Mycobacterium bovis BCG Given at Birth Followed by Inactivated Respiratory Syncytial Virus Vaccine Prevents Vaccine-Enhanced Disease by Promoting Trained Macrophages and Resident Memory T Cells.

Respiratory syncytial virus (RSV) infects more than 60% of infants in their first year of life. Since an experimental formalin-inactivated (FI) RSV vaccine tested in the 1960s caused enhanced respiratory disease (ERD), few attempts have been made to vaccinate infants. ERD is characterized by Th2-biased responses, lung inflammation, and poor protective immune memory. Innate immune memory displays an increased nonspecific effector function upon restimulation, a process called trained immunity, or a repressed effector function upon restimulation, a process called tolerance, which participates in host defense and inflammatory disease. Mycobacterium bovis bacillus Calmette-Guérin (BCG) given at birth can induce trained immunity as well as heterologous Th1 responses. We speculate that BCG given at birth followed by FI-RSV may alleviate ERD and enhance protection through promoting trained immunity and balanced Th immune memory. Neonatal mice were given BCG at birth and then vaccinated with FI-RSV+Al(OH)3. BCG/FI-RSV+Al(OH)3 induced trained macrophages, tissue-resident memory T cells (TRM), and specific cytotoxic T lymphocytes (CTL) in lungs and inhibited Th2 and Th17 cell immune memory, all of which contributed to inhibition of ERD and increased protection. Notably, FI-RSV+Al(OH)3 induced tolerant macrophages, while BCG/FI-RSV+Al(OH)3 prevented the innate tolerance through promoting trained macrophages. Moreover, inhibition of ERD was attributed to trained macrophages or TRM in lungs but not memory T cells in spleens. Therefore, BCG given at birth to regulate trained immunity and TRM may be a new strategy for developing safe and effective RSV killed vaccines for young infants. IMPORTANCE RSV is the leading cause of severe lower respiratory tract infection of infants. ERD, characterized by Th2-biased responses, inflammation, and poor immune memory, has been an obstacle to the development of safe and effective killed RSV vaccines. Innate immune memory participates in host defense and inflammatory disease. BCG given at birth can induce trained immunity as well as heterologous Th1 responses. Our results showed that BCG/FI-RSV+Al(OH)3 induced trained macrophages, TRM, specific CTL, and balanced Th cell immune memory, which contributed to inhibition of ERD and increased protection. Notably, FI-RSV+Al(OH)3 induced tolerant macrophages, while BCG/FI-RSV+Al(OH)3 prevented tolerance through promoting trained macrophages. Moreover, inhibition of ERD was attributed to trained macrophages or TRM in lungs but not memory T cells in spleens. BCG at birth as an adjuvant to regulate trained immunity and TRM may be a new strategy for developing safe and effective RSV killed vaccines for young infants.

Silencing geranylgeranyltransferase I inhibits the migration and invasion of salivary adenoid cystic carcinoma through RhoA/ROCK1/MLC signaling and suppresses proliferation through cell cycle regulation.

Geranylgeranyltransferase type I (GGTase-I) significantly affects Rho proteins, such that the malignant progression of several cancers may be induced. Nevertheless, the effect and underlying mechanism of GGTase-I in the malignant progression of salivary adenoid cystic carcinoma (SACC) remain unclear. This study primarily aimed to investigate the role and mechanism of GGTase-I in mediating the malignant progression of SACC. The level of GGTase-I gene in cells was stably knocked down by short hairpin RNA-EGFP-lentivirus. The effects of GGTase-I silencing on the migration, invasion, and spread of cells were examined, the messenger RNA levels of GGTase-I and RhoA genes of SACC cells after GGTase-I knockdown were determined, and the protein levels of RhoA and RhoA membrane of SACC cells were analyzed. Moreover, the potential underlying mechanism of silencing GGTase-I on the above-mentioned aspects in SACC cells was assessed by examining the protein expression of ROCK1, MLC, p-MLC, E-cadherin, Vimentin, MMP2, and MMP9. Furthermore, the underlying mechanism of SACC cells proliferation was investigated through the analysis of the expression of cyclinD1, MYC, E2F1, and p21CIP1/WAF1 . Besides, the change of RhoA level in SACC tissues compared with normal paracancer tissues was demonstrated through quantitative reverse-transcription polymerase chain reaction and western blot experiments. Next, the effect after GGTase-I silencing was assessed through the subcutaneous tumorigenicity assay. As indicated by the result of this study, the silencing of GGTase-I significantly reduced the malignant progression of tumors in vivo while decreasing the migration, invasion, and proliferation of SACC cells and RhoA membrane, Vimentin, ROCK1, p-MLC, MMP2, MMP9, MYC, E2F1, and CyclinD1 expression. However, the protein expression of E-cadherin and p21CIP1/WAF1 was notably upregulated. Subsequently, no significant transform of RhoA and MLC proteins was identified. Furthermore, RhoA expression in SACC tissues was significantly higher than that in paracancerous tissues. As revealed by the results of this study, GGTase-I shows a correlation with the proliferation of SACC through the regulation of cell cycle and may take on vital significance in the migration and invasion of SACC by regulating RhoA/ROCK1/MLC signaling pathway. GGTase-I is expected to serve as a novel exploration site of SACC.

