The uncoating of EV71 in mature late endosomes requires CD-M6PR

Enterovirus 71 (EV71) is one of the causative agents of hand-foot-and-mouth disease, which in some circumstances could lead to severe neurological diseases. Despite of its importance for human health, little is known about the early stages of EV71 infection. EV71 starts uncoating with its receptor, human scavenger receptor B2 (hSCARB2), at low pH. We show that EV71 was not targeted to lysosomes in human rhabdomyosarcoma cells overexpressing hSCARB2 and that the autophagic pathway is not essential for EV71 productive uncoating. Instead, EV71 was efficiently uncoated 30 minutes after infection in late endosomes (LEs) containing hSCARB2, mannose-6-phosphate receptor (M6PR), RAB9, bis(monoacylglycero)phosphate and lysosomal associated membrane protein 2 (LAMP2). Furthering the notion that mature LEs are crucial for EV71 uncoating, cation-dependent (CD)-M6PR knockdown impairs EV71 infection. Since hSCARB2 interacts with cation-independent (CI)-M6PR through M6P-binding sites and CD-M6PR also harbor a M6P-binding site, CD-M6PR is likely to play important roles in EV71 uncoating in LEs

Enhanced Recombinant Protein Productivity by Genome Reduction in Bacillus subtilis

The emerging field of synthetic genomics is expected to facilitate the generation of microorganisms with the potential to achieve a sustainable society. One approach towards this goal is the reduction of microbial genomes by rationally designed deletions to create simplified cells with predictable behavior that act as a platform to build in various genetic systems for specific purposes. We report a novel Bacillus subtilis strain, MBG874, depleted of 874 kb (20%) of the genomic sequence. When compared with wild-type cells, the regulatory network of gene expression of the mutant strain is reorganized after entry into the transition state due to the synergistic effect of multiple deletions, and productivity of extracellular cellulase and protease from transformed plasmids harboring the corresponding genes is remarkably enhanced. To our knowledge, this is the first report demonstrating that genome reduction actually contributes to the creation of bacterial cells with a practical application in industry. Further systematic analysis of changes in the transcriptional regulatory network of MGB874 cells in relation to protein productivity should facilitate the generation of improved B. subtilis cells as hosts of industrial protein production

Oligodendrocyte dynamics dictate cognitive performance outcomes of working memory training in mice

Previous work has shown that motor skill learning stimulates and requires generation of myelinating oligodendrocytes (OLs) from their precursor cells (OLPs) in the brains of adult mice. In the present study we ask whether OL production is also required for non-motor learning and cognition, using T-maze and radial-arm-maze tasks that tax spatial working memory. We find that maze training stimulates OLP proliferation and OL production in the medial prefrontal cortex (mPFC), anterior corpus callosum (genu), dorsal thalamus and hippocampal formation of adult male mice; myelin sheath formation is also stimulated in the genu. Genetic blockade of OL differentiation and neo-myelination in Myrf conditional-knockout mice strongly impairs training-induced improvements in maze performance. We find a strong positive correlation between the performance of individual wild type mice and the scale of OLP proliferation and OL generation during training, but not with the number or intensity of c-Fos+ neurons in their mPFC, underscoring the important role played by OL lineage cells in cognitive processing

Analysis of genetic factors associated with fatty liver disease‐related hepatocellular carcinoma

Abstract Aim Single‐nucleotide polymorphisms (SNPs) in PNPLA3 and hydroxysteroid 17‐beta dehydrogenase 13 (HSD17B13) genes are associated with fatty liver disease (FLD) progression and carcinogenesis. In the present study, we evaluated the characteristics of Japanese FLD patients according to HSD17B13 polymorphisms. Methods We enrolled 402 patients who were clinically and pathologically diagnosed with FLD (alcoholic: 63 cases, nonalcoholic: 339 cases) at our hospital in 1990–2018 (228 males; median age: 54.9 [14.6–83.6] years). FLD patients with HSD17B13 A/A (212 cases) and others (A/AA or AA/AA; 190 cases) were compared. Results Compared to patients with HSD17B13 A/A and others, those with the A/A genotype showed increased incidence of hepatocellular carcinoma (HCC) (A/A vs. others; 18.4% vs. 9.5%, p = 0.01), cardiovascular diseases (14.2% vs. 4.2%, p < 0.01), and hypertension (56.6% vs. 47.4%, p = 0.06). In patients without A/A, the HCC incidence was significantly reduced in those with alcohol‐related FLD, fibrosis‐4 index <2.67, and the PNPLA3 CC genotype; however, there was no significant difference in nonalcoholic‐FLD. Patients without HSD17B13 A/A showed severe steatosis (77% vs. 88.6%, p < 0.01). New HCC developed in 11 cases and the 5–year incidence rate of HCC was 3.3% in patients with both PNPLA3 GG/GC and HSD17B13 A/A, which was significantly higher than the rate for those with other SNP profiles (0.6%, p = 0.03). Conclusions Inhibiting HSD17B13 activity may prevent HCC development, particularly in alcohol‐related FLD and low‐risk patients. Therefore, combinations of SNPs and other risk factors can be used for screening FLD–HCC

