a novel adp mutant that acts as a loss-of-function mutant based on buoyancy assay results. We found that despite a change in fat content in the body overall and a decrease in the number of larger (>5 µm) brain lipid droplets, there was no change in the brain lobe volume of mutant larvae. Overall, our work describes a novel adp mutant that can functionally replace the long-standing adp60 mutant and shows that the adp gene has no obvious involvement in brain growth.

Transcription Factors AhR and ARNT Regulate the Expression of CYP6SX1 and CYP3828A1 Involved in Insecticide Detoxification in Bradysia odoriphaga.

Aryl hydrocarbon receptor (AhR) and aryl hydrocarbon receptor nuclear translocator (ARNT) mediate the responses of adaptive metabolism to various xenobiotics. Here, we found that BoAhR and BoARNT are highly expressed in the midgut of Bradysia odoriphaga larvae. The expression of BoAhR and BoARNT was significantly increased after exposure to imidacloprid and phoxim. The knockdown of BoAhR and BoARNT significantly decreased the expression of CYP6SX1 and CYP3828A1 as well as P450 enzyme activity and caused a significant increase in the sensitivity of larvae to imidacloprid and phoxim. Exposure to ß-naphthoflavone (BNF) significantly increased the expression of BoAhR, BoARNT, CYP6SX1, and CYP3828A1 as well as P450 activity and decreased larval sensitivity to imidacloprid and phoxim. Furthermore, CYP6SX1 and CYP3828A1 were significantly induced by imidacloprid and phoxim, and the silencing of these two genes significantly reduced larval tolerance to imidacloprid and phoxim. Taken together, the BoAhR/BoARNT pathway plays key roles in larval tolerance to imidacloprid and phoxim by regulating the expression of CYP6SX1 and CYP3828A1.

Genetic Disruption of cyp21a2 Leads to Systemic Glucocorticoid Deficiency and Tissues Hyperplasia in the Teleost Fish Medaka (Oryzias latipes).

cytochrome P-450, 21-hydroxylase (cyp21a2), encodes an enzyme required for cortisol biosynthesis, and its mutations are the major genetic cause of congenital adrenal hyperplasia (CAH) in humans. Here, we have generated a null allele for the medaka cyp21a2 with a nine base-pair insertion which led to a truncated protein. We have observed a delay in hatching and a low survival rate in homozygous mutants. The interrenal gland (adrenal counterpart in teleosts) exhibits hyperplasia and the number of pomca-expressing cells in the pituitary increases in the homozygous mutant. A mass spectrometry-based analysis of whole larvae confirmed a lack of cortisol biosynthesis, while its corresponding precursors were significantly increased, indicating a systemic glucocorticoid deficiency in our mutant model. Furthermore, these phenotypes at the larval stage are rescued by cortisol. In addition, females showed complete sterility with accumulated follicles in the ovary while male homozygous mutants were fully fertile in the adult mutants. These results demonstrate that the mutant medaka recapitulates several aspects of cyp21a2-deficiency observed in humans, making it a valuable model for studying steroidogenesis in CAH.

Leptin signaling promotes blood vessel formation in the Xenopus tail during the embryo-larval transition.

The signals that regulate peripheral blood vessel formation during development are still under investigation. The hormone leptin promotes blood vessel formation, adipose tissue establishment and expansion, tumor growth, and wound healing, but the underlying mechanisms for these actions are currently unknown. We investigated whether leptin promotes angiogenesis in the developing tail fin using embryonic transgenic xflk-1:GFP Xenopus laevis, which express a green fluorescent protein on vascular endothelial cells to mark blood vessels. We found that leptin protein is expressed in endothelial cells of developing blood vessels and that leptin treatment via injection increased phosphorylated STAT3 signaling, which is indicative of leptin activation of its receptor, in blood vessels of the larval tail fin. Leptin administration via media increased vessel length, branching, and reconnection with the cardinal vein, while decreased leptin signaling via immunoneutralization had an opposing effect on vessel development. We also observed disorganization of major vessels and microvessels of the tail fin and muscle when leptin signaling was decreased. Reduced leptin signaling lowered mRNA expression of cenpk, gpx1, and mmp9, markers for cell proliferation, antioxidation, and extracellular matrix remodeling/cell migration, respectively, in the developing tail, providing insight into three possible mechanisms underlying leptin's promotion of angiogenesis. Together these results illustrate that leptin levels are correlated with embryonic angiogenesis and that leptin coordinates multiple aspects of blood vessel growth and development, showing that leptin is an important morphogen during embryonic development.