Oligomerization Mechanism of Methylglyoxal Regulated by the Methyl Groups in Reduced Nitrogen Species: Implications for Brown Carbon Formation.

Uncertain chemical mechanisms leading to brown carbon (BrC) formation affect the drivers of the radiative effects of aerosols in current climate predictions. Herein, the aqueous-phase reactions of methylglyoxal (MG) and typical reduced nitrogen species (RNSs) are systematically investigated by using combined quantum chemical calculations and laboratory experiments. Imines and diimines are identified from the mixtures of methylamine (MA) and ammonia (AM) with MG, but not from dimethylamine (DA) with the MG mixture under acidic conditions, because deprotonation of DA cationic intermediates is hindered by the amino groups occupied by two methyl groups. It leads to N-heterocycle (NHC) formation in the MG + MA (MGM) and MG + AM (MGA) reaction systems but to N-containing chain oligomer formation in the MG + DA (MGD) reaction system. Distinct product formation is attributed to electrostatic attraction and steric hindrance, which are regulated by the methyl groups of RNSs. The light absorption and adverse effects of NHCs are also strongly related to the methyl groups of RNSs. Our finding reveals that BrC formation is mainly contributed from MG reaction with RNSs with less methyl groups, which have more abundant and broad sources in the urban environments.

Comparative Study on the Effects of Four Plant Protein Sources on the Liver and Intestinal Health of Largemouth Bass, Micropterus salmoides.

The effects of plant protein sources (PPSs) on the health of the liver and intestine of the largemouth bass, Micropterus salmoides, were compared to verify the potential damaging effects of dietary fiber (DF). A diet containing 55% fish meal (FM) was used as the control. The test diets contained 25% soybean meal (SBM), rapeseed meal (RSM), cottonseed meal, or peanut meal, and the FM content was decreased to 30%. The protein and lipid contents of these five diets were balanced by casein and oil. Fish were raised for 8 weeks. The fish fed the diet containing PPS showed a trend of decreasing growth and apparent digestibility coefficients. The contents of total bile acid, lipid, and collagen in the liver were increased, and the mRNA expression levels of genes encoding inflammatory factors and enzymes involved in de novo fatty acid synthesis and bile acid synthesis were upregulated. Both the lipid and collagen contents in the liver were positively correlated with the DF content in the diet significantly. Morphology and histology showed reduced liver size, hepatic steatosis, and fibrosis in fish fed diets containing PPS. The lowest hepatosomatic index was observed in fish fed the SBM diet, and the most severe damage was observed in fish fed the RSM diet. No obvious histological abnormalities were observed in the hindgut. The bile acid profile in the liver could be used to distinguish the types of PPS very well by Fisher discriminant analysis. These results indicated that 25% of each of the four PPSs in the diet exceeded the tolerance range of largemouth bass and caused liver damage, which might be mediated by bile acid. DF in PPS might be an important agent contributing to liver damage.

Activation of the Chemokine Receptor CCR1 and Preferential Recruitment of Gαi Suppress RSV Replication: Implications for Developing Novel Respiratory Syncytial Virus Treatment Strategies.

Respiratory syncytial virus (RSV) is a major pathogen that can cause acute respiratory infectious diseases of the upper and lower respiratory tract, especially in children, elderly individuals, and immunocompromised people. Generally, following viral infection, respiratory epithelial cells secrete cytokines and chemokines to recruit immune cells and initiate innate and/or adaptive immune responses. However, whether chemokines affect viral replication in nonimmune cells is rarely clear. In this study, we detected that chemokine CCL5 was highly expressed, while expression of its receptor, CCR1, was downregulated in respiratory epithelial cells after RSV infection. When we overexpressed CCR1 on respiratory epithelial cells in vivo or in vitro, viral load was significantly suppressed, which can be restored by the neutralizing antibody for CCR1. Interestingly, the antiviral effect of CCR1 was not related to type I interferon (IFN-I), apoptosis induction, or viral adhesion or entry inhibition. In contrast, it was related to the preferential recruitment and activation of the adaptor Gαi, which promoted inositol 1,4,5-triphosphate receptor type 3 (ITPR3) expression, leading to inhibited STAT3 phosphorylation; explicitly, phosphorylated STAT3 (p-STAT3) was verified to be among the important factors regulating the activity of HSP90, which has been previously reported to be a chaperone of RSV RNA polymerase. In summary, we are the first to reveal that CCR1 on the surface of nonimmune cells regulates RSV replication through a previously unknown mechanism that does not involve IFN-I induction. IMPORTANCE Our results revealed a novel mechanism by which RSV escapes innate immunity. That is, although it induces high CCL5 expression, RSV might attenuate the binding of CCL5 by downregulating the expression of CCR1 in respiratory epithelial cells to weaken the inhibitory effect of CCR1 on HSP90 activity and thereby facilitate RSV replication in nonimmune cells. This study provides a new target for the development of co-antiviral inhibitors against other components of the host and co-molecular chaperone/HSP90 and provides a scientific basis for the search for effective broad-spectrum antiviral drugs.

