Abscisic acid prevents the coalescence of protein storage vacuoles by upregulating expression of a tonoplast intrinsic protein gene in barley aleurone.

Tonoplast intrinsic proteins (TIPs) are integral membrane proteins that are known to function in plants as aquaporins. Here, we propose another role for TIPs during the fusion of protein storage vacuoles (PSVs) in aleurone cells, a process that is promoted by gibberellic acid (GA) and prevented by abscisic acid (ABA). Studies of the expression of barley (Hordeum vulgare) TIP genes (HvTIP) showed that GA specifically decreased the abundance of HvTIP1;2 and HvTIP3;1 transcripts, while ABA strongly increased expression of HvTIP3;1. Increased or decreased expression of HvTIP3;1 interfered with the hormonal effects on vacuolation in aleurone protoplasts. HvTIP3;1 gain-of-function experiments delayed GA-induced vacuolation, whereas HvTIP3;1 loss-of-function experiments promoted vacuolation in ABA-treated aleurone cells. These results indicate that TIP plays a key role in preventing the coalescence of small PSVs in aleurone cells. Hormonal regulation of the HvTIP3;1 promoter is similar to the regulation of the endogenous gene, indicating that induction of the transcription of HvTIP3;1 by ABA is a critical factor in the prevention of PSV coalescence in response to ABA. Promoter analysis using deletions and site-directed mutagenesis of sequences identified three cis-acting elements that are responsible for ABA responsiveness in the HvTIP3;1 promoter. Promoter analysis also showed that ABA responsiveness of the HvTIP3;1 promoter is likely to occur via a unique regulatory system distinct from that involving the ABA-response promoter complexes.

OsbZIP38/87-mediated activation of OsHXK7 improves the viability of rice cells under hypoxic conditions.

Maintenance of energy metabolism is critical for rice (Oryza sativa) tolerance under submerged cultivation. Here, OsHXK7 was the most actively induced hexokinase gene in the embryos of hypoxically germinating rice seeds. Suspension-cultured cells established from seeds of T-DNA null mutants for the OsHXK7 locus did not regrow after 3-d-hypoxic stress and showed increased susceptibility to low-oxygen stress-in terms of viability-and decreased alcoholic fermentation activities compared to those of the wild-type. The promoter element containing the TGACG-motif, a well-known target site for the basic leucine zipper (bZIP) transcription factors, was responsible for sugar regulation of the OsHXK7 promoter activity. Systematic screening of the OsbZIP genes showing the similar expression patterns to that of OsHXK7 in the transcriptomic datasets produced two bZIP genes, OsbZIP38 and 87, belonging to the S1 bZIP subfamily as the candidate for the activator for this gene expression. Gain- and loss-of-function experiments through transient expression assays have demonstrated that these two bZIP proteins are indeed involved in the induction of OsHXK7 expression under starvation or low-energy conditions. Our finding suggests that C/S1 bZIP network-mediated hypoxic deregulation of sugar-responsive genes may work in concert for the molecular adaptation of rice cells to submergence.

Enhancing spatial resolution in Fourier transform infrared spectral image via machine learning algorithms.

Owing to the intrinsic signal noise in the characterization of chemical structures through Fourier transform infrared (FT-IR) spectroscopy, the determination of the signal-to-noise ratio (SNR) depends on the level of the concentration of the chemical structures. In situations characterized by limited concentrations of chemical structures, the traditional approach involves mitigating the resulting low SNR by superimposing repetitive measurements. In this study, we achieved comparable high-quality results to data scanned 64 times and superimposed by employing machine learning algorithms such as the principal component analysis and non-negative matrix factorization, which perform the dimensionality reduction, on FT-IR spectral image data that was only scanned once. Furthermore, the spatial resolution of the mapping images correlated to each chemical structure was enhanced by applying both the machine learning algorithms and the Gaussian fitting simultaneously. Significantly, our investigation demonstrated that the spatial resolution of the mapping images acquired through relative intensity is further improved by employing dimensionality reduction techniques. Collectively, our findings imply that by optimizing research data through noise reduction enhancing spatial resolution using the machine learning algorithms, research processes can be more efficient, for instance by reducing redundant physical measurements.

Enhancement of the Anticancer Ability of Natural Killer Cells through Allogeneic Mitochondrial Transfer.