Synchronizing Drosophila larvae with the salivary gland reporter Sgs3-GFP for discovery of phenotypes in the late third instar stage.

The larval stage of the Drosophila melanogaster life cycle is characterized by rapid growth and nutrient storage that occur over three instar stages separated by molts. In the third instar, the steroid hormone ecdysone drives key developmental processes and behaviors that occur in a temporally-controlled sequence and prepare the animal to undergo metamorphosis. Accurately staging Drosophila larvae within the final third instar is critical due to the rapid developmental progress at this stage, but it is challenging because the rate of development varies widely across a population of animals even if eggs are laid within a short period of time. Moreover, many methods to stage third instar larvae are cumbersome, and inherent variability in the rate of development confounds some of these approaches. Here we demonstrate the usefulness of the Sgs3-GFP transgene, a fusion of the Salivary gland secretion 3 (Sgs3) and GFP proteins, for staging third instar larvae. Sgs3-GFP is expressed in the salivary glands in an ecdysone-dependent manner from the midpoint of the third instar, and its expression pattern changes reproducibly as larvae progress through the third instar. We show that Sgs3-GFP can easily be incorporated into experiments, that it allows collection of developmentally-equivalent individuals from a mixed population of larvae, and that its use enables precise assessment of changing levels of hormones, metabolites, and gene expression during the second half of the third instar.

Hedgehog signaling is required for larval muscle development and larval metamorphosis of the mussel Mytilus coruscus.

Understanding the developmental processes and signaling pathways involved in larval myogenesis and metamorphosis is crucial for comprehending the life history and adaptive strategies of marine organisms. In this study, we investigated the temporal and spatial patterns of myogenesis in the mussel Mytilus coruscus (Mc), focusing on the emergence and transformation of major muscle groups during different larval stages. We also explored the role of the Hedgehog (Hh) signaling pathway in regulating myogenesis and larval metamorphosis. The results revealed distinct developmental stages characterized by the emergence of specific muscular components, such as velum retractor muscles and anterior adductor muscles, in D-veliger and umbo larvae, which are responsible for the planktonic stage. In the pediveliger stage, posterior ventral, posterior adductor, and foot muscles appeared. After larval metamorphosis, the velum structure and its corresponding retractor muscles degenerate, indicating the transition from planktonic to benthic life. We observed a conserved pattern of larval musculature development and revealed a high degree of conservation across bivalve species, with comparable emergence times during myogenesis. Furthermore, exposure to the Hh signaling inhibitor cyclopamine impaired larval muscle development, reduced larval swimming activity, and inhibited larval metamorphosis in M. coruscus. Cyclopamine-mediated inhibition of Hh signaling led to reduced expression of four key genes within the Hh signaling pathway (McHh, McPtc, McSmo, and McGli) and the striated myosin heavy chain gene (McMHC). It is hypothesised that the abnormal larval muscle development in cyclopamine-treated groups may be an indirect effect due to disrupted McMHC expression. We provide evidence for the first time that cyclopamine treatment inhibited larval metamorphosis in bivalves, highlighting the potential involvement of Hh signaling in mediating larval muscle development and metamorphosis in M. coruscus. The present study provides insights into the dynamic nature of myogenesis and the regulatory role of the Hh signaling pathway during larval development and metamorphosis in M. coruscus. The results obtained in this study contribute to a better understanding of the evolutionary significance of Hh signaling in bivalves and shed light on the mechanisms underlying larval muscle development and metamorphosis in marine invertebrates.

