The mitochondrial genome of a social wasp, Vespula rufa (Linnaeus, 1758) (Hymenoptera: Vespidae).

For an ongoing systematic study of Korean Vespidae, we analyzed the complete mitochondrial genome of a social wasp, Vespula rufa (Linnaeus 1758), from the South Korea. The mitogenome is 17,521 bp in length, comprising 13 protein-coding genes (PCGs), 22 transfer RNA (tRNA) genes, and two ribosomal RNA (rRNA) genes. The nucleotide composition is 40.5% adenines, 43.0% thymines, 6.0% guanines, and 10.5% cytosines. The GC content is 16.5%. A maximum-likelihood analysis was conducted to understand phylogenetic relationships, based on 13 complete mitogenome sequences of Vespinae species. We recognized that V. rufa is being placed basal within the genus Vespula. The complete mitochondrial genome of V. rufa provides useful genetic information for further studies.

A 70% Ethanol Neorhodomela munita Extract Attenuates RANKL-Induced Osteoclast Activation and H2O2-Induced Osteoblast Apoptosis In Vitro.

The rapid aging of the population worldwide presents a significant social and economic challenge, particularly due to osteoporotic fractures, primarily resulting from an imbalance between osteoclast-mediated bone resorption and osteoblast-mediated bone formation. While conventional therapies offer benefits, they also present limitations and a range of adverse effects. This study explores the protective impact of Neorhodomela munita ethanol extract (EN) on osteoporosis by modulating critical pathways in osteoclastogenesis and apoptosis. Raw264.7 cells and Saos-2 cells were used for in vitro osteoclast and osteoblast models, respectively. By utilizing various in vitro methods to detect osteoclast differentiation/activation and osteoblast death, it was demonstrated that the EN's potential to inhibit RANKL induced osteoclast formation and activation by targeting the MAPKs-NFATc1/c-Fos pathway and reducing H2O2-induced cell death through the downregulation of apoptotic signals. This study highlights the potential benefits of EN for osteoporosis and suggests that EN is a promising natural alternative to traditional treatments.

Structure-based functional analysis of a novel NADPH-producing glyceraldehyde-3-phosphate dehydrogenase from Corynebacterium glutamicum.

Corynebacterium glutamicum is an industrial workhorse applied in the production of valuable biochemicals. In the process of bio-based chemical production, improving cofactor recycling and mitigating cofactor imbalance are considered major solutions for enhancing the production yield and efficiency. Although, glyceraldehyde-3-phosphate dehydrogenase (GapDH), a glycolytic enzyme, can be a promising candidate for a sufficient NADPH cofactor supply, however, most microorganisms have only NAD-dependent GapDHs. In this study, we performed functional characterization and structure determination of novel NADPH-producing GapDH from C. glutamicum (CgGapX). Based on the crystal structure of CgGapX in complex with NADP cofactor, the unique structural features of CgGapX for NADP stabilization were elucidated. Also, N-terminal additional region (Auxiliary domain, AD) appears to have an effect on enzyme stabilization. In addition, through structure-guided enzyme engineering, we developed a CgGapX variant that exhibited 4.3-fold higher kcat, and 1.2-fold higher kcat/KM values when compared with wild-type. Furthermore, a bioinformatic analysis of 100 GapX-like enzymes from 97 microorganisms in the KEGG database revealed that the GapX-like enzymes possess a variety of AD, which seem to determine enzyme stability. Our findings are expected to provide valuable information for supplying NADPH cofactor pools in bio-based value-added chemical production.

Exploration for Asian longhorned beetle parasitoids in Korea using an improved sentinel log trap.

The Asian longhorned beetle (ALB), Anoplophora glabripennis (Motschulsky) (Coleoptera: Cerambycidae), is a destructive invasive woodboring insect pest, and efforts are being made to find parasitoids for ALB biological control. Through a four-year survey in Korea using a sentinel log trap associated with host chemical cues potentially important for host finding by parasitoids, two parasitoid species were discovered attacking ALB. One species is Spathius ibarakius Belokobylskij & Maetô, which is known to also parasitize citrus longhorned beetle, Anoplophora chinensis (Forster). The other parasitoid species, whose offspring were dead before imago, could not be morphologically identified at the adult stage. We attempted molecular and morphological identification of the larvae/pupae of the unidentified parasitoid; however, only superfamily-level identification was possible. The parasitism rate recovered in the logs was 0.3% by the unidentified parasitoid in Gapyeong-gun in 2019, while it reached 29.2% by S. ibarakius in Busan city in 2022. Future efforts for exploring ALB natural enemies in the pest's native range may focus on parasitoids with high parasitism rates.