An in vitro culture period of at least 2 weeks is required to produce sufficient natural killer (NK) cells for immunotherapy, which are the key effectors in hematological malignancy treatment. Mitochondrial damage and fragmentation reduce the NK cell immune surveillance capacity. Thus, we hypothesized that the transfer of healthy mitochondria to NK cells could enhance their anticancer effects. Allogeneic healthy mitochondria isolated from WRL-68 cells were transferred to NK cells. We evaluated NK cells' proliferative capacity, cell cycle, and cytotoxic capacity against various cancer cell types by analyzing specific lysis and the cytotoxic granules released. The relationship between the transferred allogenic mitochondrial residues and NK cell function was determined. After mitochondrial transfer, the NK cell proliferation rate was 1.2-fold higher than that of control cells. The mitochondria-treated NK cells secreted a 2.7-, 4.1-, and 5-fold higher amount of granzyme B, perforin, and IFN-γ, respectively, when co-cultured with K562 cells. The specific lysis of various solid cancer cells increased 1.3-1.6-fold. However, once allogeneic mitochondria were eliminated, the NK cell activity returned to the pre-mitochondrial transfer level. Mitochondria-enriched NK cells have the potential to be used as a novel solid cancer treatment agent, without the need for in vitro cytokine-induced culture.

Comparison of transcriptomic adjustments to availability of sugar, cellular energy, and oxygen in germinating rice embryos.

During germination, the availability of sugars, oxygen, or cellular energy fluctuates under dynamic environmental conditions, likely affecting the global RNA profile of rice genes. Most genes that exhibit sugar-regulation in rice embryos under aerobic conditions are responsive to low energy and anaerobic conditions, indicating that sugar regulation is strongly associated with energy and anaerobic signaling. The interference pattern of sugar regulation by either anaerobic or low energy conditions indicates that induction is likely the more prevalent regulatory mechanism than repression for altering the expression of sugar-regulated genes. Among the aerobically sugar-regulated genes, limited genes exhibit sugar regulation under anaerobic conditions, indicating that anaerobic conditions strongly influence sugar regulated gene expression. Anaerobically responsive genes substantially overlap with low energy responsive genes. In particular, the expression levels of anaerobically downregulated genes are consistent with those provoked by low energy conditions, suggesting that anaerobic downregulation results from the prevention of aerobic respiration due to the absence of the final electron acceptor, i.e., molecular oxygen. It has been noted that abscisic acid (ABA) responsive genes are over representative of genes upregulated under low energy conditions, in contrast to downregulated genes. This suggests that either ABA itself or upstream signaling components of the ABA signaling pathway are likely to be involved in the signaling pathways activated by low energy conditions.

A Diarrhoeagenic EnteropathogenicEscherichia coli (EPEC) Infection Outbreak That Occurred among Elementary School Children in Gyeongsangbuk-Do Province of South Korea Was Associated with Consumption of Water-Contaminated Food Items.

(1) Background: In response to the notification made by an elementary school authority that reported a number of elementary school children being absent in three schools as a result of gastroenteritis symptoms on 4 July 2018, in Gyeongsangbuk-Do Province, South Korea, an epidemic investigation was carried out to determine the extent, cause, and source of the outbreak in order to prevent secondary cases and make recommendations to prevent future recurrences. (2) Methods: In this epidemiologic study, a total of 106 human subjects (school children, staff members, and cooks) who had consumed the possibly contaminated foodstuffs were enrolled retrospectively. Human specimens from clinically defined cases, food and drinks, supply and storage of them, and environmental and sanitary conditions were also assessed by observation, laboratory tests, and survey questionnaires-where and whatever applicable. The attack rate and positive rate for human specimens were first presented followed by the calculation of the relative risk ratio (RR) with 95% CI (confidence intervals) in order to identify the exposure and outcome relationships. (3) Results: The attack rate was 12.26% (13/106) for those who had ingested the food items at the three schools and the positive rate of enteropathogenic Escherichia coli (EPEC) was 15.38% (2/13). The relative risk (RR) of developing food poisoning of those who consumed the cucumber chili with ssamjang and seasoned cucumber and chives were 4.55 (95% CI 1.05-19.54) and 9.20 (95% CI 1.24-68.22), respectively. In addition, within the human specimens as well as the water and environmental samples different strains of diarrhoeagenic enteropathogenic Escherichia coli (EPEC) were detected. (4) Conclusions: Provision of safe and wholesome water access to all elementary schools by concerned authorities, especially during the likely seasons of water source contamination, as well as health education promotion about foodborne outbreaks to all school stakeholders is therefore recommended.

Charge-Induced Disorder Controls the Thermal Conductivity of Entropy-Stabilized Oxides.

Manipulating a crystalline material's configurational entropy through the introduction of unique atomic species can produce novel materials with desirable mechanical and electrical properties. From a thermal transport perspective, large differences between elemental properties such as mass and interatomic force can reduce the rate at which phonons carry heat and thus reduce the thermal conductivity. Recent advances in materials synthesis are enabling the fabrication of entropy-stabilized ceramics, opening the door for understanding the implications of extreme disorder on thermal transport. Measuring the structural, mechanical, and thermal properties of single-crystal entropy-stabilized oxides, it is shown that local ionic charge disorder can effectively reduce thermal conductivity without compromising mechanical stiffness. These materials demonstrate similar thermal conductivities to their amorphous counterparts, in agreement with the theoretical minimum limit, resulting in this class of material possessing the highest ratio of elastic modulus to thermal conductivity of any isotropic crystal.

