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Abstract To the best of our knowledge, enzymes that catalyse intramolecular Diels-Alder ([4+2] cycloaddition) reactions are frequently reported in natural product biosynthesis; however, no native enzymes utilising Lewis acid catalysis have been reported. Verticilactam is a representative member of polycyclic macrolactams, presumably produced by spontaneous cycloaddition. We report that the intramolecular [4+2] cycloadditions can be significantly accelerated by ferredoxins (Fds), a class of small iron-sulphur (Fe-S) proteins. Through iron atom substitution by Lewis acidic gallium (Ga) iron and computational calculations, we confirm that the ubiquitous Fe-S cluster efficiently functions as Lewis acid to accelerate the tandem [4+2] cycloaddition and Michael addition reactions by lowering free energy barriers. Our work highlights Nature’s ingenious strategy to generate complex molecule structures using the ubiquitous Fe-S protein. Furthermore, our study sheds light on the future design of Fd as a versatile Lewis acid catalyst for [4+2] cycloaddition reactions.

Acetic acid is a common inhibitor present in lignocellulose hydrolysate, which inhibits the ethanol production by yeast strains. Therefore, the cellulosic ethanol industry requires yeast strains that can tolerate acetic acid stress. Here we demonstrate that overexpressing a yeast native arginase-encoding gene, CAR1, renders Saccharomyces cerevisiae acetic acid tolerance. Specifically, ethanol yield increased by 27.3% in the CAR1-overexpressing strain compared to the control strain under 5.0 g/L acetic acid stress. The global intracellular amino acid level and compositions were further analyzed, and we found that CAR1 overexpression reduced the total amino acid content in response to acetic acid stress. Moreover, the CAR1 overexpressing strain showed increased ATP level and improved cell membrane integrity. Notably, we demonstrated that the effect of CAR1 overexpression was independent of the spermidine and proline metabolism, which indicates novel mechanisms for enhancing yeast stress tolerance. Our studies also suggest that CAR1 is a novel genetic element to be used in synthetic biology of yeast for efficient production of fuel ethanol.

The arginine transporter Can1 is a multifunctional protein of the conventional yeast Saccharomyces cerevisiae. Apart from facilitating arginine uptake, Can1 plays a pivotal role in regulating proline metabolism and maintaining cellular redox balance. Here, we report a novel function of Can1 in the control of yeast biofilm formation. First, the S. cerevisiae CAN1 gene knockout strain displayed a significant growth delay compared to the wild-type strain. Our genetic screening revealed that the slow growth of the CAN1 knockout strain is rescued by a functional deficiency of the FLO8 gene, which encodes the master transcription factor associated with biofilm formation, indicating that Can1 is involved in biofilm formation. Intriguingly, the CAN1 knockout strain promoted the Flo11-dependent aggregation, leading to higher biofilm formation. Furthermore, the CAN1 knockout strain of the pathogenic yeast Candida glabrata exhibited slower growth and higher biofilm formation, similar to S. cerevisiae. More importantly, the C. glabrata CAN1 gene knockout strain showed severe toxicity to macrophage-like cells and nematodes. The present results could help to elucidate both the molecular mechanism underlying yeast biofilm formation and the role it plays. Future investigations may offer insights that contribute to development of antibiofilm agents.

Summary: Transient anoxia causes amnesia and neuronal death. This is attributed to enhanced glutamate release and modeled as anoxia-induced long-term potentiation (aLTP). aLTP is mediated by glutamate receptors and nitric oxide (·NO) and occludes stimulation-induced LTP. We identified a signaling cascade downstream of ·NO leading to glutamate release and a glutamate-·NO loop regeneratively boosting aLTP. aLTP in entothelial ·NO synthase (eNOS)-knockout mice and blocking neuronal NOS (nNOS) activity suggested that both nNOS and eNOS contribute to aLTP. Immunostaining result showed that eNOS is predominantly expressed in vascular endothelia. Transient anoxia induced a long-lasting Ca2+ elevation in astrocytes that mirrored aLTP. Blocking astrocyte metabolism or depletion of the NMDA receptor ligand D-serine abolished eNOS-dependent aLTP, suggesting that astrocytic Ca2+ elevation stimulates D-serine release from endfeet to endothelia, thereby releasing ·NO synthesized by eNOS. Thus, the neuro-glial-endothelial axis is involved in long-term enhancement of glutamate release after transient anoxia.

Supersulfides, which are defined as sulfur species with catenated sulfur atoms, are increasingly being investigated in biology. We recently identified pyridoxal phosphate (PLP)-dependent biosynthesis of cysteine persulfide (CysSSH) and related supersulfides by cysteinyl-tRNA synthetase (CARS). Here, we investigated the physiological role of CysSSH in budding yeast (Saccharomyces cerevisiae) by generating a PLP-binding site mutation K109A in CRS1 (the yeast ortholog of CARS), which decreased the synthesis of CysSSH and related supersulfides and also led to reduced chronological aging, effects that were associated with an increased endoplasmic reticulum stress response and impaired mitochondrial bioenergetics. Reduced chronological aging in the K109A mutant could be rescued by using exogenous supersulfide donors. Our findings indicate important roles for CARS in the production and metabolism of supersulfides—to mediate mitochondrial function and to regulate longevity.

BackgroundIn recent years, technologies promoting the digitization of self-monitoring of blood glucose (SMBG) records including app-cloud cooperation systems have emerged. Studies combining these technological interventions with support from remote health care professionals have reported improvements in glycemic control. ObjectiveTo assess the use of an app-cloud cooperation system linked with SMBG devices in clinical settings, we evaluated its effects on outpatient management of diabetes without remote health care professional support. MethodsIn this multicenter, open-label, and single-armed prospective study, 48 patients with diabetes (including type 1 and type 2) at 3 hospitals in Japan treated with insulin or glucagon-like peptide 1 receptor agonists and performing SMBG used the app-cloud cooperation system for 24 weeks. The SMBG data were automatically uploaded to the cloud via the app. The patients could check their data, and their attending physicians reviewed the data through the cloud prior to the patients’ regular visits. The primary outcome was changes in glycated hemoglobin (HbA1c) levels. ResultsAlthough HbA1c levels did not significantly change in all patients, the frequency of daily SMBG following applying the system was significantly increased before induction at 12 (0.60 per day, 95% CI 0.19-1.00; P=.002) and 24 weeks (0.43 per day, 95% CI 0.02-0.84; P=.04). In the subset of 21 patients whose antidiabetic medication had not been adjusted during the intervention period, a decrease in HbA1c level was observed at 12 weeks (P=.02); however, this significant change disappeared at 24 weeks (P=.49). The Diabetes Treatment Satisfaction Questionnaire total score and “Q4: convenience” and “Q5: flexibility” scores significantly improved after using the system (all P

Abstract Breeding yeast strains for industrial alcoholic fermentation requires laborious screening due to the lack of in vivo modification strategies. Here we show that quiescence-specific cell wall thickening via synthesis of a major component, 1,3-β-glucan, critically antagonizes cellular fermentation ability by sequestering the available cytoplasmic carbon sources. This study provides insights into glycolytic control and reports an effective and reliable rational fermentation design.