High external pH enables more efficient secretion of alkaline α-amylase AmyK38 by <it>Bacillus subtilis</it>

Abstract Background Bacillus subtilis genome-reduced strain MGB874 exhibits enhanced production of exogenous extracellular alkaline cellulase Egl-237 and subtilisin-like alkaline protease M-protease. Here, we investigated the suitability of strain MGB874 for the production of α-amylase, which was anticipated to provoke secretion stress responses involving the CssRS (Control secretion stress Regulator and Sensor) system. Results Compared to wild-type strain 168, the production of a novel alkaline α-amylase, AmyK38, was severely decreased in strain MGB874 and higher secretion stress responses were also induced. Genetic analyses revealed that these phenomena were attributable to the decreased pH of growth medium as a result of the lowered expression of rocG, encoding glutamate dehydrogenase, whose activity leads to NH3 production. Notably, in both the genome-reduced and wild-type strains, an up-shift of the external pH by the addition of an alkaline solution improved AmyK38 production, which was associated with alleviation of the secretion stress response. These results suggest that the optimal external pH for the secretion of AmyK38 is higher than the typical external pH of growth medium used to culture B. subtilis. Under controlled pH conditions, the highest production level (1.08 g l-1) of AmyK38 was obtained using strain MGB874. Conclusions We demonstrated for the first time that RocG is an important factor for secretory enzyme production in B. subtilis through its role in preventing acidification of the growth medium. As expected, a higher external pH enabled a more efficient secretion of the alkaline α-amylase AmyK38 in B. subtilis. Under controlled pH conditions, the reduced-genome strain MGB874 was demonstrated to be a beneficial host for the production of AmyK38.</p

Frequency distribution of fibril diameter.

<p>A: dermis. Dermis in S_a (stippled bars), in S_b (diagonally hatched bars) and in S_c (filled bars) are compared. B: Dermis treated with Triton. Triton-treated dermis (diagonally hatched bars) and Triton-FT dermis (stippled bars) are compared. C: Collagen suspensions. The samples aggregated by tensilin (diagonally hatched bars), dispersed by softenin (stippled bars) or dispersed by FT-treatment (stippled bars with horizontal stripes) are compared. The diameter class 20, for example, includes the fibrils with the diameter equal to or larger than 20 nm and less than 40 nm. The total numbers of measured fibrils were: 180 for each state in A, 180 for each treatment in B, and 30 for each solution. In this and Figs <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0155673#pone.0155673.g006" target="_blank">6</a>, <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0155673#pone.0155673.g007" target="_blank">7</a> and <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0155673#pone.0155673.g008" target="_blank">8</a> the slant hatching and the stippling denote that the samples were in the state that mechanically corresponded to the standard state and to the soft state respectively.</p

Electron micrographs of collagen-fibril suspensions.

<p>Samples were aggregated by tensilin (C), which were then dispersed either by FT treatment (A) or by softenin (B). The arrowhead indicates a fibril that appears as if 4 or more thinner fibrils were embedded in a less electron-dense matrix to form a fibril. Scale bar, 100 nm.</p

Electron micrograph of dermis in the standard state cut transversely to the strain direction.

<p>Collagen fibrils in cross sections are observed. In this, and Figs <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0155673#pone.0155673.g004" target="_blank">4</a> and <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0155673#pone.0155673.g009" target="_blank">9</a>, unlabelled arrows denote cross-bridges between fibrils. Arrow marked a: fibril with a meandering perimeter that folds deeply towards the center of the fibril, suggesting that the fibril is possibly made of 4 subfibrils. Arrow marked b: thick fibril containing an electron-lucent area in which an electron-dense spot is observed at the border of the electron-lucent area. Arrow marked c: electron-dense spot on the perimeter of fibrils. Scale bar, 100 nm.</p