Title: Exploration des parasitoïdes du longicorne asiatique en Corée à l'aide d'un piège à bûche sentinelle amélioré. Abstract: Le longicorne asiatique (LA), Anoplophora glabripennis (Motschulsky) (Coleoptera : Cerambycidae), est un insecte ravageur envahissant et destructeur du bois, et des efforts sont déployés pour trouver des parasitoïdes pour la lutte biologique contre lui. Au cours d'une étude de quatre ans en Corée utilisant un piège à bûche sentinelle associé à des signaux chimiques de l'hôte potentiellement importants pour la détection de l'hôte par les parasitoïdes, deux espèces de parasitoïdes ont été découvertes attaquant le longicorne. Une espèce est Spathius ibarakius Belokobylskij & Maetô, qui est connue pour parasiter également le longicorne des agrumes, Anoplophora chinensis (Forster). Les autres espèces de parasitoïdes, dont les descendants sont morts avant l'imago, n'ont pu être identifiées morphologiquement au stade adulte. Nous avons tenté une identification moléculaire et morphologique des larves/pupes du parasitoïde non identifié, mais seule une identification au niveau de la superfamille a été possible. Le taux de parasitisme observé dans les bûches était de 0,3 % par le parasitoïde non identifié à Gapyeong-gun en 2019, tandis qu'il atteignait 29,2 % par S. ibarakius dans la ville de Busan en 2022. Les efforts futurs pour explorer les ennemis naturels du capricorne dans l'aire de répartition naturelle du ravageur pourraient se concentrer sur les parasitoïdes à taux de parasitisme élevés.

Enhanced Osteocyte Differentiation: Cathepsin D and L Secretion by Human Adipose-Derived Mesenchymal Stem Cells.

Adipose-derived mesenchymal stem cells (ASCs) have the potential to differentiate into bone, cartilage, fat, and neural cells and promote tissue regeneration and healing. It is known that they can have variable responses to hypoxic conditions. In the present study, we aimed to explore diverse changes in the cells and secretome of ASCs under a hypoxic environment over time and to present the possibility of ASCs as therapeutic agents from a different perspective. The expression differences of proteins between normoxic and hypoxic conditions (6, 12, or 24 h) were specifically investigated in human ASCs using 2-DE combined with MALDI-TOF MS analysis, and secreted proteins in ASC-derived conditioned media (ASC-derived CM) were examined by an adipokine array. In addition, genetic and/or proteomic interactions were assessed using a DAVID and miRNet functional annotation bioinformatics analysis. We found that 64 and 5 proteins were differentially expressed in hypoxic ASCs and in hypoxic ASC-derived CM, respectively. Moreover, 7 proteins among the 64 markedly changed spots in hypoxic ASCs were associated with bone-related diseases. We found that two proteins, cathepsin D (CTSD) and cathepsin L (CTSL), identified through an adipokine array independently exhibited significant efficacy in promoting osteocyte differentiation in bone-marrow-derived mesenchymal stem cells (BM-MSCs). This finding introduces a promising avenue for utilizing hypoxia-preconditioned ASC-derived CM as a potential therapeutic approach for bone-related diseases.

Structure-Guided Protein Engineering of Glyceraldehyde-3-phosphate Dehydrogenase from Corynebacterium glutamicum for Dual NAD/NADP Cofactor Specificity.