Abstract The Greenhouse gases Observing SATellite 2 (GOSAT-2) was launched in October 2018 as a successor to GOSAT (launched in 2009), the first satellite to specialize in greenhouse gas observations. Compared to the GOSAT sensors, the sensors of GOSAT-2 offer higher performance in most respects. The quality and quantity of data from observations are expected to be improved accordingly. The signal-to-noise ratio (SNR) is better in both the SWIR and TIR bands of TANSO-FTS-2, which is the main sensor of GOSAT-2. This improvement ultimately enhances the accuracy of greenhouse gas concentration analysis. Furthermore, because of the improved SNR in the SWIR band, the northern limit at which data are obtainable in high-latitude regions of the Northern Hemisphere in winter, where observation data have remained unavailable because of weak signal strength, has moved to higher latitudes. As better data are obtained in greater quantities, progress in carbon cycle research for high-latitude regions is anticipated. Moreover, the improvement of SNR in the TIR band is expected to be considerable. Particularly, the resolutions of the vertical concentration distributions of CO2 and CH4 have been improved drastically. The first function introduced for GOSAT-2 that is not in GOSAT is an intelligent pointing mechanism: a cloud area avoidance function using the in-field camera of TANSO-FTS-2. This function can increase the amounts of observation data globally and can improve the accuracy of CO2 emissions estimation and measurements of uptake intensity. The effects are expected to be strong, especially for the tropics because cumulus clouds are the most common cloud type. The intelligent pointing system can avoid the clouds effectively. Another important benefit of TANSO-FTS-2 is that the wavelength range of Band 3 of SWIR has been expanded for measuring carbon monoxide (CO). Because CO originates from combustion, it is used to evaluate some effects of human activities in urban areas and biomass burning in fields. Particularly, black carbon-type aerosols can be measured by the sub-sensor, TANSO-CAI-2, to assess biomass burning along with CO2 and CO by TANSO-FTS-2.

The baking industry is experiencing significant growth, primarily due to the widespread adoption of frozen dough baking. However, this process can negatively impact the fermentation ability of yeast, as freezing can induce stress in yeast cells. This study reports the molecular interplay between the ubiquitin–proteasome system and freezing stress tolerance in the yeast Saccharomyces cerevisiae. Using the proteasome inhibitor MG132, we first screened mutants with enhanced freezing stress tolerance. Three mutants showed elevated activity of the intracellular proteasome, particularly trypsin-like activity (more than threefold) and reduced sensitivity to MG132 inhibition of chymotrypsin-like activity (less than 0.125-fold). Genomic analysis of these mutants revealed mutations in the ROX1 gene, a heme-dependent repressor of hypoxic genes. Importantly, the ROX1 deletion strain displayed slightly improved freezing stress tolerance (about 1.5-fold). Comprehensive transcription analysis identified the ANB1 gene as a potential downstream target of Rox1. Overexpression of ANB1 enhanced freezing stress tolerance (about 1.5-fold) with increased the proteasome’s activity, indicating that Rox1 contributes to changes in the proteasome’s activity and freezing stress tolerance through the function of Anb1. The present data provide new insights into the mechanisms of freezing stress tolerance and help us improve the baking of frozen dough to produce higher-quality bread.

The efforts of mangrove plantations are not always successful, and the reasons for their failure remain unclear. In this study, the growth of planted mangroves (Kandelia sp.) was monitored in a mangrove forest on Amami Island, Japan, to investigate their survival over the first three years. A strong typhoon passed near the island 16 months after planting. Although the mangroves were planted at the frontal edge of the intertidal zone, most of young mangroves survived without visible damage. The external forces acting on the plants, such as wave forces, storm surge forces, wind drag forces, and oscillatory acceleration, were estimated by applying an integrated analysis model for tropical cyclones, storm surges, wind waves, and plant kinematics. These analyses yielded three main results. First, although large waves over 10 m in height were generated offshore, the mangrove forest was located in the innermost part of the bay, resulting in considerable wave attenuation. Second, the wave period in the bay was much shorter than that in the open ocean, which may have promoted resonance between waves and mangrove plants. The oscillatory acceleration of the young mangroves was estimated to have approximately the same magnitude as that of the gravitational force. Third, the young mangroves could withstand a fairly strong typhoon, but this may have been fortuitous because the timing of the adverse wind wave conditions coincided with low tide. It is estimated that if the tides had been higher, the oscillatory acceleration would have been five times that of the gravitational acceleration. Although this study only presents examples of specific mangrove species in a particular location, the findings can be used to improve the attempts to restore mangrove forests and thereby protect coastal settlements from the adverse effects of tropical cyclones.

Abstract Objective: Resting energy expenditure (REE) decreases if there is reduced energy intake and body weight (BW). The decrease in REE could make it difficult for patients with obesity to maintain decreased BW. This study aimed to investigate the correlation among changes in REE, energy intake, and BW during the weight loss process in patients with obesity. Materials and methods: We conducted a retrospective cohort study of patients hospitalized for the treatment of obesity in Japan. Patients received fully controlled diet during hospitalization and performed exercises if able. REE was measured once a week using a hand-held indirect calorimetry. Energy intake was determined by actual dietary intake. Results: Of 44 inpatients with obesity, 17 were included in the analysis. Their BW decreased significantly after 1 week (−4.7 ± 2.0 kg, P < 0.001) and 2 weeks (−5.7 ± 2.2 kg, P < 0.001). The change in REE after 1 and 2 weeks was positively correlated with the energy intake/energy expenditure ratio (r = 0.66, P = 0.004 at 1 week, r = 0.71, P = 0.002 at 2 weeks). Using a regression equation (y = 0.5257x – 43.579), if the energy intake/energy expenditure ratio within the second week was 82.9%, the REE after 2 weeks was similar to the baseline level. There was no significant correlation between the change in REE and BW. Conclusions: Our data suggest that changes in REE depend on energy intake/energy expenditure ratio and that the decrease in REE can be minimized by matching energy intake to energy expenditure, even during the weight loss process.

Nitric oxide (NO) is a natural reactive nitrogen species (RNS) that alters proteins, DNA, and lipids and damages biological activities. Although microorganisms respond to and detoxify NO, the regulation of the cellular metabolic mechanisms that cause cells to tolerate RNS toxicity is not completely understood. We found that the proline and arginine auxotrophic proA5 and argB2 mutants of the fungus Aspergillus nidulans require more arginine and proline for normal growth under RNS stress that starves cells by accumulating fewer amino acids. Fungal transcriptomes indicated that RNS stress upregulates the expression of the biosynthetic genes required for global amino acids, including proline and arginine. A mutant of the gene disruptant, cpcA, which encodes the transcriptional regulation of the cross-pathway control of general amino acid synthesis, did not induce these genes, and cells accumulated fewer amino acids under RNS stress. These results indicated a novel function of CpcA in the cellular response to RNS stress, which is mediated through amino acid starvation and induces the transcription of genes for general amino acid synthesis. Since CpcA also controls organic acid biosynthesis, impaired intermediates of such biosynthesis might starve cells of amino acids. These findings revealed the importance of the mechanism regulating amino acid homeostasis for fungal responses to and survival under RNS stress.

Abstract Familial neurohypophyseal diabetes insipidus (FNDI) is a degenerative disease of vasopressin (AVP) neurons. Studies in mouse in vivo models indicate that accumulation of mutant AVP prehormone is associated with FNDI pathology. However, studying human FNDI pathology in vivo is technically challenging. Therefore, an in vitro human model needs to be developed. When exogenous signals are minimized in the early phase of differentiation in vitro, mouse embryonic stem cells (ESCs)/induced pluripotent stem cells (iPSCs) differentiate into AVP neurons, whereas human ESCs/iPSCs die. Human ESCs/iPSCs are generally more similar to mouse epiblast stem cells (mEpiSCs) compared to mouse ESCs. In this study, we converted human FNDI-specific iPSCs by the naive conversion kit. Although the conversion was partial, we found improved cell survival under minimal exogenous signals and differentiation into rostral hypothalamic organoids. Overall, this method provides a simple and straightforward differentiation direction, which may improve the efficiency of hypothalamic differentiation.

