Legionella uses host Rab GTPases and BAP31 to create a unique ER niche.

Upon entry into host cells, the facultative intracellular bacterium Legionella pneumophila ( L.p .) uses its type IV secretion system, Dot/Icm, to secrete ~330 bacterial effector proteins into the host cell. Some of these effectors hijack endoplasmic reticulum (ER)-derived vesicles to form the Legionella -containing vacuole (LCV). Despite extensive investigation over decades, the fundamental question persists: Is the LCV membrane distinct from or contiguous with the host ER network? Here, we employ advanced photobleaching techniques, revealing a temporal acquisition of both smooth and rough ER (sER and rER) markers on the LCV. In the early stages of infection, the sER intimately associates with the LCV. Remarkably, as the infection progresses, the LCV evolves into a distinct niche comprising an rER membrane that is independent of the host ER network. We discover that the L.p. effector LidA binds to and recruits two host proteins of the Rab superfamily, Rab10, and Rab4, that play significant roles in acquiring sER and rER membranes, respectively. Additionally, we identify the pivotal role of a host ER-resident protein, BAP31, in orchestrating the transition from sER to rER. While previously recognized for shuttling between sER and rER, we demonstrate BAP31's role as a Rab effector, mediating communication between these ER sub-compartments. Furthermore, using genomic deletion strains, we uncover a novel L.p. effector, Lpg1152, essential for recruiting BAP31 to the LCV and facilitating its transition from sER to rER. Depletion of BAP31 or infection with an isogenic L.p. strain lacking Lpg1152 results in a growth defect. Collectively, our findings illuminate the intricate interplay between molecular players from both host and pathogen, elucidating how L.p. orchestrates the transformation of its residing vacuole membrane from a host-associated sER to a distinct rER membrane that is not contiguous with the host ER network.

A MAP1B-cortactin-Tks5 axis regulates TNBC invasion and tumorigenesis.

The microtubule-associated protein MAP1B has been implicated in axonal growth and brain development. We found that MAP1B is highly expressed in the most aggressive and deadliest breast cancer subtype, triple-negative breast cancer (TNBC), but not in other subtypes. Expression of MAP1B was found to be highly correlated with poor prognosis. Depletion of MAP1B in TNBC cells impairs cell migration and invasion concomitant with a defect in tumorigenesis. We found that MAP1B interacts with key components for invadopodia formation, cortactin, and Tks5, the latter of which is a PtdIns(3,4)P2-binding and scaffold protein that localizes to invadopodia. We also found that Tks5 associates with microtubules and supports the association between MAP1B and α-tubulin. In accordance with their interaction, depletion of MAP1B leads to Tks5 destabilization, leading to its degradation via the autophagic pathway. Collectively, these findings suggest that MAP1B is a convergence point of the cytoskeleton to promote malignancy in TNBC and thereby a potential diagnostic and therapeutic target for TNBC.

Effectiveness of primary series, first, and second booster vaccination of monovalent mRNA COVID-19 vaccines against symptomatic SARS-CoV-2 infections and severe diseases during the SARS-CoV-2 omicron BA.5 epidemic in Japan: vaccine effectiveness real-time surveillance for SARS-CoV-2 (VERSUS).

BACKGROUND: This study aimed to evaluate VE of primary, first, and second booster ancestral-strain monovalent mRNA COVID-19 vaccination against symptomatic infections and severe diseases in Japan. METHODS: We conducted a test-negative case-control study. We included medically attended episodes and hospitalizations involving individuals aged ≥16 with signs and symptoms from July to November 2022, when Omicron BA.5 was dominant nationwide. To evaluate VE, we calculated adjusted ORs of vaccination among test-positive versus test-negative individuals using a mixed-effects logistic regression. RESULTS: For VE against symptomatic infections among individuals aged 16 to 59, VE of primary vaccination at > 180 days was 26.1% (95% CI: 10.6-38.8%); VE of the first booster was 58.5% (48.4-66.7%) at ≤90 days, decreasing to 41.1% (29.5-50.8%) at 91 to 180 days. For individuals aged ≥60, VE of the first booster was 42.8% (1.7-66.7%) at ≤90 days, dropping to 15.4% (-25.9-43.2%) at 91 to 180 days, and then increasing to 44.0% (16.4-62.5%) after the second booster. For VE against severe diseases, VE of the first and second booster was 77.3% (61.2-86.7%) at ≤90 days and 55.9% (23.4-74.6%) afterward. CONCLUSION: mRNA booster vaccination provided moderate protection against symptomatic infections and high-level protection against severe diseases during the BA.5 epidemic in Japan.

