Information Integration and Communication in Plant Growth Regulation.

Plants are equipped with the capacity to respond to a large number of diverse signals, both internal ones and those emanating from the environment, that are critical to their survival and adaption as sessile organisms. These signals need to be integrated through highly structured intracellular networks to ensure coherent cellular responses, and in addition, spatiotemporal actions of hormones and peptides both orchestrate local cell differentiation and coordinate growth and physiology over long distances. Further, signal interactions and signaling outputs vary significantly with developmental context. This review discusses our current understanding of the integrated intracellular and intercellular signaling networks that control plant growth.

Mapping the signaling network of BIN2 kinase using TurboID-mediated biotin labeling and phosphoproteomics.

Elucidating enzyme-substrate relationships in posttranslational modification (PTM) networks is crucial for understanding signal transduction pathways but is technically difficult because enzyme-substrate interactions tend to be transient. Here, we demonstrate that TurboID-based proximity labeling (TbPL) effectively and specifically captures the substrates of kinases and phosphatases. TbPL-mass spectrometry (TbPL-MS) identified over 400 proximal proteins of Arabidopsis thaliana BRASSINOSTEROID-INSENSITIVE2 (BIN2), a member of the GLYCOGEN SYNTHASE KINASE 3 (GSK3) family that integrates signaling pathways controlling diverse developmental and acclimation processes. A large portion of the BIN2-proximal proteins showed BIN2-dependent phosphorylation in vivo or in vitro, suggesting that these are BIN2 substrates. Protein-protein interaction network analysis showed that the BIN2-proximal proteins include interactors of BIN2 substrates, revealing a high level of interactions among the BIN2-proximal proteins. Our proteomic analysis establishes the BIN2 signaling network and uncovers BIN2 functions in regulating key cellular processes such as transcription, RNA processing, translation initiation, vesicle trafficking, and cytoskeleton organization. We further discovered significant overlap between the GSK3 phosphorylome and the O-GlcNAcylome, suggesting an evolutionarily ancient relationship between GSK3 and the nutrient-sensing O-glycosylation pathway. Our work presents a powerful method for mapping PTM networks, a large dataset of GSK3 kinase substrates, and important insights into the signaling network that controls key cellular functions underlying plant growth and acclimation.

The F-box Protein KIB1 Mediates Brassinosteroid-Induced Inactivation and Degradation of GSK3-like Kinases in Arabidopsis.

The glycogen synthase kinase-3 (GSK3) family kinases are central cellular regulators highly conserved in all eukaryotes. In Arabidopsis, the GSK3-like kinase BIN2 phosphorylates a range of proteins to control broad developmental processes, and BIN2 is degraded through unknown mechanism upon receptor kinase-mediated brassinosteroid (BR) signaling. Here we identify KIB1 as an F-box E3 ubiquitin ligase that promotes the degradation of BIN2 while blocking its substrate access. Loss-of-function mutations of KIB1 and its homologs abolished BR-induced BIN2 degradation and caused severe BR-insensitive phenotypes. KIB1 directly interacted with BIN2 in a BR-dependent manner and promoted BIN2 ubiquitination in vitro. Expression of an F-box-truncated KIB1 caused BIN2 accumulation but dephosphorylation of its substrate BZR1 and activation of BR responses because KIB1 blocked BIN2 binding to BZR1. Our study demonstrates that KIB1 plays an essential role in BR signaling by inhibiting BIN2 through dual mechanisms of blocking substrate access and promoting degradation.

M-Sn-Q (M = Zn, Cd; Q = S, Se) Compounds Templated by (Alkyl)ammonium Species: Synthesis, Crystal Structure, and Sr2+ Adsorption Property.