Abstract Fungi are ubiquitously present in our living environment and are responsible for crop and infectious diseases. Developing new antifungal agents is constantly needed for their effective control. Here, we investigated fungal cellular responses to an array of antifungal compounds, including plant- and bacteria-derived antifungal compounds. The pathogenic fungus Aspergillus fumigatus generated reactive oxygen species in its hyphae after exposure to the antifungal compounds thymol, farnesol, citral, nerol, salicylic acid, phenazine-1-carbonic acid, and pyocyanin, as well as under oxidative and high-temperature stress conditions. The production of nitric oxide (NO) was determined using diaminofluorescein-FM diacetate (DAF-FM DA) and occurred in response to antifungal compounds and stress conditions. The application of reactive oxygen species or NO scavengers partly suppressed the inhibitory effects of farnesol on germination. However, NO production was not detected in the hyphae using the Greiss method. An LC/MS analysis also failed to detect DAF-FM-T, a theoretical product derived from DAF-FM DA and NO, in the hyphae after antifungal treatments. Thus, the cellular state after exposure to antifungal agents may be more complex than previously believed, and the role of NO in fungal cells needs to be investigated further.

Metallothionein (MT), which is a small metal-binding protein with cysteine-rich motifs, functions in the detoxification of heavy metals in a variety of organisms. Even though previous studies suggest that MT is in-volved in the tolerance mechanisms against nitrosative stress induced by toxic levels of nitric oxide (NO) in mammalian cells, the physiological functions of MT in relation to NO have not been fully understood. In this study, we analyzed the functions of MT in nitrosative stress tolerance in the yeast Saccharomyces cerevisiae. Our phenotypic analyses showed that deletion or overexpression of the MT-encoding gene, CUP1, led to higher sensitivity or tolerance to nitrosative stress in S. cerevisiae cells, respectively. We further examined whether the yeast MT Cup1 in the cell-free lysate scavenges NO. These results showed that the cell-free lysate containing a higher level of Cup1 degraded NO more efficiently. On the other hand, the transcription level of CUP1 was not affected by nitrosative stress treatment. Our findings suggest that the yeast MT Cup1 contributes to nitrosative stress tolerance, possibly as a constitutive rather than an inducible defense mechanism.

Background: Cases with varus knee osteoarthritis (OA) often present concomitant ankle joint deformity and poor tibial plafond inclination (TPI) in the coronal plane. Although changes to the TPI with respect to the ground after open wedge high tibial osteotomy (OWHTO) have been described, reports on the relationship between the correction angle and severity of knee joint line obliquity (KJLO) are scarce. This study aimed to examine the correlation between the correction angle after OWHTO and the change in knee and ankle inclination with respect to the floor. Methods: Between January 2016 and December 2019, 39 knees in 39 patients (mean age, 58.3 ± 9.7 years; male, 14 knees; female, 25 knees) underwent OWHTO for varus knee OA. The mean duration of follow up was 30.1 ± 9.8 months. Radiological severity of OA was evaluated for those with Kellgren-Lawrence grade 1–3. Preoperative and 1-year postoperative radiographs were used to examine the postoperative femorotibial angle, hip-knee angle, KJLO, TPI, talar tilt (TT), position of weight bearing line of the tibial plateau width (WBL ratio; %), and the relationship between the pre-to postoperative changes in each measured value and intraoperative correction angle. The relationship between the pre-to postoperative changes in each measured value and intraoperative correction angle were evaluated. Clinical evaluations were conducted before and 2 years after OWHTO using the Knee Injury and Osteoarthritis Outcome Score (KOOS) subscale of pain. Results: The mean pre-to post-operative changes were 3.2 ± 2.2 for KJLO and 3.1 ± 2.5 for TPI, both demonstrating significant differences as well as significant correlations with the correction angle. The knee joint demonstrated postoperative changes in the valgus direction and none in the varus direction with respect to the floor in all cases. Furthermore, the sum of changes to the KJLO and TPI showed a positive correlation that approximates the change in the correction angle. The mean pre- and postoperative 2-year KOOS subscale of pain were 50.2 ± 10.2 and 82.3 ± 9.1. There were no correlations between preoperative TPI, pre-to postoperative change in TPI, and 2-year postoperative of KOOS subscale of pain. Conclusion: For those who underwent OWHTO for varus OA, the knee joint changed from a preoperative varus malalignment to mild valgus malalignment, and the ankle joint tended to improve from a preoperative valgus malalignment to the postoperative neutral alignment with respect to the floor. Although a positive correlation was found in which the correction angle approximates the sum of changes to the KJLO and TPI, no significant difference was found between changes in KJLO and TPI nor in the ratio between the correction angle and changes in each measured value.

A cell population characterized by the release of glucose repression and known as [GAR+] emerges spontaneously in the yeast Saccharomyces cerevisiae. This study revealed that the [GAR+] variants exhibit retarded alcoholic fermentation when glucose is the sole carbon source. To identify the key to the altered glucose response, the gene expression profile of [GAR+] cells was examined. Based on RNA-seq data, the [GAR+] status was linked to impaired function of the Cyc8p–Tup1p complex. Loss of Cyc8p led to a decrease in the initial rate of alcoholic fermentation under glucose-rich conditions via the inactivation of pyruvate decarboxylase, an enzyme unique to alcoholic fermentation. These results suggest that Cyc8p can become inactive to attenuate alcoholic fermentation. These findings may contribute to the elucidation of the mechanism of non-genetic heterogeneity in yeast alcoholic fermentation.

Although proline is the most or second most abundant amino acid in wort and grape must, it is not fully consumed by the yeast Saccharomyces cerevisiae during alcoholic fermentation, unlike other amino acids. Our previous studies showed that arginine, the third most abundant amino acid in wort, inhibits the utilization of proline in most strains of S. cerevisiae. Furthermore, we found that some non-Saccharomyces yeasts utilized proline in a specific artificial medium with arginine and proline as the only nitrogen source, but these yeasts were not suitable for beer fermentation due to their low alcohol productivity. For yeasts to be useful for brewing, they need to utilize proline and produce alcohol during fermentation. In this study, 11 S. cerevisiae strains and 10 non-Saccharomyces yeast strains in the Phaff Yeast Culture Collection were identified that utilize proline effectively. Notably, two of these S. cerevisiae strains, UCDFST 40-144 and 68-44, utilize proline and produce sufficient alcohol in the beer fermentation model used. These strains have the potential to create distinctive beer products that are specifically alcoholic but with a reduction in proline in the finished beer.

Abstract Protein tyrosine nitration (PTN), in which tyrosine (Tyr) residues on proteins are converted into 3-nitrotyrosine (NT), is one of the post-translational modifications mediated by reactive nitrogen species (RNS). Many recent studies have reported that PTN contributed to signaling systems by altering the structures and/or functions of proteins. This study aimed to investigate connections between PTN and the inhibitory effect of nitrite-derived RNS on fermentation ability using the yeast Saccharomyces cerevisiae. The results indicated that RNS inhibited the ethanol production of yeast cells with increased intracellular pyruvate content. We also found that RNS decreased the activities of pyruvate decarboxylase (PDC) as a critical enzyme involved in ethanol production. Our proteomic analysis revealed that the main PDC isozyme Pdc1 underwent the PTN modification at Tyr38, Tyr157, and Tyr344. The biochemical analysis using the recombinant purified Pdc1 enzyme indicated that PTN at Tyr157 or Tyr344 significantly reduced the Pdc1 activity. Interestingly, the substitution of Tyr157 or Tyr344 to phenylalanine, which is no longer converted into NT, recovered the ethanol production under the RNS treatment conditions. These findings suggest that nitrite impairs the fermentation ability of yeast by inhibiting the Pdc1 activity via its PTN modification at Tyr157 and Tyr344 of Pdc1.

