Exendin-4 Increases Scavenger Receptor Class BI Expression via Activation of AMPK/FoxO1 in Human Vascular Endothelial Cells.

Glucagon-like peptide-1 receptor agonist (GLP-1RA) has been clinically proven to protect endothelial function. Previously, we demonstrated that endothelial NO synthase (eNOS) was activated by high-density lipoprotein (HDL) via its scavenger receptor of the B class/human homologue of SR-BI, CD36 and LIMPII analogous-1(hSR-BI/CLA-1). Here, we investigated the effect of GLP-1RA and exendin-4 on the expression of hSR-BI/CLA-1 in HUVECs. Our results confirmed that GLP-1R was expressed in HUVECs by PCR and exendin-4 significantly enhanced HDL-induced eNOS activation. Next, exendin-4 increased the expression of hSR-BI/CLA-1 and a blockade of GLP-1R cancelled this effect. Further, the hSR-BI/CLA-1 transcriptional activity was enhanced by exendin-4, which was diminished by the inhibition of AMPK or dominant-negative AMPK-α-subunit. Moreover, AMPK was phosphorylated by the activation of GLP-1R. Next, ChIP assay demonstrated that exendin-4 increased the FoxO1-binding in the hSR-BI/CLA-1 promoter by upregulation of FoxO1. Mutation of FoxO1-binding or silencing of FoxO1 cancelled the effect of exendin-4 on hSR-BI/CLA-1 expression. Exendin-4 reduced FoxO1 phosphorylation and induced its nuclear accumulation, while this effect was altered by the blocking of GLP-1R or inhibition of AMPK pathway. In summary, our results proved that exendin-4 increased hSR-BI/CLA-1 expression via the AMPK/FoxO1 pathway to activate eNOS, providing a basic mechanism underlining the protective effect of GLP-1RA on endothelial function.

Comparative Physiology and Genomics of Hydrogen-Producing Vibrios.

The Hyf-type formate hydrogen lyase (FHL) complex was first proposed based on sequence comparisons in Escherichia coli in 1997 (Andrews et al. in Microbiology 143:3633-3647, 1997). The hydrogenase in the Hyf-type FHL was estimated to be a proton-translocating energy-conserving [NiFe]-hydrogenase. Although the structure of FHL is similar to that of complex I, silent gene expression in E. coli has caused delays in unveiling the genetic and biochemical features of the FHL. The entire set of genes required for Hyf-type FHL synthesis has also been found in the genome sequences of Vibrio tritonius in 2015 (Matsumura et al. in Int J Hydrog Energy 40:9137-9146, 2015), which produces more hydrogen (H2) than E. coli. Here we investigate the physiological characteristics, genome comparisons, and gene expressions to elucidate the genetic backgrounds of Hyf-type FHL, and how Hyf-type FHL correlates with the higher H2 production of V. tritonius. Physiological comparisons among the seven H2-producing vibrios reveal that V. porteresiae and V. tritonius, grouped in the Porteresiae clade, show greater capacity for H2 production than the other species. The structures of FHL-Hyp gene clusters were closely related in both Porteresiae species, but differed from those of the other species with the presence of hupE, a possible nickel permease gene. Interestingly, deeper genome comparisons revealed the co-presence of nickel ABC transporter genes (nik) with the Hyf-type FHL gene only on the genome of the Porteresiae clade species. Therefore, active primary Ni transport might be one of the key factors characterizing higher H2 production in V. tritonius. Furthermore, the expression of FHL gene cluster was significantly up-regulated in V. tritonius cells stimulated with formate, indicating that formate is likely to be a control factor for the gene expression of V. tritonius FHL in a similar way to the formate regulon encoding the E. coli FHL.

A highly photostable and bright green fluorescent protein.

The low photostability of fluorescent proteins is a limiting factor in many applications of fluorescence microscopy. Here we present StayGold, a green fluorescent protein (GFP) derived from the jellyfish Cytaeis uchidae. StayGold is over one order of magnitude more photostable than any currently available fluorescent protein and has a cellular brightness similar to mNeonGreen. We used StayGold to image the dynamics of the endoplasmic reticulum (ER) with high spatiotemporal resolution over several minutes using structured illumination microscopy (SIM) and observed substantially less photobleaching than with a GFP variant optimized for stability in the ER. Using StayGold fusions and SIM, we also imaged the dynamics of mitochondrial fusion and fission and mapped the viral spike proteins in fixed cells infected with severe acute respiratory syndrome coronavirus 2. As StayGold is a dimer, we created a tandem dimer version that allowed us to observe the dynamics of microtubules and the excitatory post-synaptic density in neurons. StayGold will substantially reduce the limitations imposed by photobleaching, especially in live cell or volumetric imaging.

Nasal delivery of single-domain antibody improves symptoms of SARS-CoV-2 infection in an animal model.