Deep investigations on the synthetic and structural chemistry of heterometallic chalcogenidostannates bear fundamental significance for the establishment of the structure-property relationship that would offer guidance on the functional material innovation. Presented here are four ammonium- and/or alkylammonium-directed M-Sn-Q (M = Zn, Cd; Q = S, Se) compounds, namely, [NH4]7[H3O]3Zn4Sn4S17 (1), [NH4]5[(CH3)2NH2]Zn4Sn5S17 (2), [CH3CH2NH3]22Zn16Sn12Se51(H2O)4·16H2O (3), and [NH4]2CdSnSe4 (4). All four compounds were synthesized in deep eutectic solvents (DESs) or ethylamine aqueous solution, both of which function simultaneously as reaction media and structure-directing agents. Compound 1 consists of discrete P1-[Zn4Sn4S17]10- clusters templated by mixed [NH4]+/[H3O]+ cations. In compound 2, such P1 clusters are bridged by Sn4+ ions in a 4,4-connection mode to form a [Zn4Sn5S17]n6n- framework with three types of cavities (I-III) varying in size. The two smaller cavities (I and II) accommodate NH4+ while the larger one(III) is occupied by [(CH3)2NH2]+, reflecting the rational size-dependence of cations on cavities. Compound 3 features an [Zn16Sn12Se51(H2O)4]n22n- open framework constructed from the 4,3-connection of P1-[Zn4Sn4Se17]10- clusters and {Zn(H2O)}2+ bridges. This linkage mode contributes to a large cage-like subunit (inner dimension: 21.99 × 9.06 Å2) and therefore an ultrahigh porosity that are occupied by [CH3CH2NH3]+ cations and water molecules (volume fraction: 57.7%). Compound 4 exists as a stacking of [CdSnSe4]n2n- chains, which are composed of alternatively arranged {CdSe4} and {SnSe4} tetrahedra, in combination with [NH4]+ cations as both charge-compensating and space-filling agents. Detailed synthetic, structural, and topological analyses were performed on these solid materials, coupled with extensive investigations on their optical and thermal properties. Compound 3 exhibits an efficient Sr2+ adsorption performance, featuring ultrafast kinetics (94.69% in 5 min), high removal rate (98.57% in 20 min) at equilibrium, and high capacity (104.17 ± 23.53 mg g-1).

Comparative genomic analysis of ABC transporter genes in Tenebrio molitor and four other tenebrionid beetles (Coleoptera: Tenebrionidea).

ATP-binding cassette (ABC) transporters, one of the largest transmembrane protein families, transport a diverse number of substate across membranes. Details of their diverse physiological functions have not been established. Here, we identified 87 ABC transporter genes in the genomes of Tenebrio molitor along with those from Asbolus verrucosus (104), Hycleus cichorii (65), and Hycleus phaleratus (80). Combining these genes (336 in total) with genes reported in Tribolium castaneum (73), we analyzed the phylogeny of ABC transporter genes in all five Tenebrionids. They are assigned into eight subfamilies (ABCA-H). In comparison to other species, the ABCC subfamily in this group of beetles appears expanded. The expression profiles of the T. molitor genes at different life stages and in various tissues were also investigated using transcriptomic analysis. Most of them display developmental specific expression patterns, suggesting to us their possible roles in development. Most of them are highly expressed in detoxification-related tissues including gut and Malpighian tubule, from which we infer their roles in insecticide resistance. We detected specific or abundant expressions of many ABC transporter genes in various tissues such as salivary gland, ovary, testis, and antenna. This new information helps generate new hypotheses on their biological significance within tissues.

Characterization and expression profiling of serine protease inhibitors in the yellow mealworm Tenebrio molitor.

Serine protease inhibitors (SPIs) act in diverse biological processes in insects such as immunity, development, and digestion by preventing the unwanted proteolysis. So far, the repertoire of genes encoding SPIs has been identified from few insect species. In this study, 62 SPI genes were identified from the genome of the yellow mealworm, Tenebrio molitor. According to their modes of action, they were classified into three families, serpin (26), canonical SPI (31), and α-macroglobulins (A2M) (5). These SPIs feature eight domains including serpin, Kazal, TIL, Kunitz, WAP, Antistasin, pacifastin, and A2M. In total, 39 SPIs contain a single SPI domain, while the others encode at least two inhibitor units. Based on the amino acids in the cleaved reactive sites, the abilities of these SPIs to inhibit trypsin, chymotrypsin, or elastase-like enzymes are predicted. The expression profiling based on the RNA-seq data showed that these genes displayed stage-specific expression patterns during development, suggesting to us their significance in development. Some of the SPI genes were exclusively expressed in particular tissues such as hemocyte, fat body, gut, ovary, and testis, which may be involved in biological processes specific to the indicated tissues. These findings provide necessary information for further investigation of insect SPIs.

Genomic and transcriptomic analyses of chitin metabolism enzymes in Tenebrio molitor.