We investigated whether peripheral combination treatment of a sodium–glucose cotransporter 2 (SGLT2) inhibitor and leptin improves glucose metabolism in insulin-dependent diabetes mellitus (IDDM) model mice. Twelve-week-old male C57BL6 mice were intraperitoneally administered a high dose of streptozotocin to produce IDDM. IDDM mice were then divided into five groups: SGLT2 inhibitor treatment alone, leptin treatment alone, leptin and SGLT2 inhibitor co-treatment, untreated IDDM mice, and healthy mice groups. The blood glucose (BG) level at the end of the dark cycle was measured, and a glucose tolerance test (GTT) was performed and compared between the five groups. Leptin was peripherally administered at 20 μg/day using an osmotic pump, and an SGLT2 inhibitor, ipragliflozin, was orally administered at 3 mg/kg/day. Monotherapy with SGLT2 inhibitor or leptin significantly improved glucose metabolism in mice as evaluated by BG and GTT compared with the untreated group, whereas the co-treatment group with SGLT2 inhibitor and leptin further improved glucose metabolism as compared with the monotherapy group. Notably, glucose metabolism in the co-treatment group improved to the same level as that in the healthy mice group. Thus, peripheral combination treatment with leptin and SGLT2 inhibitor improved glucose metabolism in IDDM mice without the use of insulin.

Abstract Previous studies suggest that signaling by the gamma-aminobutyric acid (GABA) type B receptor (GABABR) is involved in the regulation of binge eating, a disorder which might contribute to the development of obesity. Here, we show that intermittent access to a high fat diet (HFD) induced binge-like eating behavior with activation of dopamine receptor d1 (drd1)-expressing neurons in the caudate putamen (CPu) and nucleus accumbens (NAc) in wild-type (WT) mice. The activation of drd1-expressing neurons during binge-like eating was substantially increased in the CPu, but not in the NAc, in corticostriatal neuron-specific GABABR-deficient knockout (KO) mice compared to WT mice. Treatment with the GABABR agonist, baclofen, suppressed binge-like eating behavior in WT mice, but not in KO mice, as reported previously. Baclofen also suppressed the activation of drd1-expressing neurons in the CPu, but not in the NAc, during binge-like eating in WT mice. Thus, our data suggest that GABABR signaling in CPu neurons expressing drd1 suppresses binge-like consumption during a HFD in mice.

Abstract This study presents a new storm surge hazard potential index (SSHPI) for estimating tropical cyclone (TC) induced peak surge levels at a coast. The SSHPI incorporates parameters that are often readily available at real-time: intensity in 10-min maximum wind speed, radius of 50-kt wind, translation speed, coastal geometry, and bathymetry information. The inclusion of translation speed and coastal geometry information lead to improvements of the SSHPI to other existing surge indices. A retrospective analysis of SSHPI using data from 1978–2019 in Japan suggests that this index captures historical events reasonably well. In particular, it explains ~ 66% of the observed variance and ~ 74% for those induced by TCs whose landfall intensity was larger than 79-kt. The performance of SSHPI is not sensitive to the type of coastal geometry (open coasts or semi-enclosed bays). Such a prediction methodology can decrease numerical computation requirements, improve public awareness of surge hazards, and may also be useful for communicating surge risk.

ABSTRACT The unambiguous application of fungal names is important to communicate scientific findings. Names are critical for (clinical) diagnostics, legal compliance, and regulatory controls, such as biosafety, food security, quarantine regulations, and industrial applications. Consequently, the stability of the taxonomic system and the traceability of nomenclatural changes is crucial for a broad range of users and taxonomists. The unambiguous application of names is assured by the preservation of nomenclatural history and the physical organisms representing a name. Fungi are extremely diverse in terms of ecology, lifestyle, and methods of study. Predominantly unicellular fungi known as yeasts are usually investigated as living cultures. Methods to characterize yeasts include physiological (growth) tests and experiments to induce a sexual morph; both methods require viable cultures. Thus, the preservation and availability of viable reference cultures are important, and cultures representing reference material are cited in species descriptions. Historical surveys revealed drawbacks and inconsistencies between past practices and modern requirements as stated in the International Code of Nomenclature for Algae, Fungi, and Plants (ICNafp). Improper typification of yeasts is a common problem, resulting in a large number invalid yeast species names. With this opinion letter, we address the problem that culturable microorganisms, notably some fungi and algae, require specific provisions under the ICNafp. We use yeasts as a prominent example of fungi known from cultures. But viable type material is important not only for yeasts, but also for other cultivable Fungi that are characterized by particular morphological structures (a specific type of spores), growth properties, and secondary metabolites. We summarize potential proposals which, in our opinion, will improve the stability of fungal names, in particular by protecting those names for which the reference material can be traced back to the original isolate.

Abstract The reward system, which consists of dopaminergic neurons projecting from the ventral tegmental area (VTA) to the nucleus accumbens and caudate-putamen in the striatum, has an important role in the pathogenesis of not only drug addiction but also diet-induced obesity. In the present study, we examined whether signaling through glucocorticoid receptors (GRs) in the reward system affects the rewarding value of a high-fat diet (HFD). To do so, we generated mice that lack functional GRs specifically in dopaminergic neurons (D-KO mice) or corticostriatal neurons (CS-KO mice), subjected the mice to caloric restriction stress conditions, and evaluated the rewarding value of a HFD by conditioned place preference (CPP) test. Caloric restriction induced increases in serum corticosterone to similar levels in all genotypes. While CS-KO as well as WT mice exhibited a significant preference for HFD in the CPP test, D-KO mice exhibited no such preference. There were no differences between WT and D-KO mice in consumption of HFD after fasting or cognitive function evaluated by a novel object recognition test. These data suggest that glucocorticoid signaling in the VTA increases the rewarding value of a HFD under restricted caloric stress.

ABSTRACT In the yeast Saccharomyces cerevisiae, N-acetyl glutamate kinase (NAGK), which catalyzes the phosphorylation of N-acetyl glutamate to form N-acetyl glutamyl-5-phosphate, is one of the rate-limiting enzymes in the ornithine and arginine biosynthetic pathways. NAGK activity is strictly regulated via feedback inhibition by the end product, arginine. We previously reported that the Thr340Ile variant of NAGK was insensitive to arginine feedback inhibition and that the interaction between Lys336 and Thr340 in NAGK may be important for arginine recognition. In the present study, we demonstrated that amino acid changes of Thr340 to Ala, Leu, Arg, Glu, Ile, and Asn removed arginine feedback inhibition, although the Thr340Ser variant was subject to the feedback inhibition. Therefore, these results indicate that the arginine-binding cavity formed via the interaction between the carbonyl group in the main chain of Lys336 and the hydroxyl group in the side chain of the residue at position 340 is critical for arginine recognition of NAGK. In addition, we newly identified two mutations in the ARG5,6 gene encoding the Cys119Tyr or Val267Ala variant of NAGK of sake yeast mutants with intracellular ornithine accumulation. Although it is unlikely that Cys119 and Val267 are directly involved in arginine recognition, we found here that two variants of NAGK were insensitive to arginine feedback inhibition and contributed to high-level production of ornithine. Structural analysis of NAGK suggests that these two amino acid substitutions influence the sensitivity to Arg feedback inhibition through alterations in local conformation around each residue. IMPORTANCE Ornithine has a number of physiological benefits in humans. Thus, an Orn-rich alcoholic beverage is expected to relieve feelings of fatigue after drinking. In the yeast Saccharomyces cerevisiae, N-acetyl glutamate kinase (NAGK) encoded by the ARG5,6 gene catalyzes the second step in ornithine and arginine biosynthesis, and its activity is subjected to feedback inhibition by arginine. Here, we revealed a role of key residues in the formation of the arginine-binding cavity which is critical for arginine recognition of NAGK. In addition, we analyzed novel arginine feedback inhibition-insensitive variants of NAGK in sake yeast mutants with ornithine overproduction and proposed that the amino acid substitutions in the NAGK variants destabilize the arginine-binding cavity, leading to the lower sensitivity to arginine feedback inhibition of NAGK activity. These findings provide new insight into the allosteric regulation of NAGK activity and will help to construct superior industrial yeast strains for high-level production of ornithine.