Dissection of cis-regulatory element architecture of the rice oleosin gene promoters to assess abscisic acid responsiveness in suspension-cultured rice cells.

Oleosins are the most abundant proteins in the monolipid layer surrounding neutral storage lipids that form oil bodies in plants. Several lines of evidence indicate that they are physiologically important for the maintenance of oil body structure and for mobilization of the lipids stored inside. Rice has six oleosin genes in its genome, the expression of all of which was found to be responsive to abscisic acid (ABA) in our examination of mature embryo and aleurone tissues. The 5'-flanking region of OsOle5 was initially characterized for its responsiveness to ABA through a transient expression assay system using the protoplasts from suspension-cultured rice cells. A series of successive deletions and site-directed mutations identified five regions critical for the hormonal induction of its promoter activity. A search for cis-acting elements in these regions deposited in a public database revealed that they contain various promoter elements previously reported to be involved in the ABA response of various genes. A gain-of-function experiment indicated that multiple copies of all five regions were sufficient to provide the minimal promoter with a distinct ABA responsiveness. Comparative sequence analysis of the short, but still ABA-responsive, promoters of OsOle genes revealed no common modular architecture shared by them, indicating that various distinct promoter elements and independent trans-acting factors are involved in the ABA responsiveness of rice oleosin multigenes.

Differential anoxic expression of sugar-regulated genes reveals diverse interactions between sugar and anaerobic signaling systems in rice.

The interaction between the dual roles of sugar as a metabolic fuel and a regulatory molecule was unveiled by examining the changes in sugar signaling upon oxygen deprivation, which causes the drastic alteration in the cellular energy status. In our study, the expression of anaerobically induced genes is commonly responsive to sugar, either under the control of hexokinase or non-hexokinase mediated signaling cascades. Only sugar regulation via the hexokinase pathway was susceptible for O2 deficiency or energy deficit conditions evoked by uncoupler. Examination of sugar regulation of those genes under anaerobic conditions revealed the presence of multiple paths underlying anaerobic induction of gene expression in rice, subgrouped into three distinct types. The first of these, which was found in type-1 genes, involved neither sugar regulation nor additional anaerobic induction under anoxia, indicating that anoxic induction is a simple result from the release of sugar repression by O2-deficient conditions. In contrast, type-2 genes also showed no sugar regulation, albeit with enhanced expression under anoxia. Lastly, expression of type-3 genes is highly enhanced with sugar regulation sustained under anoxia. Intriguingly, the inhibition of the mitochondrial ATP synthesis can reproduce expression pattern of a specific set of anaerobically induced genes, implying that rice cells may sense O2 deprivation, partly via perception of the perturbed cellular energy status. Our study of interaction between sugar signaling and anaerobic conditions has revealed that sugar signaling and the cellular energy status are likely to communicate with each other and influence anaerobic induction of gene expression in rice.

Hexokinase-mediated sugar signaling controls expression of the calcineurin B-like interacting protein kinase 15 gene and is perturbed by oxidative phosphorylation inhibition.

Calcineurin B-like (CBL) interacting protein kinase 15 (CIPK15) is a newly identified positive regulator which is critical to directing the O(2) deficiency signal to the sugar signaling cascade as part of Amy3D (representative Amy3 gene) regulation in rice. It is located upstream and probably contributes to reserve mobilization under anoxia. In isolated starving embryos, the temporal pattern of accumulation of CIPK15 transcripts and leaky suppression of this gene suggests that factors other than CIPK15 may also be involved in the regulation of Amy3D expression. Probing of a variety of sugars and sugar analogs has shown that hexokinase mediates the sugar regulation of CIPK15. For example, hexokinase substrates, such as mannose, 2-deoxyglucose, and other metabolizable sugars, repressed CIPK15 expression, whereas 3-O-methylglucose and 6-deoxyglucose did not. By using glucosamine, a hexokinase inhibitor, to release glucose-dependent CIPK15 suppression, we confirmed that hexokinase mediates regulation of this gene. Chemical inhibitors of mitochondrial electron transfer, proton separation or ATP synthase also effectively abolished sugar-induced repression of CIPK15. This type of interference, the release from glucose-induced repression of gene expression by inhibition of oxidative phosphorylation, was previously identified for the Amy3D gene, which suggests that hexokinase-mediated sugar signaling may be coordinated with the cellular energy status. Analysis of a transgenic rice cell line harboring the GUS reporter gene under the control of the CIPK15 promoter, and transient expression assay for 3' UTR of the CIPK15 gene indicate that sugar regulation of the rice CIPK15 gene is likely mediated by 2548-bp 5'-flanking region, with no additional post-transcriptional control.