Arbuscular Mycorrhizal Fungi Can Inhibit the Allocation of Microplastics from Crop Roots to Aboveground Edible Parts.

Microplastics are emerging pollutants that threaten soil health and food safety. Recently, there has been increasing interest in understanding the behavior of these particles in the rhizosphere, specifically regarding the potential uptake of microplastics into crops. Arbuscular mycorrhizal (AM) fungi are widespread soil fungi, forming symbiotic associations with most terrestrial plants. Therefore, it is essential to investigate if AM fungi could protect crops from microplastics in soil. Here, we grew vegetables (Lactuca sativa) inoculated with/without the AM fungus Rhizophagus irregularis at various levels of poly(methyl methacrylate) (PMMA) soil pollution (0, 0.05, 0.1, 0.2, and 0.4%, mass ratio of the pollutant to soil). Our findings revealed that the proportion of transport of PMMA from roots to shoots decreased significantly in mycorrhizal crops. This reduction occurred because some PMMA particles were immobilized by AM vesicles and intraradical fungal hyphae. However, AM symbiosis did not substantially reduce the uptake of microplastics by crops from soil. Mycorrhizal fungi might enhance the resistance of crops to microplastics through transforming the chemical properties of microplastics, reducing their complexation to crop components, and promoting crop phosphorus nutrition at high microplastic addition levels. Our study is the first report to achieve rapid quantification of microplastics in mycorrhizal crops using microscale combustion calorimetry, demonstrating that AM fungi have the ability to immobilize microplastics. The study allows a deeper insight into microplastic behavior in AM-associated crops and supports the potential application of AM fungi in crop cultivation under microplastic contamination.

Molecular Characterization and Expression Analysis of YABBY Genes in Chenopodium quinoa.

Plant-specific YABBY transcription factors play an important role in lateral organ development and abiotic stress responses. However, the functions of the YABBY genes in quinoa remain elusive. In this study, twelve YABBY (CqYAB) genes were identified in the quinoa genome, and they were distributed on nine chromosomes. They were classified into FIL/YAB3, YAB2, YAB5, INO, and CRC clades. All CqYAB genes consist of six or seven exons, and their proteins contain both N-terminal C2C2 zinc finger motifs and C-terminal YABBY domains. Ninety-three cis-regulatory elements were revealed in CqYAB gene promoters, and they were divided into six groups, such as cis-elements involved in light response, hormone response, development, and stress response. Six CqYAB genes were significantly upregulated by salt stress, while one was downregulated. Nine CqYAB genes were upregulated under drought stress, whereas six CqYAB genes were downregulated under cadmium treatment. Tissue expression profiles showed that nine CqYAB genes were expressed in seedlings, leaves, and flowers, seven in seeds, and two specifically in flowers, but no CqYAB expression was detected in roots. Furthermore, CqYAB4 could rescue the ino mutant phenotype in Arabidopsis but not CqYAB10, a paralog of CqYAB4, indicative of functional conservation and divergence among these YABBY genes. Taken together, these results lay a foundation for further functional analysis of CqYAB genes in quinoa growth, development, and abiotic stress responses.

Arbuscular Mycorrhizal Fungi Alter Arsenic Translocation Characteristics of Iris tectorum Maxim.

Arsenic (As) pollution in wetlands, mainly as As(III) and As(V), has threatened wetland plant growth. It has been well documented that arbuscular mycorrhizal (AM) fungi can alleviate As stress in terrestrial plants. However, whether AM fungi can protect natural wetland plants from As stress remains largely unknown. Therefore, three hydroponic experiments were conducted in which Iris tectorum Maxim. (I. tectorum) plants were exposed to As(III) or As(V) stresses, to investigate the effects of mycorrhizal inoculation on As uptake, efflux, and accumulation. The results suggested that short-term kinetics of As influx in I. tectorum followed the Michaelis-Menten function. Mycorrhizal inoculation decreased the maximum uptake rate (Vmax) and Michaelis constant (Km) of plants for As(III) influx, while yielding no significant difference in As(V) influx. Generally, mycorrhizal plants released more As into environments after 72 h efflux, especially under As(V) exposure. Moreover, mycorrhizal plants exhibited potential higher As accumulation capacity, probably due to more active As reduction, which was one of the mechanisms through which AM fungi mitigate As phytotoxicity. Our study has revealed the role of aerobic microorganism AM fungi in regulating As translocation in wetland plants and supports the involvement of AM fungi in alleviating plant As stress in anaerobic wetlands.