Chitin is of great importance in the cuticle and inner cuticular linings of insects. Chitin synthases (CHSs), chitin deacetylases (CDAs), chitinases (CHTs), and ß-N-acetylhexosaminidases (HEXs) are important enzymes required for chitin metabolism, and play essential roles in development and metamorphosis. Although chitin metabolism genes have been well characterized in limited insects, the information in the yellow mealworm, Tenebrio molitor, a model insect, is presently still unavailable. With the help of bioinformatics, we identified 54 genes that encode putative chitin metabolism enzymes, including 2 CHSs, 10 CDAs, 32 CHTs, and 10 HEXs in the genome of T. molitor. All these genes have the conserved domains and motifs of their corresponding protein family. Phylogenetic analyses indicated that CHS genes were divided into two groups. CDA genes were clustered into five groups. CHT genes were phylogenetically grouped into 11 clades, among which 1 in the endo-ß-N-acetylglucosaminidases group and the others were classified in the glycoside hydrolase family 18 groups. HEX genes were assorted into six groups. Developmental and tissue-specific expression profiling indicated that the identified chitin metabolism genes showed dynamical expression patterns concurrent with specific instar during molting period, suggesting their significant roles in molting and development. They were predominantly expressed in different tissues or body parts, implying their functional specialization and diversity. The results provide important information for further clarifying their biological functions using the yellow mealworm as an ideal experimental insect.

Identification and expression profiling of serine protease-related genes in Tenebrio molitor.

In insects, serine proteases and serine protease homologs (SPs/SPHs) are involved in a variety of physiological processes including digestion, development, and immunity. Here, we identified 112 SP and 88 SPH genes in the genome of the yellow mealworm, Tenebrio molitor. Based on the features of domain structure, they were divided into "S" group containing single Tryp-SPc or Tryp-SPHc domain, "C" group containing 1-4 CLIP domain (CLIPA-D) and "M" group containing the CBD, CUB, EGF, Fz, Gd, LDLa, PAN, SEA, SR, Sushi, and TSP domains, and have 115, 48, and 37 gene members, respectively. According to the active sites in the catalytic triad, the putative trypsin, chymotrypsin, or elastase-like enzyme specificity of the identified SPs/SPHs were predicted. Phylogenetic and genomic location analyses revealed that gene duplication exists in the large amount of SPs/SPHs. Gene expression profiling using RNA-seq data along with real time reverse transcription-polymerase chain reaction analysis showed that most SP/SPH genes display life stage specific expression patterns, indicating their important roles in development. Many SP/SPH genes are specifically or highly expressed in the gut, salivary gland, fat body, hemocyte, ovary, and testis, suggesting that they participate in digestion, immunity, and reproduction. The findings lay the foundation for further functional characterization of SPs/SPHs in T. molitor.

Genome-wide and expression-profiling analyses of the cytochrome P450 genes in Tenebrionidea.

Cytochrome P450 monooxygenases (CYPs) are present in almost all areas of the tree of life. As one of the largest and most diverse superfamilies of multifunctional enzymes, they play important roles in the metabolism of xenobiotics and biosynthesis of endogenous compounds, shaping the success of insects. In this study, the CYPome (an omics term for all the CYP genes in a genome) diversification was examined in the four Tenebrionidea species through genome-wide analysis. A total of 483 CYP genes were identified, of which 103, 157, 122, and 101 were respectively deciphered from the genomes of Tebebrio molitor, Asbolus verucosus, Hycleus cichorii and Hycleus phaleratus. These CYPs were classified into four major clans (mitochondrial, CYP2, CYP3, and CYP4), and clans CYP3 and CYP4 are most diverse. Phylogenetic analysis showed that most CYPs of these Tenebrionidea beetles from each clan had a very close 1:1 orthology to each other, suggesting that they originate closely and have evolutionally conserved function. Expression analysis at different developmental stages and in various tissues showed the life stage-, gut-, salivary gland-, fat body-, Malpighian tubule-, antennae-, ovary- and testis-specific expression patterns of T. molitor CYP genes, implying their various potential roles in development, detoxification, immune response, digestion, olfaction, and reproduction. Our studies provide a platform to understand the evolution of Tenebrionidea CYP gene superfamily, and a basis for further functional investigation of the T. molitor CYPs involved in various biological processes.

Comparative genomic analysis of carboxylesterase genes in Tenebrio molitor and other four tenebrionids.

