NKT cells are responsible for the clearance of murine norovirus through the virus-specific secretory IgA pathway

Norovirus infection cause epidemic nonbacterial gastroenteritis in patients. The immune mechanisms responsible for the clearance of virus are not completely understood. We examined whether NKT cells are effective against norovirus infection using CD1d KO mice. The body weights of 4-weeks-old CD1d KO mice that were infected with murine norovirus-S7 (MNV-S7) were significantly lower than those of non-infected CD1d KO mice. On the other hand, the body weights of infected WT mice were comparable to those of non-infected WT mice. Correspondingly, CD1d KO mice had an almost 1000-fold higher MNV-S7 burden in the intestine after infection in comparison to WT mice. The mechanism responsible for the insufficient MNV-S7 clearance in CD1d KO mice was attributed to reduced IFN-γ production early during MNV-S7 infection. In addition, the markedly impaired IL-4 production in CD1d KO mice resulted in an impaired MNV-S7-specific secretory IgA production after MNV-S7 infection which is associated with mucosal immunity. Thus, the present results provide evidence that NKT cells play an essential role in MNV-S7 clearance.     

Correction to: Comparative analysis of sperm motility in liquid and seminal coagulum portions between Bornean orangutan (Pongo pygmaeus) and chimpanzee (Pan troglodytes)

Primates - In the original publication of the article, the coauthor “Takashi Hayakawa” was wrongly assigned as co-corresponding author.     

Molecular characterization of mouse CREB3 regulatory factor in Neuro2a cells

We performed expression and functional analysis of mouse CREB3 regulatory factor (CREBRF) in Neuro2a cells by constructing several expression vectors. Overexpressed full-length (FL) CREBRF protein was stabilized by MG132; however, the intrinsic CREBRF expression in Neuro2a cells was negligible under all conditions. On the other hand, N- or C-terminal deletion of CREBRF influenced its stability. Cotransfection of CREBRF together with GAL4-tagged FL CREB3 increased luciferase reporter activity, and only the N-terminal region of CREBRF was sufficient to potentiate luciferase activity. Furthermore, this positive effect of CREBRF was also observed in cells expressing GAL4-tagged cleaved CREB3, although CREBRF hardly influenced the protein stability of NanoLuc-tagged cleaved CREB3 or intracellular localization of EGFP-tagged one. In conclusion, this study suggests that CREBRF, a quite unstable proteasome substrate, positively regulates the CREB3 pathway, which is distinct from the canonical ER stress pathway in Neuro2a cells.

     

Site-directed mutagenesis by biolistic transformation efficiently generates inheritable mutations in a targeted locus in soybean somatic embryos and transgene-free descendants in the T1 generation

The clustered regularly interspaced short palindromic repeat (CRISPR)/CRISPR-associated endonuclease 9 (Cas9) system is being rapidly developed for mutagenesis in higher plants. Ideally, foreign DNA introduced by this system is removed in the breeding of edible crops and vegetables. Here, we report an efficient generation of Cas9-free mutants lacking an allergenic gene, Gly m Bd 30K, using biolistic transformation and the CRISPR/Cas9 system. Five transgenic embryo lines were selected on the basis of hygromycin resistance. Cleaved amplified polymorphic sequence analysis detected only two different mutations in e all of the lines. These results indicate that mutations were induced in the target gene immediately after the delivery of the exogenous gene into the embryo cells. Soybean plantlets (T₀ plants) were regenerated from two of the transgenic embryo lines. The segregation pattern of the Cas9 gene in the T₁ generation, which included Cas9-free plants, revealed that a single copy number of transgene was integrated in both lines. Immunoblot analysis demonstrated that no Gly m Bd 30K protein accumulated in the Cas9-free plants. Gene expression analysis indicated that nonsense mRNA decay might have occurred in mature mutant seeds. Due to the efficient induction of inheritable mutations and the low integrated transgene copy number in the T₀ plants, we could remove foreign DNA easily by genetic segregation in the T₁ generation. Our results demonstrate that biolistic transformation of soybean embryos is useful for CRISPR/Cas9-mediated site-directed mutagenesis of soybean for human consumption.