Integrative application of biochar and arbuscular mycorrhizal fungi for enhanced chromium resistance in Medicago sativa.

Soil chromium (Cr) contamination has become an environmental problem of global concern. However, the joint effects of combined utilization of biochar and arbuscular mycorrhizal (AM) fungal inoculum, which are considered as two promising remediation strategies of soil heavy metal pollutions, on plant Cr resistance are still poorly understood. In this study, a two-factor pot experiment was conducted to investigate how biochar and AM fungus Rhizophagus irregularis regulate Medicago sativa growth, physiological trait, nutrient and Cr uptake, relevant gene expressions, soil properties, and Cr speciation, independently or synergistically. The results showed that biochar notably decreased AM colonization, while biochar and AM fungus could simultaneously increase plant dry biomass. The greatest growth promotion was observed in mycorrhizal shoots at the highest biochar level (50 g kg-1 soil) by 91 times. Both biochar application and AM fungal inoculation enhanced plant photosynthesis and P nutrition, but the promoting effects of AM fungus on them were significantly greater than that of biochar. In addition, the combined application of biochar and AM fungus dramatically reduced shoot and root Cr concentrations by up to 92 % and 78 %, respectively, compared to the non-amended treatment. Meanwhile, down-regulated expressions were observed for metal chelating-related genes. Furthermore, Cr translocation from roots to shoots was reduced by both two soil amendments. Transcriptional levels of genes involved in reactive oxygen species and proline metabolisms were also regulated by biochar application and AM fungal colonization, leading to alleviation of Cr phytotoxicity. Furthermore, AM fungal inoculation slightly elevated soil pH but decreased plant-available soil P, which was, by contrast, lifted by biochar addition. The combined application reduced soil acid-extractable Cr concentration by 40 %. This study provides new insights into comprehensively understanding of the mechanisms of biochar and AM fungi combination on improving plant Cr tolerance.

The mediating role of diabetes stigma and self-efficacy in relieving diabetes distress among patients with type 2 diabetes mellitus: a multicenter cross-sectional study.

Background: Patients with diabetes mellitus often suffer from diabetes distress. Social support and certain psychological factors potentially influence diabetes distress, but studies exploring the mechanisms underlying these relationships are scarce. Objectives: To reveal the associations between social support, diabetes stigma, diabetes self-efficacy, and diabetes distress among patients with type 2 diabetes and the underlying mechanisms linking these variables. Design and methods: A multicenter cross-sectional study was adopted and a sample of 431 patients with type 2 diabetes was investigated. Social support, diabetes stigma, diabetes self-efficacy, and diabetes distress were surveyed with the Perceived Social Support Scale, Type 2 Diabetes Stigma Assessment Scale, Self-Efficacy for Diabetes Scale, and Diabetes Distress Scale, respectively. The hypothesized model was verified using structural equation modeling. Results: Social support and diabetes stigma had direct associations with diabetes distress. Diabetes stigma mediated the association between social support and diabetes distress, and the association between diabetes self-efficacy and diabetes distress. Diabetes stigma and self-efficacy exerted a chain mediation effect on the association between social support and diabetes distress. Conclusion: Social support and diabetes stigma were significant predictors of diabetes distress. Diabetes stigma and self-efficacy play essential mediating roles in relieving diabetes distress. This can provide guidance for the development of evidence- and theory-based interventions. Culturally sensitive interventions that aim to provide ongoing social support, decrease diabetes stigma, and enhance self-efficacy have the potential to relieve diabetes distress.