Carboxylesterases (COEs) have various functions in wide taxons of organisms. In insects, COEs are important enzymes involved in the hydrolysis of a variety of ester-containing xenobiotics, neural signal transmission, pheromone degradation, and reproductive development. Understanding the diversity of COEs is basic to illustrate their functions. In this study, we identified 53, 105, 37, and 39 COEs from the genomes of Tenebrio molitor, Asbolus verucosus, Hycleus cichorii, and H. phaleratus in the superfamily of Tenebrionidea, respectively. Phylogenetic analysis showed that 234 COEs from these four species and those reported in Tribolium castaneum (63) could be divided into 12 clades and three major classes. The α-esterases significantly expanded in T. molitor, A. verucosus, and T. castaneum compared to dipteran and hymenopteran insects. In T. molitor, most COEs showed tissue and stage-specific but not a sex-biased expression. Our results provide insights into the diversity and evolutionary characteristics of COEs in tenebrionids, and lay a foundation for the functional characterization of COEs in the yellow mealworm.

The Wnt gene family in Tenebrio molitor and other coleopterans.

The Wnt gene family is involved in a wide range of developmental processes. Despite its significance, the evolution and function of Wnt genes remain largely unclear. Here, an exhaustive survey of Wnt genes was conducted in Tenebrio molitor and 17 other beetle genomes. A total of 146 Wnt genes were identified, creating a comprehensive coleopteran Wnt gene catalog. Comparative genomics indicates that dynamic evolutionary patterns of Wnt gene loss and duplication occurred in Coleoptera, leading to the diverse Wnt gene repertoire in various beetles. A striking loss of particular Wnt gene subfamilies occurs in Coleoptera. Remarkably, Wnt gene duplication was discovered for the first time in insects. Further analysis of Wnt gene expression in T. molitor indicates that each Wnt gene, including the duplicated ones, has a unique spatial or temporal expression pattern. The current study provides valuable insight into the evolution and functional validation of Wnt genes in Coleoptera.

The ionotropic receptor gene family in Lepidoptera and Trichoptera: Annotation, evolutionary and functional perspectives.

Lepidoptera (moths and butterflies) and Trichoptera (caddisflies), belonging to the superorder Amphiesmenoptera, are the most diverse insect orders as representatives of the terrestrial and aquatic insects, respectively. The insects of the two orders possess different biological and behavioral characteristics, especially their larvae, presumably resulting in the differences of the ionotropic receptor (IR) genes in numbers, sequence characteristics or gene structure. Here, we employed genomics, transcriptomics, bioinformatics, phylogenetics and molecular biology strategies to characterize the IR gene repertoire in Lepidoptera and Trichoptera. Genome and transcriptome analyses with exhaustive homology-based searches and manual efforts, in 32 lepidopterans and five trichopterans, led to the identification of 1449 genes encoding IRs with 1170 full-length sequences, representing the most comprehensive set of chemoreceptor superfamilies across the Amphiesmenoptera. Analysis of gene gains and losses in orthologous groups implied that some IRs were lost in related species, and multiple gene copies occurred mainly in divergent IRs (D-IRs) by gene duplications. Phylogenetic analysis of 2442 IR proteins from 67 species revealed that Lepidoptera and Trichoptera IRs could be classified into three subfamilies, i.e., 14 antennal IRs (A-IRs), five Lepidoptera-specific IRs (LS-IRs) and four D-IRs. Of the three subfamilies, A-IRs and LS-IRs members within orthologous groups exhibited high conservation of gene structure, but D-IRs shared extremely low amino acid identities (below 30%). Expression profiles revealed functional diversities of IRs from Bombyx mori and Papilio xuthus involving smell, taste or reproduction, in which some genes displayed sex-biased expression in antennae associated with specific chemosensory behaviors of female or male adults. Our current study has provided insights into the evolution, conservation and divergence of IRs between/within Lepidoptera and Trichoptera, and allows for further experiments to investigate IR functions.

Local climate adaptation and gene flow in the native range of two co-occurring fruit moths with contrasting invasiveness.