Effectiveness of mRNA COVID-19 vaccines against symptomatic SARS-CoV-2 infections during the SARS-CoV-2 Omicron BA.1 and BA.2 epidemic in Japan: vaccine effectiveness real-time surveillance for SARS-CoV-2 (VERSUS).

BACKGROUND: Evaluating COVID-19 vaccine effectiveness (VE) domestically is crucial for assessing and determining national vaccination policy. This study aimed to evaluate VE of mRNA COVID-19 vaccines in Japan. METHODS: We conducted a multicenter test-negative case-control study. The study comprised individuals aged ≥16 visiting medical facilities with COVID-19-related signs or symptoms from 1 January to 26 June 2022, when Omicron BA.1 and BA.2 were dominant nationwide. We evaluated VE of primary and booster vaccination against symptomatic SARS-CoV-2 infections and relative VE of booster compared with primary. RESULTS: We enrolled 7,931 episodes, including 3,055 test positive. The median age was 39, 48.0% were male, and 20.5% had underlying medical conditions. In individuals aged 16 to 64, VE of primary vaccination within 90 days was 35.6% (95% CI, 19.0-48.8%). After booster, VE increased to 68.7% (60.6-75.1%). In individuals aged ≥65, VE of primary and booster was 31.2% (-44.0-67.1%) and 76.5% (46.7-89.7%), respectively. Relative VE of booster compared with primary vaccination was 52.9% (41.0-62.5%) in individuals aged 16 to 64 and 65.9% (35.7-81.9%) in individuals aged ≥65. CONCLUSIONS: During BA.1 and BA.2 epidemic in Japan, mRNA COVID-19 primary vaccination provided modest protection. Booster vaccination was necessary to protect against symptomatic infections.

Association between oral function and nutrition in gastric cancer patients undergoing chemotherapy.

OBJECTIVES: This study aimed to clarify the relationship between the oral assessment guide (OAG), a simple method for assessing oral function and poor nutrition in gastric cancer patients and investigate the reduction of oral mucositis through appropriate oral function management. SUBJECTS AND METHODS: Gastric cancer patients who underwent chemotherapy at the Nagoya Ekisaikai Hospital between January 2015 and December 2020 were evaluated. The prognostic nutritional index (PNI), as the objective variable, was used to assess nutritional status. The explanatory variables were sex, age, smoking status, body mass index (BMI), number of remaining teeth, cancer stage, albumin level, C-reactive protein level, white blood cell count and the OAG score. RESULTS: PNI was significantly associated with age, number of remaining teeth, cancer stage and the OAG score (p < 0.05) among the 217 patients. There were significant differences in age, BMI, cancer stage and the OAG score among the patients. An abnormal BMI and an advanced cancer stage were more common in older patients, and abnormal OAG scores were associated with a lower PNI. CONCLUSIONS: For gastric cancer patients undergoing postoperative chemotherapy, worse oral functional status is associated with worse PNI and nutritional status.

NANOS3 suppresses premature spermatogonial differentiation to expand progenitors and fine-tunes spermatogenesis in mice.

In the mouse testis, sperm originate from spermatogonial stem cells (SSCs). SSCs give rise to spermatogonial progenitors, which expand their population until entering the differentiation process that is precisely regulated by a fixed time-scaled program called the seminiferous cycle. Although this expansion process of progenitors is highly important, its regulatory mechanisms remain unclear. NANOS3 is an RNA-binding protein expressed in the progenitor population. We demonstrated that the conditional deletion of Nanos3 at a later embryonic stage results in the reduction of spermatogonial progenitors in the postnatal testis. This reduction was associated with the premature differentiation of progenitors. Furthermore, this premature differentiation caused seminiferous stage disagreement between adjacent spermatogenic cells, which influenced spermatogenic epithelial cycles, leading to disruption of the later differentiation pathway. Our study suggests that NANOS3 plays an important role in timing progenitor expansion to adjust to the proper differentiation timing by blocking the retinoic acid (RA) signaling pathway.