Response of soil fungal community to chromium contamination in agricultural soils with different physicochemical properties.

Chromium (Cr) contamination has been of great concern in agricultural soil health due to its persistence, toxicity and bioaccumulation. Fungi, as an essential regulator of soil remediation and biochemical processes, had an unclear response to Cr contamination. In this study, the composition, diversity and interaction mechanisms of fungal communities in agricultural soils from ten different provinces of China were investigated in order to elucidate the fungal community response to varying soil properties and Cr concentrations. The results showed that high concentrations of Cr led to substantial alterations in the fungal community composition. The complex soil properties had a far greater impact on the fungal community structure than the single factor of Cr concentration, with soil available phosphorus (AP) and pH being most influential. Function predictions based on FUNGuild indicated that high concentrations of Cr have a significant impact on certain functional groups of fungi, including mycorrhizal fungi and plant saprotroph. The fungal community tended to resist Cr stress by enhancing interactions and clustering among network modules, while generating new keystone taxa. This study allowed insights into the response of soil fungal community to Cr contamination in different agricultural soils from different provinces and provided a theoretical basis for soil Cr ecological risk assessment and the development of bioremediation techniques for Cr-contaminated soils.

Target gene selection for RNAi-based biopesticides against the hawthorn spider mite, Amphitetranychus viennensis (Acari: Tetranychidae).

BACKGROUND: Recently, RNA interference (RNAi)-based biopesticide, a species-specific pest control alternative, has been deregulated and commercialized in the US and Canada. The hawthorn spider mite, Amphitetranychus viennensis Zacher, is a major pest for rosaceous plants, which has been controlled primarily by synthetic pesticides. To address the emerging resistance issues in A. viennensis, we initiated a project to develop RNAi-based biopesticides. RESULTS: In this study, we (i) developed a dietary RNAi system for A. viennensis using leaf disc, (ii) assessed the suitability of multiple control genes to distinguish sequence-specific silencing from non-specific effects within this RNAi system, and (iii) screened for the target gene candidates. As a result, ß-Glucuronidase (GUS), an enzyme derived from E. coli and a broadly used reporter for plants is the appropriate control for A. viennensis RNAi, while green fluorescent protein (GFP), is not suitable due to its significantly higher mortality than the other controls. For target gene screening, suppression was confirmed for all the candidates, including two housekeeping genes (Vacuolar-type H + -ATPase subunit A (V-ATPase A) and Glyceraldehyde 3-phosphate dehydrogenase, (GAPDH)), and three genes associated with development (ATP-dependent RNA Helicase DDX3Y (Belle), CREB-binding protein (CBP), and Farnesoic acid O-methyltransferase (FaMet)). Knocking down of V-ATPase A resulted in the highest mortality (~ 90%) and reduced fecundity (over 90%) than other candidates. As for the genes associated with development, suppression of Belle and CBP, led to approximately 65% mortality, as well as 86% and 40% reduction in fecundity, respectively. Silencing of FaMet, however, had negligible biological impacts on A. viennensis. CONCLUSION: The combined efforts not only establish an effective dsRNA delivery method, but also provide potential target genes for RNAi-based biopesticides against A. viennensis, a devastating invasive pest for fruit trees and woody ornamental plants throughout Asia and Europe. © 2023 Society of Chemical Industry.

Community response of soil microorganisms to combined contamination of polycyclic aromatic hydrocarbons and potentially toxic elements in a typical coking plant.