The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) that causes the disease COVID-19 can lead to serious symptoms, such as severe pneumonia, in the elderly and those with underlying medical conditions. While vaccines are now available, they do not work for everyone and therapeutic drugs are still needed, particularly for treating life-threatening conditions. Here, we showed nasal delivery of a new, unmodified camelid single-domain antibody (VHH), termed K-874A, effectively inhibited SARS-CoV-2 titers in infected lungs of Syrian hamsters without causing weight loss and cytokine induction. In vitro studies demonstrated that K-874A neutralized SARS-CoV-2 in both VeroE6/TMPRSS2 and human lung-derived alveolar organoid cells. Unlike other drug candidates, K-874A blocks viral membrane fusion rather than viral attachment. Cryo-electron microscopy revealed K-874A bound between the receptor binding domain and N-terminal domain of the virus S protein. Further, infected cells treated with K-874A produced fewer virus progeny that were less infective. We propose that direct administration of K-874A to the lung could be a new treatment for preventing the reinfection of amplified virus in COVID-19 patients.

Understanding the NaCl-dependent behavior of hydrogen production of a marine bacterium, Vibrio tritonius.

Biohydrogen is one of the most suitable clean energy sources for sustaining a fossil fuel independent society. The use of both land and ocean bioresources as feedstocks show great potential in maximizing biohydrogen production, but sodium ion is one of the main obstacles in efficient bacterial biohydrogen production. Vibrio tritonius strain AM2 can perform efficient hydrogen production with a molar yield of 1.7 mol H2/mol mannitol, which corresponds to 85% theoretical molar yield of H2 production, under saline conditions. With a view to maximizing the hydrogen production using marine biomass, it is important to accumulate knowledge on the effects of salts on the hydrogen production kinetics. Here, we show the kinetics in batch hydrogen production of V. tritonius strain AM2 to investigate the response to various NaCl concentrations. The modified Han-Levenspiel model reveals that salt inhibition in hydrogen production using V. tritonius starts precisely at the point where 10.2 g/L of NaCl is added, and is critically inhibited at 46 g/L. NaCl concentration greatly affects the substrate consumption which in turn affects both growth and hydrogen production. The NaCl-dependent behavior of fermentative hydrogen production of V. tritonius compared to that of Escherichia coli JCM 1649 reveals the marine-adapted fermentative hydrogen production system in V. tritonius. V. tritonius AM2 is capable of producing hydrogen from seaweed carbohydrate under a wide range of NaCl concentrations (5 to 46 g/L). The optimal salt concentration producing the highest levels of hydrogen, optimal substrate consumption and highest molar hydrogen yield is at 10 g/L NaCl (1.0% (w/v)).

A novel P2X4 receptor-selective antagonist produces anti-allodynic effect in a mouse model of herpetic pain.

Accumulating evidence indicates that purinergic P2X4 receptors (P2X4R: cation channels activated by extracellular ATP) expressed in spinal microglia are crucial for pathological chronic pain caused by nerve damage, suggesting a potential target for drug discovery. We identified NP-1815-PX (5-[3-(5-thioxo-4H-[1,2,4]oxadiazol-3-yl)phenyl]-1H-naphtho[1, 2-b][1,4]diazepine-2,4(3H,5H)-dione) as a novel antagonist selective for P2X4R with high potency and selectivity compared with other P2XR subtypes. In in vivo assay for acute and chronic pain, intrathecal administration of NP-1815-PX produced an anti-allodynic effect in mice with traumatic nerve damage without affecting acute nociceptive pain and motor function (although its oral administration did not produce the effect). Furthermore, in a mouse model of herpetic pain, P2X4R upregulation in the spinal cord exclusively occurred in microglia, and intrathecal NP-1815-PX suppressed induction of mechanical allodynia. This model also showed K(+)/Cl(-) cotransporter 2 (KCC2) downregulation, which is implicated in dorsal horn neuron hyperexcitability; this downregulation was restored by intrathecal treatment with NP-1815-PX or by interfering with brain-derived neurotrophic factor (BDNF) signaling, a P2X4R-activated microglial factor implicated in KCC2 downregulation. Taken together, the newly developed P2X4R antagonist NP-1815-PX produces anti-allodynic effects in chronic pain models without altering acute pain sensitivity, suggesting that microglial P2X4R could be an attractive target for treating chronic pain.

Complete Genome Sequence of Moraxella osloensis Strain KMC41, a Producer of 4-Methyl-3-Hexenoic Acid, a Major Malodor Compound in Laundry.

We report the complete genome sequence of Moraxella osloensis strain KMC41, isolated from laundry with malodor. The KMC41 genome comprises a 2,445,556-bp chromosome and three plasmids. A fatty acid desaturase and at least four ß-oxidation-related genes putatively associated with 4-methyl-3-hexenoic acid generation were detected in the KMC41 chromosome.