Invasive species pose increasing threats to global biodiversity and ecosystems. While previous studies have characterized successful invaders based on ecological traits, characteristics related to evolutionary processes have rarely been investigated. Here we compared gene flow and local adaptation using demographic analyses and outlier tests in two co-occurring moth pests across their common native range of China, one of which (the peach fruit moth, Carposina sasakii) has maintained its native distribution, while the other (the oriental fruit moth, Grapholita molesta) has expanded its range globally during the past century. We found that both species showed a pattern of genetic differentiation and an evolutionary history consistent with a common southwestern origin and northward expansion in their native range. However, for the noninvasive species, genetic differentiation was closely aligned with the environment, and there was a relatively low level of gene flow, whereas in the invasive species, genetic differentiation was associated with geography. Genome scans indicated stronger patterns of climate-associated loci in the noninvasive species. While strong local adaptation and reduced gene flow across its native range may have decreased the invasiveness of C. sasakii, this requires further validation with additional comparisons of invasive and noninvasive species across their native range.

Mixed Solvothermal Synthesis of Tn Cluster-Based Indium and Gallium Sulfides Using Versatile Ammonia or Amine Structure-Directing Agents.

Metal chalcogenide supertetrahedral Tn clusters are of current interest for their unique compositions and structures, which rely highly on the structure-directing agents. Herein, we report four novel Tn cluster-based indium and gallium sulfides, namely, [NH(CH3)3]4In4S10H4 (1), (NH3)4Ga4S6 (2), [NH3CH2CH3]5(NH2CH2CH3)2Ga11S19 (3), and [NH3CH2CH2OH]6Ga10S18·2NH2CH2CH2OH (4). All four compounds were solvothermally synthesized in mixed amine-ethanol solutions or deep eutectic solvent (DES), where ammonia/amine molecules play significant structure-directing roles in the speciation and crystal growth. (1) Being protonated, the trimethylamine and ethanolamine molecules surround the T2-[In4S10H4]4- clusters (for 1) and [Ga10S18]n6n- open framework (for 4), respectively, compensating for the negative charge of the inorganic moieties. (2) With the lone pair of electrons, the ammonia molecules in 2 coordinate directly to corner Ga3+ ions of the {Ga4S6} cage to give a neutral T2-(NH3)4Ga4S6 cluster. (3) For compound 3, part of the ethylamine molecules act as terminating ligands for the T1 and T3 units in the [Ga11S19(NH2CH2CH3)2]n5n- layer, while the rest act as interlamellar countercations upon protonation. Theoretical studies reveal the contributions of N, C, and H to the density of states (DOS) for 2 and 3 because of their hybrid structures that combine the ammonia/amine ligands with sulfide moieties together.

Genome sequence of a novel member of the order Picornavirales from the endoparasitoid wasp Diversinervus elegans.

Here, we report a novel RNA virus from an encyrtid endoparasitoid wasp (Diversinervus elegans). This virus has a genome of 8845 nucleotides in length with a poly(A) tail. It contains one open reading frame (ORF) encoding a single polyprotein that shares the most significant similarity to the polyproteins of dicistroviruses. Phylogenetic analysis suggested that this virus belongs to the family Dicistroviridae from the order Picornavirales, but its genomic organization is distinct from that of the other known dicistroviruses, which have two ORFs. Consequently, we propose that this virus is a member of a new species in the order Picornavirales, and have named it "Diversinervus elegans virus" (DEV).

Genome-based analysis reveals a novel SNMP group of the Coleoptera and chemosensory receptors in Rhaphuma horsfieldi.

Through an exhaustive homology-based approach, coupled with manual efforts, we annotated and characterized 128 sensory neuron membrane proteins (SNMPs) from genomes and transcriptomes of 22 coleopteran species, with 107 novel candidates. Remarkably, we discovered, for the first time, a novel SNMP group, defined as Group 4 based on the phylogeny, sequence characteristics, gene structure and organization. The lineage-specific expansions in SNMPs occurred mainly in the family Scarabaeidae, harboring 12 representatives in Onthophagus taurus as a typical gene duplication and the most massive set of SNMPs in insects to date. Transcriptome sequencing of Rhaphuma horsfieldi resulted in the yields of approximately 611.9 million clean reads that were further assembled into 543,841 transcripts and 327,550 unigenes, respectively. From the transcriptome, 177 transcripts encoding 84 odorant (ORs), 62 gustatory (GRs), 20 ionotropic (IRs), and 11 ionotropic glutamate (iGluRs) receptors were identified. Phylogenetic analysis classified RhorORs into six groups, RhorGRs into four subfamilies, and RhorIRs into 10 conserved antennal IRs and one divergent IRs. Expression profiles revealed that over 80% of chemosensory genes were specifically or highly transcribed in antennae or tarsi, suggestive of their olfactory and/or gustatory roles. This study has greatly complemented the resources for chemosensory genes in the cerambycid beetles, and most importantly, identifies a novel group of SNMPs in Coleoptera.