Bisphenol A-sulfate conjugate disrupts AURKA transcription and cell cycle in BeWo cytotrophoblasts.

Bisphenol A (BPA) has been shown to exhibit various toxic effects, including the induction of reproductive disorders. Generally, BPA is converted to conjugated metabolites, leading to bio-inactivation. On the other hand, the toxicity of conjugated metabolites is not fully understood. Notably, the placenta develops the sulfate-sulfatase pathway, which transports and reactivates sulfated steroids. Therefore, we investigated the potential adverse effects of the BPA-sulfate conjugate (BPA-S) on human placenta-derived BeWo cytotrophoblasts. In the present study, high-concentration BPA-S (100 µM) induced significant inhibition of BeWo growth, with effects similar to those seen with unconjugated BPA (100 µM and 100 nM). This growth inhibition was restored by treatment of the cells with an inhibitor of the organic anion-transporting peptides (OATPs) (bromosulphophthalein) or with a sulfatase (STS) inhibitor (STX64). BeWo exhibits expression of the genes encoding OATP1A2 and OATP4A1 as known sulfated steroid transporters and STS, suggesting that BPA-S suppresses cell growth activity via the sulfate-sulfatase pathway. In addition, cell cycle analysis revealed that BPA-S (100 µM) increased the fraction of cytotrophoblasts in the G2/M phases and significantly decreased the accumulation of the transcript encoding Aurora kinase A (AURKA), which is a critical regulator of cellular division. These results suggested that BPA-S triggers cell cycle arrest and inhibits proliferation of BeWo cytotrophoblasts by decreased AURKA, an effect that is mediated by the sulfate-sulfatase pathway. Overall, these findings provide insights into the reactivation of sulfated endocrine-disrupting chemicals and subsequent adverse effects.

Syntaxin 17, an ancient SNARE paralog, plays different and conserved roles in different organisms.

Mammalian syntaxin 17 (Stx17) has several roles in processes other than membrane fusion, including in mitochondrial division, autophagosome formation and lipid droplet expansion. In contrast to conventional syntaxins, Stx17 has a long C-terminal hydrophobic region with a hairpin-like structure flanked by a basic amino acid-enriched C-terminal tail. Although Stx17 is one of the six ancient SNAREs and is present in diverse eukaryotic organisms, it has been lost in multiple lineages during evolution. In the present study, we compared the localization and function of fly and nematode Stx17s expressed in HeLa cells with those of human Stx17. We found that fly Stx17 predominantly localizes to the cytosol and mediates autophagy, but not mitochondrial division. Nematode Stx17, on the other hand, is predominantly present in mitochondria and facilitates mitochondrial division, but is irrelevant to autophagy. These differences are likely due to different structures in the C-terminal tail. Non-participation of fly Stx17 and nematode Stx17 in mitochondrial division and autophagy, respectively, was demonstrated in individual organisms. Our results provide an insight into the evolution of Stx17 in metazoa. This article has an associated First Person interview with the first author of the paper.

The interaction of ATP11C-b with ezrin contributes to its polarized localization.

ATP11C, a member of the P4-ATPase family, translocates phosphatidylserine and phosphatidylethanolamine at the plasma membrane. We previously revealed that its C-terminal splice variant ATP11C-b exhibits polarized localization in motile cell lines, such as MDA-MB-231 and Ba/F3. In the present study, we found that the C-terminal cytoplasmic region of ATP11C-b interacts specifically with ezrin. Notably, the LLxY motif in the ATP11C-b C-terminal region is crucial for its interaction with ezrin as well as its polarized localization on the plasma membrane. A constitutively active, C-terminal phosphomimetic mutant of ezrin was colocalized with ATP11C-b in polarized motile cells. ATP11C-b was partially mislocalized in cells depleted of ezrin alone, and exhibited greater mislocalization in cells simultaneously depleted of the family members ezrin, radixin and moesin (ERM), suggesting that ERM proteins, particularly ezrin, contribute to the polarized localization of ATP11C-b. Furthermore, Atp11c knockout resulted in C-terminally phosphorylated ERM protein mislocalization, which was restored by exogenous expression of ATP11C-b but not ATP11C-a. These observations together indicate that the polarized localizations of ATP11C-b and the active form of ezrin to the plasma membrane are interdependently stabilized.