Since the discovery of l-glutamate-producing Corynebacterium glutamicum, it has evolved to be an industrial workhorse. For biobased chemical production, suppling sufficient amounts of the NADPH cofactor is crucial. Glyceraldehyde-3-phosphate dehydrogenase (GAPDH), a glycolytic enzyme that converts glyceraldehyde-3-phosphate (G3P) to 1,3-bisphosphoglycerate and produces NADH, is a major prospective solution for the cofactor imbalance issue. In this study, we determined the crystal structure of GAPDH from C. glutamicum ATCC13032 (CgGAPDH). Based on the structural information, we generated six CgGAPDH variants, CgGAPDHL36S, CgGAPDHL36S/T37K, CgGAPDHL36S/T37K/P192S, CgGAPDHL36S/T37K/F100V/P192S, CgGAPDHL36S/T37K/F100L/P192S, and CgGAPDHL36S/T37K/F100I/P192S, that can produce both NADH and NAPDH. The final CgGAPDHL36S/T37K/F100V/P192S variant showed a 212-fold increase in enzyme activity for NADP as well as 200% and 30% increased activity for the G3P substrate under NAD and NADP cofactor conditions, respectively. In addition, crystal structures of CgGAPDH variants in complex with NAD(P) permit the elucidation of differences between wild-type CgGAPDH and variants in relation to cofactor stabilization.

Structural and Biochemical Studies on Product Inhibition of S-Adenosylmethionine Synthetase from Corynebacterium glutamicum.

S-Adenosylmethionine (SAM) acts as a methyl donor in living organisms, and S-adenosylmethionine synthetase (MetK) is an essential enzyme for cells, as it synthesizes SAM from methionine and adenosine triphosphate (ATP). This study determined the crystal structures of the apo form and adenosine/triphosphate complex form of MetK from Corynebacterium glutamicum (CgMetK). Results showed that CgMetK has an allosteric inhibitor binding site for the SAM product in the vicinity of the active site and is inhibited by SAM both competitively and noncompetitively. Through structure-guided protein engineering, the CgMetKE68A variant was developed that exhibited an almost complete release of inhibition by SAM with rather enhanced enzyme activity. The crystal structure of the CgMetKE68A variant revealed that the formation of a new hydrogen bond between Tyr66 and Glu102 by the E68A mutation disrupted the allosteric SAM binding site and also improved the protein thermal stability by strengthening the tetramerization of the enzyme.

Crystal Structures of 6-Phosphogluconate Dehydrogenase from Corynebacterium glutamicum.

Corynebacterium glutamicum (C. glutamicum) has been considered a very important and meaningful industrial microorganism for the production of amino acids worldwide. To produce amino acids, cells require nicotinamide adenine dinucleotide phosphate (NADPH), which is a biological reducing agent. The pentose phosphate pathway (PPP) can supply NADPH in cells via the 6-phosphogluconate dehydrogenase (6PGD) enzyme, which is an oxidoreductase that converts 6-phosphogluconate (6PG) to ribulose 5-phosphate (Ru5P), to produce NADPH. In this study, we identified the crystal structure of 6PGD_apo and 6PGD_NADP from C. glutamicum ATCC 13032 (Cg6PGD) and reported our biological research based on this structure. We identified the substrate binding site and co-factor binding site of Cg6PGD, which are crucial for understanding this enzyme. Based on the findings of our research, Cg6PGD is expected to be used as a NADPH resource in the food industry and as a drug target in the pharmaceutical industry.

Crystal Structure and Functional Characterization of Acetylornithine Aminotransferase from Corynebacterium glutamicum.

The amino acids l-arginine and l-ornithine are widely used in animal feed and as health supplements and pharmaceutical compounds. In arginine biosynthesis, acetylornithine aminotransferase (AcOAT) uses pyridoxal-5'-phosphate (PLP) as a cofactor for amino group transfer. Here, we determined the crystal structures of the apo and PLP complex forms of AcOAT from Corynebacterium glutamicum (CgAcOAT). Our structural observations revealed that CgAcOAT undergoes an order-to-disorder conformational change upon binding with PLP. Additionally, we observed that unlike other AcOATs, CgAcOAT exists as a tetramer. Subsequently, we identified the key residues involved in PLP and substrate binding based on structural analysis and site-directed mutagenesis. This study might provide structural insights on CgAcOAT, which can be utilized for the development of improved l-arginine production enzymes.

The Role of Epicardial Adipose Tissue-Derived MicroRNAs in the Regulation of Cardiovascular Disease: A Narrative Review.