Both polycyclic aromatic hydrocarbons (PAHs) and potentially toxic elements (PTEs) of coking industries impose negative effects on the stability of soil ecosystem. Soil microbes are regarded as an essential moderator of biochemical processes and soil remediation, while their responses to PAHs-PTEs combined contamination are largely unknown. In the present study, soil microbial diversity and community composition in the typical coking plant under the chronic co-exposure of PAHs and PTEs were investigated and microbial interaction networks were built to reveal microbial co-occurrence patterns. The results indicated that the concentrations of PAHs in the soil inside the coking plant were significantly higher than those outside the plant. The mean concentration of ∑16PAHs was 2894.4 ng·g-1, which is 5.58 times higher than that outside the plant. The average Hg concentration inside the coking plant was 22 times higher than the background value of Hebei province. The soil fungal community inside the coking plant showed lower richness compared with that of outside community, and there are significant difference in the bacterial and fungal community composition between inside and outside of coking plant (p < 0.01). Predicted contribution of different environmental factors to each dominant species based on random forest identified 20 and 25 biomarkers in bacteria and fungi, respectively, that were highly sensitive to coking plant soil in operation, such as Betaproteobacteria，Sordariomycetes and Dothideomycetes. Bacterial and fungal communities were shaped by the soil chemical properties (pH), PTEs (Hg), and PAHs together in the coking plant soils. Furthermore, the bacterial and fungal interaction patterns were investigated separately or jointly by intradomain and interdomain networks. Competition is the main strategy based on the co-exclusion pattern in fungal community, and the competitive relationship inside the coking plant is more complex than that outside the plant. In contrast, cooperation is the dominant strategy in bacterial networks based on the co-occurrence pattern. The present study provided insights into microbial response strategies and the interactions between bacteria and fungi under long-term combined contamination.

Genome-Wide Identification and Characterization of SPL Family Genes in Chenopodium quinoa.

SQUAMOSA PROMOTER BINDING PROTEIN-LIKE (SPL) genes encode a large family of plant-specific transcription factors that play important roles in plant growth, development, and stress responses. However, there is little information available on SPL genes in Chenopodiaceae. Here, 23 SPL genes were identified and characterized in the highly nutritious crop Chenopodium quinoa. Chromosome localization analysis indicated that the 23 CqSPL genes were unevenly distributed on 12 of 18 chromosomes. Two zinc finger-like structures and a nuclear location signal were present in the SBP domains of all CqSPLs, with the exception of CqSPL21/22. Phylogenetic analysis revealed that these genes were classified into eight groups (group I-VIII). The exon-intron structure and motif composition of the genes in each group were similar. Of the 23 CqSPLs, 13 were potential targets of miR156/7. In addition, 5 putative miR156-encoding loci and 13 putative miR157-encoding loci were predicted in the quinoa genome, and they were unevenly distributed on chromosome 1-4. The expression of several Cqu-MIR156/7 loci was confirmed by reverse transcription polymerase chain reaction in seedlings. Many putative cis-elements associated with light, stress, and phytohormone responses were identified in the promoter regions of CqSPLs, suggesting that CqSPL genes are likely involved in the regulation of key developmental processes and stress responses. Expression analysis revealed highly diverse expression patterns of CqSPLs among tissues. Many CqSPLs were highly expressed in leaves, flowers, and seeds, and their expression levels were low in the roots, suggesting that CqSPLs play distinct roles in the development and growth of quinoa. The expression of 13 of 23 CqSPL genes responded to salt treatment (11 up-regulated and 2 down-regulated). A total of 22 of 23 CqSPL genes responded to drought stress (21 up-regulated and 1 down-regulated). Moreover, the expression of 14 CqSPL genes was significantly altered following cadmium treatment (3 up-regulated and 11 down-regulated). CqSPL genes are thus involved in quinoa responses to salt/drought and cadmium stresses. These findings provide new insights that will aid future studies of the biological functions of CqSPLs in C. quinoa.

Long-term nickel contamination increased soil fungal diversity and altered fungal community structure and co-occurrence patterns in agricultural soils.