Gustatory Receptor 206 Participates in the Foraging Behavior of Larvae of Polyphagous Pest Spodoptera litura.

Insect gustatory receptors (GRs) aid in the precise identification of deterrent or stimulant compounds associated with food, mating, and egg-laying. Thus, they are promising targets for developing efficient insecticides. Here, 61 GRs in the chemosensory organs of Spodoptera litura larvae and adults were identified. Among them, SlitGR206 exhibited larval labium (LL)-specific expression characteristics. To explore the role of SlitGR206, a bacterial expression system was established to produce high-quality double-stranded RNA (dsRNA) and suppress SlitGR206 expression in LL. Subsequent behavioral assessments revealed that SlitGR206 silencing influenced larval feeding preferences and absorption. Moreover, it was found to reduce the ability of larvae to forage the five crucial host odorants. These findings demonstrate that SlitGR206 likely plays an indirect regulatory role in host recognition, consequently affecting foraging behavior. This provides a crucial foundation for the analysis of functional diversity among insect GRs and the precise development of nucleic acid pesticides in the future.

Interactions between intestinal microbiota and metabolites in zebrafish larvae exposed to polystyrene nanoplastics: Implications for intestinal health and glycolipid metabolism.

Previous studies have shown the harmful effects of nanoscale particles on the intestinal tracts of organisms. However, the specific mechanisms remain unclear. Our present study focused on examining the uptake and distribution of polystyrene nanoplastics (PS-NPs) in zebrafish larvae, as well as its toxic effects on the intestine. It was found that PS-NPs, marked with red fluorescence, primarily accumulated in the intestine section. Subsequently, zebrafish larvae were exposed to normal PS-NPs (0.2-25 mg/L) over a critical 10-day period for intestinal development. Histopathological analysis demonstrated that PS-NPs caused structural changes in the intestine, resulting in inflammation and oxidative stress. Additionally, PS-NPs disrupted the composition of the intestinal microbiota, leading to alterations in the abundance of bacterial genera such as Pseudomonas and Aeromonas, which are associated with intestinal inflammation. Metabolomics analysis showed alterations in metabolites that are primarily involved in glycolipid metabolism. Furthermore, MetOrigin analysis showed a significant correlation between bacterial flora (Pedobacter and Bacillus) and metabolites (D-Glycerate 2-phosphate and D-Glyceraldehyde 3-phosphate), which are related to the glycolysis/gluconeogenesis pathways. These findings were further validated through alterations in multiple biomarkers at various levels. Collectively, our data suggest that PS-NPs may impair the intestinal health, disrupt the intestinal microbiota, and subsequently cause metabolic disorders.

Drosophila TET acts with PRC1 to activate gene expression independently of its catalytic activity.

Enzymes of the ten-eleven translocation (TET) family play a key role in the regulation of gene expression by oxidizing 5-methylcytosine (5mC), a prominent epigenetic mark in many species. Yet, TET proteins also have less characterized noncanonical modes of action, notably in Drosophila, whose genome is devoid of 5mC. Here, we show that Drosophila TET activates the expression of genes required for larval central nervous system (CNS) development mainly in a catalytic-independent manner. Genome-wide profiling shows that TET is recruited to enhancer and promoter regions bound by Polycomb group complex (PcG) proteins. We found that TET interacts and colocalizes on chromatin preferentially with Polycomb repressor complex 1 (PRC1) rather than PRC2. Furthermore, PRC1 but not PRC2 is required for the activation of TET target genes. Last, our results suggest that TET and PRC1 binding to activated genes is interdependent. These data highlight the importance of TET noncatalytic function and the role of PRC1 for gene activation in the Drosophila larval CNS.