Cardiovascular diseases have been leading cause of death worldwide for many decades, and obesity has been acknowledged as a risk factor for cardiovascular diseases. In the present review, human epicardial adipose tissue-derived miRNAs reported to be differentially expressed under pathologic conditions are discussed and summarized. The results of the literature review indicate that some of the epicardial adipose tissue-derived miRNAs are believed to be cardioprotective, while some others show quite the opposite effects depending on the underlying pathologic conditions. Furthermore, they suggest that that the epicardial adipose tissue-derived miRNAs have great potential as both a diagnostic and therapeutic modality. Nevertheless, mainly due to highly limited availability of human samples, it is very difficult to make any generalized claims on a given miRNA in terms of its overall impact on the cardiovascular system. Therefore, further functional investigation of a given miRNA including, but not limited to, the study of its dose effect, off-target effects, and potential toxicity is required. We hope that this review can provide novel insights to transform our current knowledge on epicardial adipose tissue-derived miRNAs into clinically viable therapeutic strategies for preventing and treating cardiovascular diseases.

A Low Concentration of Citreoviridin Prevents Both Intracellular Calcium Deposition in Vascular Smooth Muscle Cell and Osteoclast Activation In Vitro.

Vascular calcification (VC) and osteoporosis are age-related diseases and significant risk factors for the mortality of elderly. VC and osteoporosis may share common risk factors such as renin-angiotensin system (RAS)-related hypertension. In fact, inhibitors of RAS pathway, such as angiotensin type 1 receptor blockers (ARBs), improved both vascular calcification and hip fracture in elderly. However, a sex-dependent discrepancy in the responsiveness to ARB treatment in hip fracture was observed, possibly due to the estrogen deficiency in older women, suggesting that blocking the angiotensin signaling pathway may not be effective to suppress bone resorption, especially if an individual has underlying osteoclast activating conditions such as estrogen deficiency. Therefore, it has its own significance to find alternative modality for inhibiting both vascular calcification and osteoporosis by directly targeting osteoclast activation to circumvent the shortcoming of ARBs in preventing bone resorption in estrogen deficient individuals. In the present study, a natural compound library was screened to find chemical agents that are effective in preventing both calcium deposition in vascular smooth muscle cells (vSMCs) and activation of osteoclast using experimental methods such as Alizarin red staining and Tartrate-resistant acid phosphatase staining. According to our data, citreoviridin (CIT) has both an anti-VC effect and anti-osteoclastic effect in vSMCs and in Raw 264.7 cells, respectively, suggesting its potential as an effective therapeutic agent for both VC and osteoporosis.

Organ-on-a-chip: Its use in cardiovascular research.

Organ-on-a-chip (OOAC) has attracted great attention during the last decade as a revolutionary alternative to conventional animal models. This cutting-edge technology has also brought constructive changes to the field of cardiovascular research. The cardiovascular system, especially the heart as a well-protected vital organ, is virtually impossible to replicate in vitro with conventional approaches. This made scientists assume that they needed to use animal models for cardiovascular research. However, the frequent failure of animal models to correctly reflect the native cardiovascular system necessitated a search for alternative platforms for preclinical studies. Hence, as a promising alternative to conventional animal models, OOAC technology is being actively developed and tested in a wide range of biomedical fields, including cardiovascular research. Therefore, in this review, the current literature on the use of OOACs for cardiovascular research is presented with a focus on the basis for using OOACs, and what has been specifically achieved by using OOACs is also discussed. By providing an overview of the current status of OOACs in cardiovascular research and its future perspectives, we hope that this review can help to develop better and optimized research strategies for cardiovascular diseases (CVDs) as well as identify novel applications of OOACs in the near future.

Nidus vespae Built by an Invasive Alien Hornet, Vespa velutina nigrithorax, Inhibits Adipose Tissue Expansion in High-Fat Diet-Induced Obese Mice

Nidus vespae, commonly known as the wasp nest, has antioxidative, anti-inflammatory, antimicrobial, and antitumor properties. However, the anti-obesity effects of Nidus vespae extract (NV) have not yet been reported. This study aimed to elucidate the potential anti-obesity effects of NV in vivo and in vitro, using a high-fat diet (HFD)-induced obese mouse model and 3T3-L1 adipocytes, respectively. NV administration to HFD-induced obese mice significantly decreased the mass and plasma lipid content of adipose tissues. Uncoupling protein-1 expression was significantly higher in the inguinal white adipose tissues of NV-treated mice than in those of HFD-fed mice. Furthermore, we found that NV inhibited the differentiation and intracellular lipid accumulation of 3T3-L1 adipocytes by regulating the insulin signaling cascade, including protein kinase B, peroxisome proliferator-activated receptor gamma, CCAAT/enhancer binding protein alpha, and adiponectin. These findings suggest that NV may exhibit therapeutic effects against obesity by suppressing adipose tissue expansion and preadipocyte differentiation, thereby providing critical information for the development of new drugs for disease prevention and treatment. To our knowledge, this study provides the first evidence of the anti-obesity effects of NV.