Saccharomyces cerevisiae is a facultative anaerobic organism that grows well under both aerobic and hypoxic conditions in media containing abundant fermentable nutrients such as glucose. In order to deeply understand the physiological dependence of S. cerevisiae on aeration, we checked endoplasmic reticulum (ER)-stress status by monitoring the splicing of HAC1 mRNA, which is promoted by the ER stress-sensor protein, Ire1. HAC1-mRNA splicing that was caused by conventional ER-stressing agents, including low concentrations of dithiothreitol (DTT), was more potent in hypoxic cultures than in aerated cultures. Moreover, growth retardation was observed by adding low-dose DTT into hypoxic cultures of ire1∆ cells. Unexpectedly, aeration mitigated ER stress and DTT-induced impairment of ER oxidative protein folding even when mitochondrial respiration was halted by the ro mutation. An ER-located protein Ero1 is known to directly consume molecular oxygen to initiate the ER protein oxidation cascade, which promotes oxidative protein folding of ER client proteins. Our further study using ero1-mutant strains suggested that, in addition to mitochondrial respiration, this Ero1-medaited reaction contributes to mitigation of ER stress by molecular oxygen. Taken together, here we demonstrate a scenario in which aeration acts beneficially on S. cerevisiae cells even under fermentative conditions.

Reactive nitrogen species (RNS) are signal molecules involved in various biological events; however, excess levels of RNS cause nitrosative stress, leading to cell death and/or cellular dysfunction. During the process of infection, pathogens are exposed to nitrosative stress induced by host-derived RNS. Therefore, the nitrosative stress resistance mechanisms of pathogenic microorganisms are important for their infection and pathogenicity, and could be promising targets for antibiotics. Previously, we demonstrated that the RIB1 gene encoding GTP cyclohydrolase II (GCH2), which catalyzes the first step of the riboflavin biosynthesis pathway, is important for nitrosative stress resistance in the yeast Saccharomyces cerevisiae. Here, we identified and characterized the RIB1 gene in the opportunistic pathogenic yeast Candida glabrata. Our genetic and biochemical analyses indicated that the open reading frame of CAGL0F04279g functions as RIB1 in C. glabrata (CgRIB1). Subsequently, we analyzed the effect of CgRIB1 on nitrosative stress resistance by a growth test in the presence of RNS. Overexpression or deletion of CgRIB1 increased or decreased the nitrosative stress resistance of C. glabrata, respectively, indicating that GCH2 confers nitrosative stress resistance on yeast cells. Moreover, we showed that the proliferation of C. glabrata in cultures of macrophage-like cells required the GCH2-dependent nitrosative stress detoxifying mechanism. Additionally, an infection assay using silkworms as model host organisms indicated that CgRIB1 is indispensable for the virulence of C. glabrata. Our findings suggest that the GCH2-dependent nitrosative stress detoxifying mechanism is a promising target for the development of novel antibiotics.

Introduction: Recently, some studies showed assessment of the component gap is important for determination of the implant-inserted condition during total knee arthroplasty (TKA). We perform the modified gap technique with adjustment of the virtual gap which estimated by computer-aided design (CAD) using navigation system. The purpose of this study was to compare the virtual gap (CAD-gap) with the actual gap after inserting a femoral trial component (Trial-gap), and examine the usefulness of the surgical technique. Materials and methods: The subjects were 35 patients who underwent primary TKA using a navigation system and posterior-stabilized type TKA. The surgical procedure was to produce an extension gap, confirm the flexed CAD-gap on the navigation screen based on CAD data, and plan osteotomy of the femur. After osteotomy, the femoral component was inserted and the gap balance was measured. A tensor was used to adjust and measure the gap balance. Initial alignment, rotation of the femoral component, soft tissue balance in extension, final alignment after fixing all components, and the CAD- and Trial-gaps in both extension and flexion were evaluated. Results: The mean initial alignment angle, rotation angle of the femoral component, soft tissue balance angle and final alignment angle were 8.1 ± 4.2° varus, 3.5 ± 1.3°external rotation, 2.7 ± 2.5° varus and 0.4 ± 1.4° varus respectively. The mean medial and lateral CAD-gaps in extension were 10.8 ± 2.5 and 13.7 ± 2.5 mm, and the mean medial and lateral CAD-gaps in flexion were 12.2 ± 2.2 and 13.9 ± 2.7 mm. The equivalent Trial-gaps in extension and flexion were 10.5 ± 2.6 and 11.4 ± 3.1 mm, and 12.2 ± 2.5 and 14.4 ± 2.8 mm. The CAD- and Trial-gaps differed significantly only for lateral gaps in extension. Conclusion: In comparing the CAD-gap and the Trial-gap, only small difference was found in the lateral gap of extension. The other gaps in both extension and flexion were well maintained. We concluded adjustment of the CAD-gap during surgery using a navigation system can be used to adjust the actual component gap especially in the medial side.

Summary: Binge eating could contribute to the development of obesity, and previous studies suggest that gamma-aminobutyric acid (GABA) type B receptor (GABABR) signaling is involved in the regulation of binge eating. Here, we show that time-restricted access to a high-fat diet (HFD) induces binge-like eating behavior in wild-type mice. HFD consumption during restricted time was significantly increased in corticostriatal neuron-specific GABABR-deficient mice compared with wild-type mice. Furthermore, the GABABR agonist baclofen suppressed HFD intake during restricted time in wild-type mice but not in corticostriatal or dopaminergic neuron-specific GABABR-deficient mice. In contrast, there were no significant differences in food consumption among genotypes under ad libitum access to HFD. Thus, our data show that the mesolimbic system regulates food consumption under time-restricted but not ad libitum access to HFD and have identified a mechanism by which GABABR signaling suppresses binge-like eating of HFD. : Animal Physiology; Neuroscience; Behavioral Neuroscience Subject Areas: Animal Physiology, Neuroscience, Behavioral Neuroscience

Binge eating is a characteristic symptom observed in obese individuals that is related to dysfunction of dopaminergic neurons (DNs). Intermittent administration of a high-fat diet (HFD) is reported to induce binge-like eating, but the underlying mechanisms remain unclear. We generated dopaminergic neuron specific IKKβ deficient mice (KO) to examine the effects of inflammation in DNs on binge-like eating under inflammatory conditions associated with HFD. After administration of HFD for 4 weeks, mice were fasted for 24 h, and then the consumption of HFD was measured for 2 h. We also evaluated that the mRNA expressions of inflammatory cytokines, glial markers, and dopamine signaling-related genes in the ventral tegmental area (VTA) and striatum. Moreover, insulin was administered intraventricularly to assess downstream signaling. The consumption of HFD was significantly reduced, and the phosphorylation of AKT in the VTA was significantly increased in female KO compared to wild-type (WT) mice. Analyses of mRNA expressions revealed that DNs activity and inflammation in the VTA were significantly decreased in female KO mice. Thus, our data suggest that HFD-induced inflammation with glial cell activation in the VTA affects DNs function and causes abnormal eating behaviors accompanied by insulin resistance in the VTA of female mice.