Nickel (Ni) contamination imposes deleterious effects on the stability of soil ecosystem. Soil fungal community as a crucial moderator of soil remediation and biochemical processes has attracted more and more research interests. In the present study, soil fungal community composition and diversity under long-term Ni contamination were investigated and fungal interaction networks were built to reveal fungal co-occurrence patterns. The results showed that moderate Ni contamination significantly increased fungal diversity and altered fungal community structure. Functional predictions based on FUNGuild suggested that the relative abundance of arbuscular mycorrhizal fungi (AMF) significantly increased at moderate Ni contamination level. Ni contamination strengthened fungal interactions. Keystone taxa at different Ni contamination levels, such as Penicillium at light contamination, were identified, which might have ecological significance in maintaining the stability of fungal community to Ni stress. The present study provided a deeper insight into the effect of long-term Ni contamination on fungal community composition and co-occurrence patterns, and was helpful to further explore ecological risk of Ni contamination in cultivated field.

TDRD5 Is Required for Spermatogenesis and Oogenesis in Locusta migratoria.

Tudor family proteins exist in all eukaryotic organisms and play a role in many cellular processes by recognizing and binding to proteins with methylated arginine or lysine residues. TDRD5, a member of Tudor domain-containing proteins (TDRDs), has been implicated in the P-element-induced wimpy testis-interacting RNA (piRNA) pathway and germ cell development in some model species, but little is known about its function in other species. Therefore, we identified and characterized LmTDRD5, the TDRD5 ortholog in Locusta migratoria, a hemimetabolous pest. The LmTdrd5 gene has 19 exons that encode a protein possessing a single copy of the Tudor domain and three LOTUS domains at its N-terminus. qRT-PCR analysis revealed a high LmTdrd5 expression level in genital glands. Using RNA interference, LmTdrd5 knockdown in males led to a lag in meiosis phase transition, decreased spermatid elongation and sperm production, and downregulated the expression of the two germ cell-specific transcription factors, LmCREM and LmACT, as well as the sperm tail marker gene LmQrich2.LmTdrd5 knockdown in females reduced the expression levels of vitellogenin (Vg) and Vg receptor (VgR) and impaired ovarian development and oocyte maturation, thus decreasing the hatchability rate. These results demonstrate that LmTdrd5 is essential for germ cell development and fertility in locusts, indicating a conserved function for TDRD5.

DEAD-Box RNA Helicase DDX47 Maintains Midgut Homeostasis in Locusta migratoria.

Tissue homeostasis is critical for maintaining organ shape, size, and function. The condition is regulated by the balance between the generation of new cells and the loss of senescent cells, and it involves many factors and mechanisms. The midgut, an important part of the intestinal tract, is responsible for digestion and nutrient absorption in insects. LmDDX47, the ortholog of DEAD-box helicase 47 from Locusta migratoria, is indispensable for sustaining a normal midgut in the nymphs. However, the underlying cellular and molecular mechanisms remain to be elucidated. In this study, LmDDX47 knockdown resulted in atrophy of the midgut and gastric cecum in both nymph and adult locusts. After LmDDX47 knockdown, the number of regenerative and columnar cells in the midgut was significantly reduced, and cell death was induced in columnar tissue. LmDDX47 was localized to the nucleolus; this was consistent with the reduction in 18S rRNA synthesis in the LmDDX47 knockdown group. In addition, the acetylation and crotonylation levels of midgut proteins were significantly increased. Therefore, LmDDX47 could be a key regulator of midgut homeostasis, regulating 18S rRNA synthesis as well as protein acetylation and crotonylation in the migratory locust.

Structure and diversity of fungal communities in long-term copper-contaminated agricultural soil.