The mitochondrial genome of Nigidius miwai Nagel (Coleoptera: Lucanidae), a carnivorous lucanid from South Korea.

The complete mitochondrial genome of carnivorous Nigidius miwai was analyzed prior to a study on evolution of carnivorous pathway within the family. This species' genome had a total length of 18,462 bp (GenBank accession number: OL597607), comprising 36 genes along with 13 protein-coding genes, 21 tRNA genes, two rRNA genes, and an A + T rich region. The nucleotide composition was 37.2% A, 33.8% T, 9.2% G, and 19.8% C (GC contents 29.0%). The phylogenetic tree indicated that N. miwai is a distinctive species within Nigidiini of Lucaninae.

Brite Adipocyte FGF21 Attenuates Cardiac Ischemia/Reperfusion Injury in Rat Hearts by Modulating NRF2.

Although the optimal therapy for myocardial infarction includes reperfusion to restore blood flow to the ischemic area, myocardial injury after ischemia/reperfusion usually leads to an inflammatory response, oxidative stress, and cardiomyocyte apoptosis. In this study, rat adipose-derived stem cells were differentiated into low-thermogenic beige adipocytes (LBACs) and high-thermogenic beige adipocytes (HBACs) to study the different cardioprotective effects of heterogeneous expression of brown adipocytes. We found that antioxidant and antiapoptotic factors in H9c2 cardiomyocytes were upregulated by high levels of secreted FGF21 in HBAC conditioned medium (HBAC-CM), whereas FGF21 in HBAC-CM did not affect antioxidative or antiapoptotic cell death in H9c2 cardiomyocytes with Nrf2 knockdown. These results show that NRF2 mediates antioxidative and antiapoptotic effects through the HBAC-secreted factor FGF21. Consistent with this finding, the expression of antioxidant and antiapoptotic genes was upregulated by highly secreted FGF21 after HBAC-CM treatment compared to LBAC-CM treatment in H9c2 cardiomyocytes via NRF2 activation. Furthermore, HBAC-CM significantly attenuated ischemic rat heart tissue injury via NRF2 activation. Based on these findings, we propose that HBAC-CM exerts beneficial effects in rat cardiac ischemia/reperfusion injury by modulating NRF2 and has potential as a promising therapeutic agent for myocardial infarction.

Structural insight into a molecular mechanism of methenyltetrahydrofolate cyclohydrolase from Methylobacterium extorquens AM1.

Bioconversion of the C1 compounds into value-added products is one of the CO2-reducing strategies. In particular, because CO2 can be easily converted into formate, the efficient and direct bioconversion of CO2 through formate assimilation is attracting attention. The tetrahydrofolate (THF) cycle is the highly efficient reconstructed formate assimilation pathway, and 5,10-methenyltetrahydrofolate cyclohydrolase (FchA) is an essential enzyme involved in the THF cycle. In this study, a kinetic analysis of FchA from Methylobacterium extorquens AM1 (MeFchA) was performed and revealed that the enzyme has much higher cyclization than hydrolyzation activity, making it an optimal enzyme for formate assimilation. The crystal structure of MeFchA in the apo- and the THF-complexed forms was also determined, revealing that the substrate-binding site of the enzyme has three differently charged regions to stabilize the three differently charged moieties of the formyl-THF substrate. The residues involved in the substrate binding were also verified through site-directed mutagenesis. This study provides a biochemical and structural basis for the molecular mechanism underlying formate assimilation.

Rational Engineering of Homoserine O-Succinyltransferase from Escherichia coli for Reduced Feedback Inhibition by Methionine.