Background: Lateral discoid meniscus (LDM) should be treated and preserved with saucerization and/or suture repair. However, repair of the meniscal hoop structure is sometimes difficult due to displacement or large defects. In this study, we aimed to examine tear patterns based on the Ahn classification in those requiring meniscal repair and those undergoing subtotal meniscectomy. Methods: Twenty-three patients were evaluated (mean age, 27.4 years; mean follow-up period, 2.5 years). The following were evaluated: displacement morphology based on the Ahn classification, site of tear under arthroscopy, morphology, surgical procedure, Lysholm score at final postoperative follow-up, and clinical outcome of meniscus using Barrett’s criteria. Result: There were 16 knees without displacement (saucerization with suture repair, 13 knees; subtotal meniscectomy, 3 knees) and 10 knees with displacement (reduction with suture repair, 3 knees; subtotal meniscectomy, 7 knees). Subtotal meniscectomy was performed more often in cases with dislocation, especially in the central shift type as defined by the Ahn classification. The mean Lysholm score was 65.0 points preoperatively and 95.3 points postoperatively. Twenty-three knees (88%) were postoperatively categorized under the Barrett’s criteria as healing and 3 knees (12%) were categorized as non-healing. The number of non-healing cases that underwent subtotal meniscectomy was relatively small (1 of 10 knees), and the short-term results were not poor. Conclusion: Localized peripheral longitudinal tears tended to be repairable even with displacement, while peripheral tears covering the entire meniscus or with severe defects/tears in the body of the meniscus tended to be difficult to repair, leading to subtotal meniscectomy.

Background Pituitary dysfunction is a life-threatening immune-related adverse event (irAE) induced by immune checkpoint inhibitors (ICIs). To date, it is not possible to identify patients who may develop pituitary irAEs prior to ICI treatment. The aim of this study was to characterize the predisposition for ICI-induced pituitary irAEs by analyzing anti-pituitary antibodies (APAs) and human leukocyte antigens (HLAs).Methods In this case–control study, APAs and HLA alleles were analyzed in 62 patients (17 who developed ICI-induced isolated adrenocorticotropic hormone deficiency (ICI-IAD), 5 who developed ICI-induced hypophysitis (ICI-H) and 40 who did not develop pituitary irAEs) treated with ICIs between November 2, 2015, and March 31, 2020, at Nagoya University Hospital. The main outcome measures in this study were the association between the development of pituitary irAEs with APAs at baseline and after treatment and HLA alleles.Results Eleven of 17 (64.7%) patients who developed ICI-IAD had APAs at baseline, whereas APAs were positive only in 1 of 40 (2.5%) control patients. Although APAs were negative at baseline in all patients who developed ICI-H, they had become positive before the onset of ICI-H in 3 of 4 patients several weeks after ipilimumab administration. At the onset of ICI-IAD and ICI-H, APAs were positive in 15 of 17 (88.2%) and 4 of 5 (80%) patients, respectively. The prevalence of HLA-Cw12, HLA-DR15, HLA-DQ7, and HLA-DPw9 was significantly higher in patients with ICI-IAD, whereas that of HLA-Cw12 and HLA-DR15 was significantly higher in patients with ICI-H than in controls.Conclusions This study showed distinct and overlapped patterns of APAs and HLA alleles between ICI-IAD and ICI-H. Our findings also showed that positive APAs at baseline and after treatment, together with susceptible HLA alleles, could become predictive biomarkers for ICI-IAD and ICI-H, respectively.Trial registration number UMIN000019024.

In many plants and microorganisms, intracellular proline has a protective role against various stresses, including heat-shock, oxidation and osmolarity. Environmental stresses induce cellular senescence in a variety of eukaryotes. Here we showed that intracellular proline regulates the replicative lifespan in the budding yeast Saccharomyces cerevisiae. Deletion of the proline oxidase gene PUT1 and expression of the γ-glutamate kinase mutant gene PRO1-I150T that is less sensitive to feedback inhibition accumulated proline and extended the replicative lifespan of yeast cells. Inversely, disruption of the proline biosynthetic genes PRO1, PRO2, and CAR2 decreased stationary proline level and shortened the lifespan of yeast cells. Quadruple disruption of the proline transporter genes unexpectedly did not change intracellular proline levels and replicative lifespan. Overexpression of the stress-responsive transcription activator gene MSN2 reduced intracellular proline levels by inducing the expression of PUT1, resulting in a short lifespan. Thus, the intracellular proline levels at stationary phase was positively correlated with the replicative lifespan. Furthermore, multivariate analysis of amino acids in yeast mutants deficient in proline metabolism showed characteristic metabolic profiles coincident with longevity: acidic and basic amino acids and branched-chain amino acids positively contributed to the replicative lifespan. These results allude to proline metabolism having a physiological role in maintaining the lifespan of yeast cells.

Abstract Background The ureides allantoin and allantoate are major metabolic intermediates of purine catabolism with high nitrogen-to-carbon ratios. Ureides play a key role in nitrogen utilization in ureide-type legumes, but their effects on growth and development in non-legume plants are poorly understood. Here, we examined the effects of knocking out genes encoding ureide-degrading enzymes, allantoinase (ALN) and allantoate amidohydrolase (AAH), on the vegetative-to-reproductive transition and subsequent growth of Arabidopsis plants. Results The ureide-degradation mutants (aln and aah) showed symptoms similar to those of nitrogen deficiency: early flowering, reduced size at maturity, and decreased fertility. Consistent with these phenotypes, carbon-to-nitrogen ratios and nitrogen-use efficiencies were significantly decreased in ureide-degradation mutants; however, adding nitrogen to irrigation water did not alleviate the reduced growth of these mutants. In addition to nitrogen status, levels of indole-3-acetic acid and gibberellin in five-week-old plants were also affected by the aln mutations. To test the possibility that ureides are remobilized from source to sink organs, we measured ureide levels in various organs. In wild-type plants, allantoate accumulated predominantly in inflorescence stems and siliques; this accumulation was augmented by disruption of its catabolism. Mutants lacking ureide transporters, ureide permeases 1 and 2 (UPS1 and UPS2), exhibited phenotypes similar to those of the ureide-degradation mutants, but had decreased allantoate levels in the reproductive organs. Transcript analysis in wild-type plants suggested that genes involved in allantoate synthesis and ureide transport were coordinately upregulated in senescing leaves. Conclusions This study demonstrates that ureide degradation plays an important role in supporting healthy growth and development in non-legume Arabidopsis during and after transition from vegetative to reproductive stages.

Excessive sodium salt (NaCl) or fat intake is associated with a variety of increased health risks. However, whether excessive NaCl intake accompanied by a high-fat diet (HFD) affects glucose metabolism has not been elucidated. In this study, C57BL/6J male mice were fed a normal chow diet (NCD), a NCD plus high-NaCl diet (NCD plus NaCl), a HFD, or a HFD plus high-NaCl diet (HFD plus NaCl) for 30 weeks. No significant differences in body weight gain, insulin sensitivity, and glucose tolerance were observed between NCD-fed and NCD plus NaCl-fed mice. In contrast, body and liver weights were decreased, but the weight of epididymal white adipose tissue was increased in HFD plus NaCl-fed compared to HFD-fed mice. HFD plus NaCl-fed mice had lower plasma glucose levels in an insulin tolerance test, and showed higher plasma glucose and lower plasma insulin levels in an intraperitoneal glucose tolerance test compared to HFD-fed mice. The β-cell area and number of islets were decreased in HFD plus NaCl-fed compared to HFD-fed mice. Increased Ki67-positive β-cells, and increased expression levels of Ki67, CyclinB1, and CyclinD1 mRNA in islets were observed in HFD-fed but not HFD plus NaCl-fed mice when compared to NCD-fed mice. Our data suggest that excessive NaCl intake accompanied by a HFD exacerbates glucose intolerance, with impairment in insulin secretion caused by the attenuation of expansion of β-cell mass in the pancreas.