Copper (Cu) contamination threatens the stability of soil ecosystems. As important moderators of biochemical processes and soil remediation, the fungal community in contaminated soils has attracted much research interest. In this study, soil fungal diversity and community composition under long-term Cu contamination were investigated based on high-throughput sequencing. The co-occurrence networks were also constructed to display the co-occurrence patterns of the soil fungal community. The results showed that the richness and Chao1 index both significantly increased at 50 mg kg-1 Cu and then significantly decreased at 1600 and 3200 mg kg-1 Cu. Soil fungal diversity was significantly and positively correlated with plant dry weight. Specific tolerant taxa under different Cu contamination gradients were illustrated by linear discriminant analysis effect size (LEfSe). Soil Cu concentration and shoot dry weight were the strongest driving factors influencing fungal composition. The relative abundance of arbuscular mycorrhizal fungi increased first and then declined along with elevating Cu concentrations via FUNGuild analysis. The interactions among fungi were enhanced under light and moderate Cu contamination but weakened under heavy Cu contamination by random matrix theory (RMT)-based molecular ecological network analysis. Penicillium, identified as a keystone taxon in Cu-contaminated soils, had the function of removing heavy metals and detoxification, which might be vital to trigger the resistance of the fungal community to Cu contamination. The results may facilitate the identification of Cu pollution indicators and the development of in situ bioremediation technology for contaminated cultivated fields.

The toxicity of hexavalent chromium to soil microbial processes concerning soil properties and aging time.

Chromium (Cr) pollution has attracted much attention due to its biological toxicity. However, little is known regarding Cr toxicity to soil microorganisms. The present study assesses the toxicity of Cr(VI) on two microbial processes, potential nitrification rate (PNR) and substrate-induced respiration (SIR), in a wide range of agricultural soils and detected the abundance of soil bacteria, fungi, ammonia-oxidizing bacteria and archaea. The toxicity thresholds of 10% and 50% effective concentrations (EC10 and EC50) for PNR varied by 32.18- and 38.66-fold among different soils, while for SIR they varied by 391.21- and 16.31-fold, respectively. Regression model analysis indicated that for PNR, CEC as a single factor explained 27% of the variation in EC10, with soil clay being the key factor explaining 47.3% of the variation in EC50. For SIR, organic matter and pH were found to be the most vital predictors for EC10 and EC50, explaining 34% and 61.1% of variation, respectively. In addition, extended aging time was found to significantly attenuate the toxicity of Cr on PNR. SIR was mainly driven by total bacteria rather than fungi, while PNR was driven by both AOA and AOB. These results were helpful in deriving soil Cr toxicity threshold based on microbial processes, and provided a theoretical foundation for ecological risk assessments and establishing a soil environmental quality criteria for Cr.

Molecular Characterizations and Functional Analyses of LmR2D2 in the Locusta migratoria siRNA Pathway.

Small interfering RNAs (siRNAs) are non-coding RNAs with a length of 21~23 nucleotides (nt) and present in almost all eukaryotes. The formation of siRNA is a highly conserved post-transcriptional gene-silencing mechanism mediated by key proteins, including Dicer2, Argonaute2 (Ago2) and R2D2. R2D2 has been identified as a double-stranded RNA (dsRNA)-binding protein and reported as an integral component of the siRNA pathway in Drosophila. However, the involvement of R2D2 in the siRNA pathway of Locusta migratoria is still unknown. In the present study, we identified an LmR2D2 gene from the transcriptome of L. migratoria. It consists of a 954-bp open reading frame that encodes a protein of 318 amino acid residues. Further sequence analysis revealed that LmR2D2 possesses two tandem dsRNA-binding domains (dsRBD) at the N-terminus. Analysis of the developmental expression profile of LmR2D2 indicated that its transcript level was stable in third-instar nymphs of L. migratoria, whereas the tissue-dependent expression profile exhibited high levels of expression of LmR2D2 in the testis and ovary. When LmR2D2 was silenced by RNAi, the RNAi efficiency against Lmß-tubulin as a marker gene was significantly diminished, as indicated by the 37.7% increased Lmß-tubulin transcript level. Additionally, the prokaryotic expression system was used to obtain the LmR2D2 supernatant protein. By incubating the LmR2D2 protein with biotin-dsRNA, we found that LmR2D2 can bind to dsRNA in vitro, which supports our conclusion that LmR2D2 plays an essential role in the siRNA pathway of L. migratoria.