     

Characterization,subsite mapping and N-terminal sequence of miliin,a serine-protease isolated from the latex of Euphorbia milii

Miliin is a serine protease purified from the latex of Euphorbia milii. This work reports the effect of pH and temperature on the catalytic activity of miliin, using fluorescence resonance energy transfer (FRET) substrates. Miliin displayed the highest activity at pH 9 and 35 °C. Subsite mapping shows that subsites S₂ to S₂′ prefer uncharged residues. The S₂ subsite prefers hydrophobic aliphatic amino acids (Val, Pro and Ile) and defines the cleavage site. This work is the first one that reports subsite mapping of Euphorbiacea proteases. The N-terminal sequence showed higher similarity (40%) with the serine protease LIM9 isolated from Lilium. The presence of Tyr, Pro and Lys at positions 2, 5 and 10 respectively, were observed for most of the serine proteases used for comparison. The N-terminal sequence has striking differences with those reported previously for milin and eumiliin, other serine proteases isolated from the latex of E. milii.     

Lateral position preference in grazing feral horses

Behavioural lateralisation is an effective way for animals to manage daily tasks by specialising behaviour to either side of the body. Many types of lateralisation are linked to the function of each brain hemisphere. Lateralisation of monitoring behaviour in mother–infant relationships occurs in a wide range of mammals, where infants frequently use their left eye to monitor their mother. However, few studies have focused on this type of spatial relationships among adults in daily life, such as during foraging. The present study focused on monitoring adult feral horse behaviour using quantitative analysis of spatial relationships, using drone technology. We found that horses form a localised spatial relationship with their nearest neighbour. Specifically, the nearest neighbour was located to the left rear of a target individual significantly more frequently than to the right rear. Furthermore, the nearest neighbour was less frequently located behind a target individual. We propose that this relationship is caused by a left-eye preference, because information via the left eye predominantly proceeds to the right hemisphere, which is dominant for social processing.     

Molecular dynamics simulation of diffusion of hydrogen in binary hydrogen–tetrahydrofuran hydrate

The binary structure II hydrogen–tetrahydrofuran (THF) hydrate was studied with molecular dynamics simulation. The simulations were carried out at 300, 310 K and 10.1 MPa, and with various contents of hydrogen and THF. The migrations of hydrogen molecules from cage to cage were observed. The migration process of hydrogen was also analysed, and the diffusion coefficients of hydrogen in the hydrate were calculated. The calculated diffusion coefficients qualitatively agreed with the experimental data. Double and quintet occupancies of hydrogen molecules were observed in the small and large cages, respectively, without changing the hydrate structure.     

Depression in Medical School: The Influence of Morningness‐Eveningness

Medical students are at higher risk for depression, affecting not only their lives but also patient care. This article studied a population of medical students engaged in lecture‐based learning regarding the presence of depressive symptoms and its relation to morningness‐eveningness. Depressive symptoms were assessed by the Beck Depressive Inventory scale (BDI>10), and diurnal preference was assessed by the Horne & Östberg Morningness/Eveningness Questionnaire (MEQ). Family history of depression and involvement in regular physical activity were also investigated. A total of 161 students, 77 (47.8%) males, aged 19 to 30 yrs (22.1±2.1) living in a city close to the equator were evaluated. Fifty‐three individuals (32.9%) had depressive symptoms. Depressive individuals showed a trend to be female (p=0.07). Also, female gender showed a non‐significant shift toward morningness. Fifty‐eight (36.0%) subjects participated in regular physical activity. In 57 cases (35.4%), there was a history of depression in the family. Fifteen individuals (9.3%) were definitely evening type, 42 (26.1%) were moderately evening type, 44 (27.3%) were indifferent, 42 (26.1%) were moderately morning type, and 18 (11.2%) were definitely morning type. Family history of depression (OR=0.29, 95% CI=1.37–6.12) and sedentary life (OR=0.28, 95% CI=0.12–0.65) were associated with depressive symptoms. Eveningness was associated with depressive symptoms (OR=0.66, 95% CI=0.50–0.88), and this association remained significant after adjusting for the presence of familial depression and physical activity (OR=0.71, 95% CI=0.52–0.95). In conclusion, depressive symptoms are independently associated with “eveningness” in medical students. These results should be confirmed by future studies involving a larger number of subjects.