Brassinosteroid signaling network and regulation of photomorphogenesis.

In plants, the steroidal hormone brassinosteroid (BR) regulates numerous developmental processes, including photomorphogenesis. Genetic, proteomic, and genomic studies in Arabidopsis have illustrated a fully connected BR signal transduction pathway from the cell surface receptor kinase BRI1 to the BZR1 family of transcription factors. Genome-wide analyses of protein-DNA interactions have identified thousands of BZR1 target genes that link BR signaling to various cellular, metabolic, and developmental processes, as well as other signaling pathways. In controlling photomorphogenesis, BR signaling is highly integrated with the light, gibberellin, and auxin pathways through both direct interactions between signaling proteins and transcriptional regulation of key components of these pathways. BR signaling also cross talks with other receptor kinase pathways to modulate stomata development and innate immunity. The molecular connections in the BR signaling network demonstrate a robust steroid signaling system that has evolved in plants to orchestrate signal transduction, genome expression, metabolism, defense, and development.

Removal of Sr2+ Ions by a High-Capacity Indium Sulfide Exchanger Containing Permeable Layers with Large Pores.

An ethylammonium-templated indium sulfide, [CH3CH2NH3]6In8S15 (InS-2), featuring anionic layers perforated with large, 24-membered rings that facilitate the accommodation of hydrated Sr2+ ions is reported. InS-2 exhibits an excellent adsorption performance toward Sr2+ with a top-ranked capacity (qm = 143.29 mg g-1), rapid kinetics, wide pH durability (3-14), ß- and γ-radiation resistances, and a facile elution.

Genome-based identification and analysis of ionotropic receptors in Spodoptera litura.

The ability to sense and recognize various classes of compounds is of particular importance for survival and reproduction of insects. Ionotropic receptor (IR), a sub-family of the ionotropic glutamate receptor family, has been identified as one of crucial chemoreceptor super-families, which mediates the sensing of odors and/or tastants, and serves as non-chemosensory functions. Yet, little is known about IR characteristics, evolution, and functions in Lepidoptera. Here, we identify the IR gene repertoire from a destructive polyphagous pest, Spodoptera litura. The exhaustive analyses with genome and transcriptome data lead to the identification of 45 IR genes, comprising 17 antennal IRs (A-IRs), 8 Lepidoptera-specific IRs (LS-IRs), and 20 divergent IRs (D-IRs). Phylogenetic analysis reveals that S. litura A-IRs generally retain a strict single copy within each orthologous group, and two lineage expansions are observed in the D-IR sub-family including IR100d-h and 100i-o, likely attributed to gene duplications. Results of gene structure analysis classify the SlitIRs into four types: I (intronless), II (1-3 introns), III (5-9 introns), and IV (10-18 introns). Extensive expression profiles demonstrate that the majority of SlitIRs (28/43) are enriched in adult antennae, and some are detected in gustatory-associated tissues like proboscises and legs as well as non-chemosensory organs like abdomens and reproductive tissues of both sexes. These results indicate that SlitIRs have diverse functional roles in olfaction, taste, and reproduction. Together, our study has complemented the information on chemoreceptor genes in S. litura, and meanwhile allows for target experiments to identify potential IR candidates for the control of this pest.

Superoxide dismutase from venom of the ectoparasitoid Scleroderma guani inhibits melanization of hemolymph.

Superoxide dismutase (SOD) known as an important antioxidative stress protein has been recently found in venoms of several parasitoid wasps. However, its functions and characteristics as a virulent factor remain scarcely described. Here, we report the characterization of two venomous SOD genes (SguaSOD1 and SguaSOD3) from the ectoparasitoid, Scleroderma guani. The metal binding sites, cysteine amino acid positions and signature sequences of the SOD family were conserved within SguaSOD1 and SguaSOD3. Relatively high levels of their transcripts were observed in pupae followed a decrease in early adults, after which they had the highest transcriptions, indicating that their productions would be regulated in venom apparatus. Although the two genes showed lower expression in venom apparatus compared to head and thorax, the enzymatic assay revealed that SOD indeed had activity in venom. Further, we showed that recombinant SguaSOD3 suppressed melanization of host hemolymph, implying that this protein used as a virulent factor uniquely impacts the prophenoloxidase cascade.