The RNA helicase DDX3 is required for ovarian development and oocyte maturation in Locusta migratoria.

DDX3 represents a well-defined subfamily of DEAD-box RNA helicase and exerts multiple functions in RNA metabolism, cell cycle, tumorigenesis, signal pathway, and fertility. Our previous study has shown that LmDDX3, the ortholog of DDX3 in Locusta migratoria, is ubiquitously expressed, and with a high abundance in testis and ovary. Knockdown of LmDDX3 results in a lethal phenotype in nymph, but it still remains unclear for its role in reproductive process. In this study, we therefore characterized LmDDX3 expression in female adult locust and analyzed its function in oocyte development. LmDDX3 was expressed in all tissues examined with significant more transcripts in ovary and hindgut. In ovary, a strong expression level was detected at the day just after adult eclosion, and a dramatic reduction then occurred during the oocyte development. LmDDX3 RNAi led to a reduced vitellogenin (Vg) expression in fat body via partially at least, the JH signaling pathway, and caused an upregulation of vitellogenin receptor (VgR) in ovary, and thus blocked the ovarian development and oocyte maturation. Sequence and phylogenetic analysis indicated that LmDDX3 was closely related to termite DDX3. Taken together, these data reveal a critical role for LmDDX3 in regulating the transcription of Vg and VgR, two major factors in vitellogenesis that is a key process required for ovary development and oocyte maturation in locust, and contribute thereof a new putative target for locust biological control.

Endoplasmic reticulum membrane receptors of the GET pathway are conserved throughout eukaryotes.

Type II tail-anchored (TA) membrane proteins are involved in diverse cellular processes, including protein translocation, vesicle trafficking, and apoptosis. They are characterized by a single C-terminal transmembrane domain that mediates posttranslational targeting and insertion into the endoplasmic reticulum (ER) via the Guided-Entry of TA proteins (GET) pathway. The GET system was originally described in mammals and yeast but was recently shown to be partially conserved in other eukaryotes, such as higher plants. A newly synthesized TA protein is shielded from the cytosol by a pretargeting complex and an ATPase that delivers the protein to the ER, where membrane receptors (Get1/WRB and Get2/CAML) facilitate insertion. In the model plant Arabidopsis thaliana, most components of the pathway were identified through in silico sequence comparison, however, a functional homolog of the coreceptor Get2/CAML remained elusive. We performed immunoprecipitation-mass spectrometry analysis to detect in vivo interactors of AtGET1 and identified a membrane protein of unknown function with low sequence homology but high structural homology to both yeast Get2 and mammalian CAML. The protein localizes to the ER membrane, coexpresses with AtGET1, and binds to Arabidopsis GET pathway components. While loss-of-function lines phenocopy the stunted root hair phenotype of other Atget lines, its heterologous expression together with the coreceptor AtGET1 rescues growth defects of Δget1get2 yeast. Ectopic expression of the cytosolic, positively charged N terminus is sufficient to block TA protein insertion in vitro. Our results collectively confirm that we have identified a plant-specific GET2 in Arabidopsis, and its sequence allows the analysis of cross-kingdom pathway conservation.

DDX6 Is Essential for Oocyte Development and Maturation in Locusta migratoria.