Methionine is an essential amino acid in all living organisms and has been used in various industrial applications such as food and feed additives. However, inhibition of enzymes involved in methionine biosynthesis is considered to be a crucial bottleneck for an efficient bio-based methionine production process. Homoserine O-succinyltransferase fromEscherichia coli (EcHST) has been reported to be feedback inhibited by the final product methionine. To understand the regulation mechanism of the enzyme and generate a feedback-resistant mutant, we determined the crystal structure of EcHST and elucidated the binding site of homoserine and succinyl-CoA. The enzyme kinetic experiments of EcHST revealed that the enzyme is noncompetitively inhibited by methionine with a Ki value of 2.44 mM, and we also identified a putative inhibitor binding site located in the vicinity of the substrate binding site. We then generated the EcHSTT242A variant with reduced feedback inhibition with a Ki value of 17.40 mM.

Cold-pressed oil from Citrus aurantifolia inhibits the proliferation of vascular smooth muscle cells via regulation of PI3K/MAPK signaling pathways.

Vascular occlusive disease is a chronic disease with significant morbidity and mortality. Although a variety of therapies and medications have been developed, the likelihood of disease re-emergence is high and this can be life-threatening. Based on a previous screening experiment related to vascular obstructive diseases using 34 types of essential oils, cold-pressed oil (CpO) from Citrus aurantifolia (lime) has been demonstrated to have the best effect for the inhibition of vascular smooth muscle cells (VSMCs) proliferation. The aim of the present study was to evaluate the effect of lime CpO on the pathological changes of VSMCs. To determine this, the effect of lime CpO on VSMC proliferation, a major cause of vascular disease, was investigated. To determine the safe concentration interval for toxicity of CpO during VSMC culture, a dilution of 1x10-5 was determined using Cell Counting Kit-8 assay, which was confirmed to be non-toxic using a lactate dehydrogenase assay. To examine the effect of lime CpO in cellular signaling pathways, changes in phosphorylation of both the PI3K/AKT/mTOR and extracellular signal-regulated MEK/ERK signaling pathways with serum were investigated. Furthermore, lime CpO with FBS also significantly decreased the expression levels of the cell cycle regulators cyclin D1 and proliferating cell nuclear antigen. Additionally, lime CpO with FBS significantly inhibited the sprouting of VSMCs in an ex vivo culture system. These results suggested that lime CpO inhibited the abnormal proliferation of VSMCs and can be developed as a nature-based therapeutic agent for obstructive vascular disease.

Temporal Variation Dominates in Local Carabid Beetle Communities in Korean Mountains.

Spatial and temporal variation in ecological environments may result in spatial and temporal variation in communities. Temporal studies of biodiversity are essential for forecasting future changes in community structure and ecosystem function. Therefore, determining the mechanisms that drive temporal change in communities remains an important and interesting challenge in ecology. We quantified spatial and temporal variations in carabid beetle communities and site-specific environmental factors for 5 years at nine study sites on three mountains in the Baekdudaegan Mountain Range, Korea. Carabid beetle communities exhibited significant temporal variation, which was larger than spatial variations between and within mountains. Environmental factors mostly varied between sites within mountains. Community variation was only weakly associated with environmental factors at wide scales, i.e., between sites on three mountains, but was strongly associated at narrow spatial scales, i.e., between sites within one mountain. Our results indicate that temporal variation in communities occurs in response to variations in the local climate, and that the patterns of temporal variation differ between mountains. Thus, temporal surveys of insect communities and climates at local scales are important for predicting temporal changes in the communities.

Crystal Structure and Functional Characterization of the Bifunctional N-(5'-Phosphoribosyl)anthranilate Isomerase-indole-3-glycerol-phosphate Synthase from Corynebacterium glutamicum.

L-Tryptophan is known as an aromatic amino acid and one of the essential amino acids that must be ingested through various additives or food. TrpCF is a bifunctional enzyme that has indole-glycerol-phosphate synthase (IGPS) and phosphoribosylanthranilate isomerase (PRAI) activity. In this report, we identified the crystal structure of TrpCF from Corynebacterium glutamicum (CgTrpCF) and successfully elucidated the active site by attaching rCdRP similar to the substrate and product of the TrpCF reaction. Also, we revealed that CgTrpCF shows a conformational change at the loops upon substrate binding. We analyzed amino acid sequences of the homologues of CgTrpCF, and the residues of the substrate-binding site in TrpCF were highly conserved except for some residues. These less conserved residues were replaced by site-directed mutagenesis experiments. Consequently, we obtained the CgTrpCFP294K (PRAICD/P294K) variant that has enhanced activity.