Corrigendum: Updating the Vibrio clades defined by multilocus sequence phylogeny: proposal of eight new clades, and the description of Vibrio tritonius sp. nov.

[This corrects the article on p. 414 in vol. 4, PMID: 24409173.].

Interferon regulatory factor 8 expressed in microglia contributes to tactile allodynia induced by repeated cold stress in rodents.

We investigated the role of interferon regulatory factor 8 (IRF8) in a model of chronic pain in which repeated cold stress (RCS) exposure produces tactile allodynia. RCS exposure produced a decrease in paw withdrawal threshold (PWT) to mechanical stimulation. Spinal microglia of RCS-exposed mice were morphologically activated. Expression of IRF8 was significantly increased in the spinal cord of RCS-exposed mice and was localized in microglia. IRF8-knockout mice failed to show the RCS-induced decrease in PWT. Thus, RCS exposure activates spinal microglia and upregulation of IRF8 in these cells is involved in the development of tactile allodynia after RCS exposure.

Zunongwangia mangrovi sp. nov., isolated from mangrove (Avicennia marina) rhizosphere, and emended description of the genus Zunongwangia.

A Gram-stain-negative, strictly aerobic, slightly halophilic, non-motile and rod-shaped bacterial strain, designated P2E16(T), was isolated from mangrove (Avicennia marina) rhizosphere, collected at Devipattinam mangroves, Tamil Nadu, India. Strain P2E16(T) grew optimally at pH 7.0-8.0, at 25-28 °C and in the presence of 2-3% (w/v) NaCl. 16S rRNA gene analysis showed that strain P2E16(T) was phylogenetically closely related to the genus Zunongwangia, with Zunongwangia profunda SM-A87(T) as the closest related type strain (98.2% 16S rRNA gene sequence similarity) and less than 93% 16S rRNA gene sequence similarity to all other members of the family Flavobacteriaceae. Strain P2E16(T) contained MK-6 as the major respiratory quinone, phosphatidylethanolamine as the predominant polar lipid and iso-C(15 : 0) (17.8%), iso-C(17 : 0) 3-OH (15.1%), C(15 : 0) (12.8%), iso-C(17 : 1)ω9c (9.8%), iso-C(15 : 1) G (9.0%), and summed feature 3 (comprising C(16 : 1)ω7c and/or iso-C(15 : 0) 2-OH; 7.1%) as the major fatty acids. The DNA G+C content was 34.3 mol%. Differential phenotypic properties, together with the phylogenetic distinctiveness and low DNA-DNA relatedness demonstrated that strain P2E16(T) was distinct from the type strain of Zunongwangia profunda. On the basis of these presented data, strain P2E16(T) is considered to represent a novel species of the genus Zunongwangia, for which the name Zunongwangia mangrovi sp. nov. is proposed. The type strain is P2E16(T) ( = DSM 24499(T) = LMG 26237(T) = KCTC 23496(T)). An emended description of the genus Zunongwangia is also provided.

Updating the Vibrio clades defined by multilocus sequence phylogeny: proposal of eight new clades, and the description of Vibrio tritonius sp. nov.

To date 142 species have been described in the Vibrionaceae family of bacteria, classified into seven genera; Aliivibrio, Echinimonas, Enterovibrio, Grimontia, Photobacterium, Salinivibrio and Vibrio. As vibrios are widespread in marine environments and show versatile metabolisms and ecologies, these bacteria are recognized as one of the most diverse and important marine heterotrophic bacterial groups for elucidating the correlation between genome evolution and ecological adaptation. However, on the basis of 16S rRNA gene phylogeny, we could not find any robust monophyletic lineages in any of the known genera. We needed further attempts to reconstruct their evolutionary history based on multilocus sequence analysis (MLSA) and/or genome wide taxonomy of all the recognized species groups. In our previous report in 2007, we conducted the first broad multilocus sequence analysis (MLSA) to infer the evolutionary history of vibrios using nine housekeeping genes (the 16S rRNA gene, gapA, gyrB, ftsZ, mreB, pyrH, recA, rpoA, and topA), and we proposed 14 distinct clades in 58 species of Vibrionaceae. Due to the difficulty of designing universal primers that can amplify the genes for MLSA in every Vibrionaceae species, some clades had yet to be defined. In this study, we present a better picture of an updated molecular phylogeny for 86 described vibrio species and 10 genome sequenced Vibrionaceae strains, using 8 housekeeping gene sequences. This new study places special emphasis on (1) eight newly identified clades (Damselae, Mediterranei, Pectenicida, Phosphoreum, Profundum, Porteresiae, Rosenbergii, and Rumoiensis); (2) clades amended since the 2007 proposal with recently described new species; (3) orphan clades of genomospecies F6 and F10; (4) phylogenetic positions defined in 3 genome-sequenced strains (N418, EX25, and EJY3); and (5) description of V. tritonius sp. nov., which is a member of the "Porteresiae" clade.