DEAD-box protein 6 (DDX6) is a member of the DDX RNA helicase family that exists in all eukaryotes. It has been extensively studied in yeast and mammals and has been shown to be involved in messenger ribonucleoprotein assembly, mRNA storage, and decay, as well as in miRNA-mediated gene silencing. DDX6 participates in many developmental processes but the biological function of DDX6 in insects has not yet been adequately addressed. Herein, we characterized the LmDDX6 gene that encodes the LmDDX6 protein in Locusta migratoria, a global, destructive pest. LmDDX6 possesses five motifs unique to the DDX6 subfamily. In the phylogenetic tree, LmDDX6 was closely related to its orthologs in Apis dorsata and Zootermopsis nevadensis. RT-qPCR data revealed high expression of LmDDX6 in the ovary, muscle, and fat body, with a declining trend in the ovary after adult ecdysis. LmDDX6 knockdown downregulated the expression levels of the juvenile hormone receptor Met, and genes encoding Met downstream targeted Grp78-1 and Grp78-2, reduced LmVg expression, and impaired ovary development and oocyte maturation. These results demonstrate that LmDDX6 plays an essential role in locust female reproduction and, thus, could be a novel target for locust biological control.

20-Hydroxyecdysone regulates the prophenoloxidase cascade to immunize Metarhizium anisopliae in Locusta migratoria.

BACKGROUND: PPO (prophenoloxidase) cascade plays an important role in resisting invasion of entomogenous fungus. The 20-hydroxyecdysone (20E) exerts potent effect on the innate immunity in many insects. However, whether 20E controls the PPO cascade system against fungi and the regulatory mechanism in insects remains unclear. RESULTS: In this study, both the proteome and transcriptome of Locusta migratoria were determined followed by the induction of 20E. Pattern recognition receptor GNBP-2 (Gram-negative binding proteins) has been identified that responded to 20E at both messenger RNA (mRNA) and protein levels. The PPO gene expression in fat body and PO (phenoloxidase) activity in plasma was found significantly induced after 20E injection and during the high-20E developmental stage. However, when 20E signal was blocked by RNA interference (RNAi) of ecdysone receptor, the expression level of PPO and PO activity failed to be increased by 20E. Thus, 20E could not significantly induce the expression of PPO gene and PO activity after RNAi of GNBP-2. Furthermore, 20E treatment notably enhanced the resistance of L. migratoria against Metarhizium anisopliae. Followed by of GNBP-2 silencing, the mortality of nymphs was significantly increased under the stress of Metarhizium anisopliae, and 20E injection could not increase the resistance. CONCLUSION: The 20E regulates the PPO system to resist fungal invasion via regulating GNBP-2 in worldwide pest L. migratoria. Our results provide insight into the mechanism of how 20E enhances the antimicrobial immunity, and will be beneficial for modification of entomogenous fungi targeting on hormones and the immune system. © 2020 Society of Chemical Industry.

Molecular characterization and RNA interference analysis of the DEAD-box gene family in Locusta migratoria.

DEAD-box (DDX) genes encode a group of RNA helicases that are highly conserved and ubiquitously expressed from prokaryotes to eukaryotes, and appear to participate in almost every aspect of RNA metabolism. Studies have been extensively done in yeast and human, in insect, beyond the flies, however, the information of these genes is limited. Here, we therefore identified and characterized 32 DDX genes from Locusta migratoria (L. migratoria), a crop pest. Overview of the gene structure and domain composition showed that the gene size varies significantly from one to fifteen exons, and the encoded proteins contain the conserved helicase core with various extensions at their amino and carboxyl termini. Phylogenetic trees informed that these locust DDX family members have orthologs in all insect species examined and can be classified into 30 subfamilies, all of them found counterparts in human, and most in yeast as well. Quantitative real-time PCR revealed that these genes are expressed in all stages and tissues examined, overall with higher expression level at second and third-instar nymphs and in the reproductive organs. RNA interference (RNAi) analyses showed that seven genes cause lethal phenotype when silenced, of which five lead to defective midgut and gastric caecum, indicating that these genes are essential for the survival and maintenance of normal digestive organs of locust. These data provide a foundation for further functional analysis of these DDX genes in locust.