Chemical Characterization of a Volatile Dubnium Compound, DbOCl3.

The formation and the chemical characterization of single atoms of dubnium (Db, element 105), in the form of its volatile oxychloride, was investigated using the on-line gas phase chromatography technique, in the temperature range 350-600 °C. Under the exactly same chemical conditions, comparative studies with the lighter homologues of Group 5 in the Periodic Table clearly indicate the volatility sequence being NbOCl3 > TaOCl3 ≥ DbOCl3 . From the obtained experimental results, thermochemical data for DbOCl3 were derived. The present study delivers reliable experimental information for theoretical calculations on chemical properties of transactinides.

Improved development of mouse somatic cell nuclear transfer embryos by chlamydocin analogues, class I and IIa histone deacetylase inhibitors†.

In mammalian cloning by somatic cell nuclear transfer (SCNT), the treatment of reconstructed embryos with histone deacetylase (HDAC) inhibitors improves efficiency. So far, most of those used for SCNT are hydroxamic acid derivatives-such as trichostatin A-characterized by their broad inhibitory spectrum. Here, we examined whether mouse SCNT efficiency could be improved using chlamydocin analogues, a family of newly designed agents that specifically inhibit class I and IIa HDACs. Development of SCNT-derived embryos in vitro and in vivo revealed that four out of five chlamydocin analogues tested could promote the development of cloned embryos. The highest pup rates (7.1-7.2%) were obtained with Ky-9, similar to those achieved with trichostatin A (7.2-7.3%). Thus, inhibition of class I and/or IIa HDACs in SCNT-derived embryos is enough for significant improvements in full-term development. In mouse SCNT, the exposure of reconstructed oocytes to HDAC inhibitors is limited to 8-10 h because longer inhibition with class I inhibitors causes a two-cell developmental block. Therefore, we used Ky-29, with higher selectivity for class IIa than class I HDACs for longer treatment of SCNT-derived embryos. As expected, 24-h treatment with Ky-29 up to the two-cell stage did not induce a developmental block, but the pup rate was not improved. This suggests that the one-cell stage is a critical period for improving SCNT cloning using HDAC inhibitors. Thus, chlamydocin analogues appear promising for understanding and improving the epigenetic status of mammalian SCNT-derived embryos through their specific inhibitory effects on HDACs.

Simple chemical detection based on a surface-modified electroosmotic pump via interval immobilization.

Environmental water quality monitoring plays an important role in human health risk assessments for pharmaceuticals in water and pollutant source control. A new chemical detection method was developed to enhance molecular selectivity and portability by combining the molecularly imprinted technique and an electroosmotic pump (EOP), which requires only a small pump, batteries and stopwatch in principle. Selective chemical adsorption on the surface-modified EOP decreases the pumping performance of EOP due to a decrease in the surface electric charge. For proof of concept, the microfabricated EOPs with chemical surface treatment were used to investigate the effects of surface chemical change on pumping performance. The microfluidic EOP of a size of 20 mm × 20 mm × 1 mm was modified by an interval immobilization method using the template of 4-(tributylammonium-methyl)-benzyltributylammonium chloride (TBTA) and evaluated by measuring EOF. The pumping performance of the surface-modified EOP was decreased by the selective adsorption of TBTA to a two-point recognition site on the EOP surfaces. The relationships between the flow rate and the TBTA concentration were fitted to the Langmuir equation. The EOP can selectively detect the model substance even in a mixture solution with a different chemical compound. This molecular imprinted EOP does not require large and expensive instruments for driving the device and chemical detection, which can be applied to a portable analytical device for onsite analysis.