This is a rare case of a patient with a double-layered lateral meniscus, undergoing surgical treatment. A 17-year-old woman who was a member of a volleyball club had a two-year history of right knee pain with episodes of locking, although she had no history of trauma. She was referred to our hospital because her condition did not improve after conservative treatment. On presentation to the hospital, she had full range of motion in the right knee. McMurray’s test revealed no clicks; however, it produced pain in the lateral part. Plain radiography revealed no abnormal findings, whereas magnetic resonance imaging showed high signal in the posterior segment of the lateral meniscus and an increase in its volume. Arthroscopic findings showed an accessory meniscus with a flat surface overlying a normal-sized lateral meniscus. It was firmly connected to the posterior root and middle segment of the lower normal meniscus. The accessory meniscus was markedly mobile as revealed by probing. The patient was diagnosed with double-layered lateral meniscus and underwent resection of the accessory meniscus. Postoperatively, she initiated strengthening of muscles and range of motion training without weight-bearing restrictions. Two months postoperatively, she had completely recovered and participated in volleyball practices. In the last follow-up at 18 months, she had no restrictions in daily or sports activities.

Four disease-specific induced pluripotent stem cell (iPSC) lines were respectively derived from peripheral blood mononuclear cells of two affected individuals in a family affected by familial neurohypophyseal diabetes insipidus carrying the c.314G>C mutation. The expression of pluripotency markers (NANOG, OCT4, and SOX2), maintenance of a normal karyotype, absence of episomal vectors used for iPSC generation, and presence of the original pathogenic mutation were confirmed for each iPSC line. The ability to differentiate into three germ layers was confirmed by a teratoma formation assay. These iPSC lines can help in disease recapitulation in vitro using organoids and elucidation of disease mechanisms.

Summary: Misfolded or unfolded proteins in the ER are said to be degraded only after translocation or isolation from the ER. Here, we describe a mechanism by which mutant proteins are degraded within the ER. Aggregates of mutant arginine vasopressin (AVP) precursor were confined to ER-associated compartments (ERACs) connected to the ER in AVP neurons of a mouse model of familial neurohypophysial diabetes insipidus. The ERACs were enclosed by membranes, an ER chaperone and marker protein of phagophores and autophagosomes were expressed around the aggregates, and lysosomes fused with the ERACs. Moreover, lysosome-related molecules were present within the ERACs, and aggregate degradation within the ERACs was dependent on autophagic-lysosomal activity. Thus, we demonstrate that protein aggregates can be degraded by autophagic-lysosomal machinery within specialized compartments of the ER.

Background Several immune-related adverse events (irAEs) are reported to be associated with therapeutic efficacy of immune checkpoint inhibitors, yet whether pituitary dysfunction, a life-threatening irAE, affects overall survival (OS) in patients with malignancies is unclear. This prospective study examined the association of pituitary dysfunction (pituitary-irAE) with OS of patients with non-small cell lung carcinoma (NSCLC) or malignant melanoma (MM).Methods A total of 174 patients (NSCLC, 108; MM, 66) treated with ipilimumab, nivolumab, pembrolizumab, or atezolizumab at Nagoya University Hospital were evaluated for OS and the development of pituitary-irAE. Kaplan-Meier curves of OS as a function of the development of pituitary-irAE were produced with the log-rank test as a primary endpoint.Results Pituitary-irAE was observed in 16 patients (4 (3.7%) with NSCLC, 12 (18.2%) with MM) having two different disease types: hypophysitis with deficiency of multiple anterior pituitary hormones accompanied by pituitary enlargement, and isolated adrenocorticotropic hormone (ACTH) deficiency without pituitary enlargement. Among these patients, 6 developed pituitary-irAE while being treated with ipilimumab (6/25 patients (24.0%) treated with ipilimumab) and 10 developed pituitary-irAE during treatment with nivolumab or pembrolizumab (10/167 (6.0%)). All 16 patients had ACTH deficiency and were treated with physiological doses of hydrocortisone. The development of pituitary-irAE was associated with better OS in patients with NSCLC (not reached vs 441 (95% CI not calculated) days, p

Overexpression of MSN2, which is the transcription factor gene in response to stress, is well-known to increase the tolerance of the yeast Saccharomyces cerevisiae cells to a wide variety of environmental stresses. Recent studies have found that the Msn2 is a feasible potential mediator of proline homeostasis in yeast. This result is based on the finding that overexpression of the MSN2 gene exacerbates the cytotoxicity of yeast to various amino acid analogs whose uptake is increased by the active amino acid permeases localized on the plasma membrane as a result of a dysfunctional deubiquitination process. Increased understanding of the cellular responses induced by the Msn2-mediated proline incorporation will provide better comprehension of how cells respond to and counteract to different kinds of stimuli and will also contribute to the breeding of industrial yeast strains with increased productivity.

Type 2 diabetes and dyslipidemia are diseases that collectively increase the risk of patients developing cardiovascular complications. Several incretin-based drugs are reported to improve lipid metabolism, and one of these medications, anagliptin, is a dipeptidyl peptidase-4 (DPP-4) inhibitor that has been shown to decrease serum triglyceride and low-density lipoproteins cholesterol. This study aimed to conduct an investigation into the effects of anagliptin on serum lipid profiles. This multicenter, open-label, randomized (1:1), parallel group study was designed to evaluate the effects of anagliptin on serum lipid profiles (triglycerides, lipoproteins, apolipoproteins, and cholesterol fractions). The study involved 24 patients with type 2 diabetes at two participating hospitals for a period of 24 weeks. Patients were randomly assigned to the anagliptin (n = 12) or control (n = 12) groups. Patients in the anagliptin group were treated with 200 mg of the drug twice daily. Patients in the control group did not receive anagliptin, but continued with their previous treatment schedules. Lipid metabolism was examined under fasting conditions at baseline and 24 weeks. Patients treated with anagliptin for 24 weeks exhibited significantly reduced levels of serum apolipoprotein B-48, a marker for lipid transport from the intestine, compared with the control group patients (P < 0.05). After 24 weeks of treatment, serum adiponectin levels were significantly raised, whereas glycated hemoglobin (HbA1c) levels were significantly lower compared with the baseline in the anagliptin group (P < 0.05), but not in the control group. This study showed that the DPP-4 inhibitor anagliptin reduces fasting apolipoprotein B-48 levels, suggesting that this drug may have beneficial effects on lipid metabolism possibly mediated by the inhibition of intestinal lipid transport.

Summary: The pituitary is a major hormone center that secretes systemic hormones responding to hypothalamus-derived-releasing hormones. Previously, we reported the independent pituitary induction and hypothalamic differentiation of human embryonic stem cells (ESCs). Here, a functional hypothalamic-pituitary unit is generated using human induced pluripotent stem (iPS) cells in vitro. The adrenocorticotropic hormone (ACTH) secretion capacity of the induced pituitary reached a comparable level to that of adult mouse pituitary because of the simultaneous maturation with hypothalamic neurons within the same aggregates. Corticotropin-releasing hormone (CRH) from the hypothalamic area regulates ACTH cells similarly to our hypothalamic-pituitary axis. Our induced hypothalamic-pituitary units respond to environmental hypoglycemic condition in vitro, which mimics a life-threatening situation in vivo, through the CRH-ACTH pathway, and succeed in increasing ACTH secretion. Thus, we generated powerful hybrid organoids by recapitulating hypothalamic-pituitary development, showing autonomous maturation on the basis of interactions between developing tissues. : Kasai et al. generate a functional hypothalamic-pituitary unit using human iPS cells. The ACTH secretion capacity of induced pituitary cells reaches the same level as adult mice anterior pituitary cells. The hypothalamic-pituitary unit responds to environmental stimulation such as low glucose through the CRH-ACTH pathway. Keywords: pituitary, hypothalamus, human iPS cells, 3D culture, self-organization, interaction, organoid, functional unit