Introducing activities of the Archives of Infectious Diseases History (AIDH) project: Historical epidemiology.

The AIDH is a project as a historical epidemiology. The AIDH aims to collect, maintain, and manage past epidemiological materials and to offer these materials to persons who are interested in the history and in the fields of tropical medicine and global health. In this paper, we introduce our purpose and activities and show a hypothesis about lymphatic filariasis with Brugia malayi in Japan as a case of historical epidemiology. We hope to build fruitful ties between historians and scholars of tropical medicine and global health workers through an interdisciplinary approach to the history of control of infectious diseases.

The ER cholesterol sensor SCAP promotes CARTS biogenesis at ER-Golgi membrane contact sites.

In response to cholesterol deprivation, SCAP escorts SREBP transcription factors from the endoplasmic reticulum to the Golgi complex for their proteolytic activation, leading to gene expression for cholesterol synthesis and uptake. Here, we show that in cholesterol-fed cells, ER-localized SCAP interacts through Sac1 phosphatidylinositol 4-phosphate (PI4P) phosphatase with a VAP-OSBP complex, which mediates counter-transport of ER cholesterol and Golgi PI4P at ER-Golgi membrane contact sites (MCSs). SCAP knockdown inhibited the turnover of PI4P, perhaps due to a cholesterol transport defect, and altered the subcellular distribution of the VAP-OSBP complex. As in the case of perturbation of lipid transfer complexes at ER-Golgi MCSs, SCAP knockdown inhibited the biogenesis of the trans-Golgi network-derived transport carriers CARTS, which was reversed by expression of wild-type SCAP or a Golgi transport-defective mutant, but not of cholesterol sensing-defective mutants. Altogether, our findings reveal a new role for SCAP under cholesterol-fed conditions in the facilitation of CARTS biogenesis via ER-Golgi MCSs, depending on the ER cholesterol.

A Rapid Method to Measure Serum Retinol Concentrations in Japanese Black Cattle Using Multidimensional Fluorescence.

Vitamin A levels in fattening Japanese Black cattle affect meat quality; therefore, it is important to monitor serum retinol concentrations. To simplify and accelerate the evaluation of serum retinol concentrations in cattle, we developed a new predictive method using excitation-emission matrix (EEM) fluorescence spectrophotometry. For analytical comparison, the concentration of serum retinol was also measured using the conventional HPLC method. We examined excitation (Ex) and emission (Em) wavelengths of cattle serum, which were 250-450 and 250-600 nm, respectively. Parallel factor analysis separated four components from EEM data, one of which was related to retinol. Next, a partial least square regression model was created using the obtained EEMs as explanatory variables and accrual measurement values as objective variables. The determination coefficient value (R2), root mean squared error of prediction (RMSEP), and the ratio of performance to deviation (RPD) of the model were determined. A comparison with reference values found that R2, RMSEP, and RPD of the calibration model were 0.95, 6.4 IU/dl, and 4.2, respectively. This implies that EEM can estimate the serum retinol concentration with high accuracy. Additionally, the fluorescent peaks that contributed to the calibration, which were extracted from the regression coefficient and variable importance in projection plots, were Ex/Em = 320/390 and 330/520 nm. Thus, we assume that this method observes not only free retinol, but also retinol-binding protein. In conclusion, multidimensional fluorescence analysis can accurately and quickly determine serum retinol concentrations in fattening cattle.

Development of Force Field Parameters for p-Carborane to Investigate the Structural Influence of Carborane Derivatives on Drug Targets by Complex Formation.

Androgen receptor (AR) has a key role in the development and progression of prostate cancer, and AR antagonists are used for its remedy. Recently, carborane derivatives, which are carbon-containing boron clusters have attracted attention as new AR ligands. Here we determined the force field parameters of 10-vertex and 12-vertex p-carborane to facilitate in silico drug design of boron clusters. Then, molecular dynamics (MD) simulations of complexes of AR-carborane derivatives were performed to evaluate the parameters and investigate the influences of carborane derivatives on the three-dimensional structure of AR. Energy profiles were obtained using quantum chemical calculations, and the force-field parameters were determined by curve fitting of the energy profiles. The results of MD simulations indicated that binding of the antagonist-BA341 changed some hydrogen-bond formations involved in the structure and location of helix 12. Those results were consistent with previously reported data. The determined parameters are also useful for refining the structure of the carborane-receptor complex obtained by docking simulations and development of new ligands with carborane cages not only for AR but also for various receptors.

Discontinuation or reduction in benzodiazepine use by treatment with the traditional herbal medicine Hangekobokuto, case reports.

In this report, we present two cases in which benzodiazepines (BZDs) were able to be successfully reduced or discontinued by treatment with traditional Japanese herbal medicine (Kampo medicine), including Hangekobokuto (HKT). These two patients with long-term use of BZDs due to mental disorders suffered epigastric symptoms. After starting Kampo therapy including HKT, their epigastric symptoms were improved and they were able to reduce or discontinue the use of BZDs. This suggests that HKT is a potentially promising substitutive pharmacotherapy for patients with long-term use of BZDs. HKT should be considered as a treatment for patients with mental disorders who have taken BZDs for a long time and suffer from medically unexplained epigastric symptoms.

Enantioselective Organocatalytic Construction of Spirochroman Derivatives.

Highly enantioselective organocatalytic construction of spirochromans containing a tetrasubstituted stereocenter was developed. Intramolecular oxy-Michael addition was catalyzed with a bifunctional cinchona alkaloid thiourea catalyst. A variety of spirochroman compounds containing a tetrasubstituted stereocenter were obtained with excellent enantioselectivities of up to 99% enantiomeric excess. The reaction was applied to the asymmetric formal synthesis of (-)-(R)-cordiachromene.

Loss of H3K27me3 imprinting in the Sfmbt2 miRNA cluster causes enlargement of cloned mouse placentas.

Somatic cell nuclear transfer (SCNT) in mammals is an inefficient process that is frequently associated with abnormal phenotypes, especially in placentas. Recent studies demonstrated that mouse SCNT placentas completely lack histone methylation (H3K27me3)-dependent imprinting, but how it affects placental development remains unclear. Here, we provide evidence that the loss of H3K27me3 imprinting is responsible for abnormal placental enlargement and low birth rates following SCNT, through upregulation of imprinted miRNAs. When we restore the normal paternal expression of H3K27me3-dependent imprinted genes (Sfmbt2, Gab1, and Slc38a4) in SCNT placentas by maternal knockout, the placentas remain enlarged. Intriguingly, correcting the expression of clustered miRNAs within the Sfmbt2 gene ameliorates the placental phenotype. Importantly, their target genes, which are confirmed to cause SCNT-like placental histology, recover their expression level. The birth rates increase about twofold. Thus, we identify loss of H3K27me3 imprinting as an epigenetic error that compromises embryo development following SCNT.

Glucuronidation as a metabolic barrier against zearalenone in rat everted intestine.

Zearalenone (ZON), produced by Fusarium fungi, exhibits estrogenic activity. Livestock can be exposed to ZON orally through contaminating feeds such as cereals, leading to reproductive disorders such as infertility and miscarriage via endocrine system disruption. However, the details of ZON metabolism remain unclear, and the mechanism of its toxicity has not been fully elucidated. In this study, we investigated the kinetics of ZON absorption and metabolism in rat segmented everted intestines. ZON absorption was confirmed in each intestine segment 60 min after application to the mucosal buffer at 10 µM. Approximately half of the absorbed ZON was metabolized to α-zearalenol, which tended to be mainly glucuronidated in intestinal cells. In the proximal intestine, most of the glucuronide metabolized by intestinal cells was excreted to the mucosal side, suggesting that the intestine plays an important role as a first drug metabolism barrier for ZON. However, in the distal intestine, ZON metabolites tended to be transported to the serosal side. Glucuronide transported to the serosal side could be carried via the systemic circulation to the local tissues, where it could be reactivated by deconjugation. These results are important with regard to the mechanism of endocrine disruption caused by ZON.