Mangrove forests are expected to act as green infrastructure against coastal hazards. However, mangroves cannot grow along urban coasts where the water is too deep and the wave is too high. This paper conceptualizes a new disaster countermeasure, coined as “adaptive mangrove hybrid platform,” which provides a solution to achieve ecosystem-based disaster risk reduction (Eco-DRR) in urban areas; particularly, those in developing countries. The proposed concept is a hybrid solution, starting from a landfill to level the seabed. A mangrove plantation is then established on the surface. The front part of the landfill is intentionally raised to form a shore-protection mound to reduce wave run-up that may wash away mangrove seedlings. Improvised technology such as wood piles and small stones can also be utilized to protect the landfill from erosion. The platform will inevitably sink owing to self-weight consolidation, land subsidence, and sea-level rise, while sediment deposition trapped by the developed mangrove forest may be the reliable avenue to raise the ground. Hence, the initial geometry of the landfill needs to be appropriately designed in order to achieve an equilibrium state whereby the ground surface is adaptively in balance with the relative sea-level rise. In this paper, many challenges as well as advantages are addressed, with the objective of promoting research to facilitate the exploration of adaptive mangrove hybrid platforms. Keywords: Mangrove, Coastal hazards, Urban areas, Land subsidence, Sea-level rise, Eco-DRR, Green infrastructure

High differentiation efficiency is one of the most important factors in developing an in vitro model from pluripotent stem cells. In this report, we improved the handling technique applied to mouse-induced pluripotent stem (iPS) cells, resulting in better differentiation into hypothalamic vasopressin (AVP) neurons. We modified the culture procedure to make the maintenance of iPS cells in an undifferentiated state much easier. Three-dimensional floating culture was demonstrated to be effective for mouse iPS cells. We also improved the differentiation method with regards to embryology, resulting in a greater number of bigger colonies of AVP neurons differentiating from mouse iPS cells. Fgf8, which was not used in the original differentiation method, increased iPS differentiation into AVP neurons. These refinements will be useful as a valuable tool for the modeling of degenerative disease in AVP neurons in vitro using disease-specific iPS cells in future studies. Keywords: AVP neuron, Hypothalamus, Mouse iPS cells, Differentiation, Maintenance

In the yeast Saccharomyces cerevisiae, the branched-chain amino acid aminotransferases (BCATs) Bat1 and Bat2 catalyze the conversion of α-ketoisovalerate, α-keto-β-methylvalerate, and α-ketoisokaproate and into valine, isoleucine, and leucine, respectively, as the final step of branched-chain amino acid biosynthesis. Bat1 and Bat2 are homologous proteins that share 77% identity, but Bat1 localizes in the mitochondria and Bat2 in the cytosol. Based on our preliminary finding that only disruption of the BAT1 gene led to slow-growth phenotype, we hypothesized that Bat1 and Bat2 play distinct roles in valine biosynthesis and the regulation of cell growth. In this study, we found that intracellular valine content was dramatically decreased in Δbat1 cells, whereas Δbat2 cells exhibited no changes in the valine level. To further examine the distinct roles of Bat1 and Bat2, we constructed two artificial genes encoding the mitochondrial-targeting signal (MTS)-deleted Bat1 (Bat1-MTS) and the MTS of Bat1-fused Bat2 (Bat2+MTS). Interestingly, Bat2+MTS was relocalized into the mitochondria, because Bat2 localization was changed to the mitochondria by addition of MTS, and could partially restore the valine content and growth in Δbat1Δbat2 cells. These results suggest that the mitochondria are the major site of valine biosynthesis, and mitochondrial BCAT is important for valine biosynthesis in S. cerevisiae.

Awamori, the traditional distilled alcoholic beverage of Okinawa, Japan, is brewed with the yeast Saccharomyces cerevisiae. During the distillation process after the fermentation, enormous quantities of distillation residues containing yeast cells must be disposed of, and this has recently been recognized as a major problem both environmentally and economically. Proline, a multifunctional amino acid, has the highest water retention capacity among amino acids. Therefore, distillation residues with large amounts of proline could be useful in cosmetics. Here, we isolated a yeast mutant with high levels of intracellular proline and found a missense mutation (Gln79His) on the PRO1 gene encoding the γ-glutamyl kinase Pro1, a limiting enzyme in proline biosynthesis. The amino acid change of Gln79 to His in Pro1 resulted in desensitization to the proline-mediated feedback inhibition of GK activity, leading to the accumulation of proline in cells. Biochemical and in silico analyses showed that the amino acid residue at position 79 is involved in the stabilization of the proline binding pocket in Pro1 via a hydrogen-bonding network, which plays an important role in feedback inhibition. Our current study, therefore, proposed a possible mechanism underlying the feedback inhibition of γ-glutamyl kinase activity. This mechanism can be applied to construct proline-accumulating yeast strains to effectively utilize distillation residues.

Proline is a pivotal and multifunctional amino acid that is used not only as a nitrogen source but also as a stress protectant and energy source. Therefore, proline metabolism is known to be important in maintaining cellular homeostasis. Here, we discovered that proline oxidation, catalyzed by the proline oxidase Put1, a mitochondrial flavin-dependent enzyme converting proline into ∆1-pyrroline-5-carboxylate, controls the chronological lifespan of the yeast Saccharomyces cerevisiae. Intriguingly, the yeast strain with PUT1 deletion showed a reduced chronological lifespan compared with the wild-type strain. The addition of proline to the culture medium significantly increased the longevity of wild-type cells but not that of PUT1-deleted cells. We next found that induction of the transcriptional factor Put3-dependent PUT1 and degradation of proline occur during the aging of yeast cells. Additionally, the lifespan of the PUT3-deleted strain, which is deficient in PUT1 induction, was shorter than that of the wild-type strain. More importantly, the oxidation of proline by Put1 helped maintain the mitochondrial membrane potential and ATP production through the aging period. These results indicate that mitochondrial energy metabolism is maintained through oxidative degradation of proline and that this process is important in regulating the longevity of yeast cells.

Abstract Background Many epidemiological studies on the health effects of air pollutants have been carried out in regions with major sources such as factories and automobiles. However, the health effects of air pollutants in regions without major sources remain unclear. This study investigated the acute effects of ambient air pollution on pulmonary function among healthy students in an isolated island without major artificial sources of air pollutants. Methods A panel study was conducted of 43 healthy subjects who attended a school in an isolated island in the Seto Inland Sea, Japan. We measured the forced expiratory volume in 1 s (FEV1) and peak expiratory flow (PEF) every morning for about 1 month in May 2014. Ambient concentrations of particulate matter ≤ 2.5 μm in diameter (PM2.5), particulate matter between 2.5 and 10 μm in diameter (PM10-2.5), black carbon (BC), ozone (O3), and nitrogen dioxide (NO2) were measured. The associations between the concentrations of air pollutants and pulmonary function were analyzed using mixed-effects models. Results A decrease in FEV1 was significantly associated with BC concentrations (−27.28 mL [95%confidence interval (CI):−54.10,−0.46] for an interquartile range (IQR) increase of 0.23 μg/m3). The decrease in PEF was significantly associated with indoor O3 concentrations (−8.03 L/min [95% CI:−13.02,−3.03] for an IQR increase of 11 ppb). Among subjects with a history of allergy, an increase in PM2.5 concentrations was significantly associated with low FEV1. In subjects with a history of asthma, an inverse association between the indoor O3 concentration and pulmonary function was observed. Conclusions Our results demonstrate that increases in BC and O3 concentrations have acute effects on the pulmonary function among students in an isolated island without major artificial sources of air pollutants.