Three-Component Tandem Amidosulfenylation of Alkenes for the Synthesis of ß-Succinimide Sulfides.

An environmentally benign multicomponent strategy for the amidosulfenylation of alkenes for the synthesis of ß-succinimide sulfides is disclosed. In this process, common disulfides smoothly act as a sulfur-based source, and N-iodosuccinimide (NIS) is used not only as a free radical initiator but also as an N-nucleophile. A broad range of functional groups are tolerated in this reaction system.

Spontaneous population oscillation of confined active granular particles.

Spontaneous collective oscillation may emerge from seemingly irregular active matter systems. Here, we experimentally demonstrate a spontaneous population oscillation of active granular particles confined in two chambers connected by a narrow channel, and verify the intriguing behavior predicted in simulation [M. Paoluzzi, R. Di Leonardo and L. Angelani, Self-sustained density oscillations of swimming bacteria confined in microchambers, Phys. Rev. Lett., 2015, 115(18), 188303]. During the oscillation, the two chambers are alternately (nearly) filled up and emptied by the self-propelled particles in a periodic manner. We show that the stable unidirectional flow induced due to the confined channel and its periodic reversal triggered by the particle concentration difference between two chambers jointly give rise to the oscillatory collective behavior. Furthermore, we propose a minimal theoretical model that properly reproduces the experimental results without free parameters. This self-sustained collective oscillation could serve as a robust active granular clock, capable of providing rhythmic signals.

Migration of an active colloidal cell in inhomogeneous environments.

Living cells on a substrate with mechanical inhomogeneities often migrate along or against the mechanical gradient, i.e., mechanotaxis, which inspires us to ask how biomimetic cells without biochemical signaling processes respond to environmental inhomogeneity. Here, we perform computer simulations to study the migration of a 2D active colloidal cell (ACC), which consists of active particles enclosed by a passive vesicle, in a heterogeneous environment composed of two adjoining uniform regions with different attributes (influencing the persistent length of the active particle). We find that the ACC can migrate unidirectionally across the interface separating the heterogeneous region and behave tactically. Interestingly, the tactic motion of the ACC is qualitatively different from that of the constituent active particles themselves. In addition, the ACC may also experience a directed drift along the interface of the heterogeneous environment. The tactic behavior of the ACC can be explained by analyzing the pressure distribution on the cell membrane exerted by the enclosed active particles. The findings provide insights into understanding the taxis of biological cells and designing biomimetic cells with environment-sensitive capabilities.

Frequency and Spectrum of Mutations Induced by Gamma Rays Revealed by Phenotype Screening and Whole-Genome Re-Sequencing in Arabidopsis thaliana.

Genetic variations are an important source of germplasm diversity, as it provides an allele resource that contributes to the development of new traits for plant breeding. Gamma rays have been widely used as a physical agent for mutation creation in plants, and their mutagenic effect has attracted extensive attention. However, few studies are available on the comprehensive mutation profile at both the large-scale phenotype mutation screening and whole-genome mutation scanning. In this study, biological effects on M1 generation, large-scale phenotype screening in M2 generation, as well as whole-genome re-sequencing of seven M3 phenotype-visible lines were carried out to comprehensively evaluate the mutagenic effects of gamma rays on Arabidopsis thaliana. A total of 417 plants with visible mutated phenotypes were isolated from 20,502 M2 plants, and the phenotypic mutation frequency of gamma rays was 2.03% in Arabidopsis thaliana. On average, there were 21.57 single-base substitutions (SBSs) and 11.57 small insertions and deletions (InDels) in each line. Single-base InDels accounts for 66.7% of the small InDels. The genomic mutation frequency was 2.78 × 10-10/bp/Gy. The ratio of transition/transversion was 1.60, and 64.28% of the C > T events exhibited the pyrimidine dinucleotide sequence; 69.14% of the small InDels were located in the sequence with 1 to 4 bp terminal microhomology that was used for DNA end rejoining, while SBSs were less dependent on terminal microhomology. Nine genes, on average, were predicted to suffer from functional alteration in each re-sequenced line. This indicated that a suitable mutation gene density was an advantage of gamma rays when trying to improve elite materials for one certain or a few traits. These results will aid the full understanding of the mutagenic effects and mechanisms of gamma rays and provide a basis for suitable mutagen selection and parameter design, which can further facilitate the development of more controlled mutagenesis methods for plant mutation breeding.

Ionizing Radiation: Effective Physical Agents for Economic Crop Seed Priming and the Underlying Physiological Mechanisms.

To overcome various factors that limit crop production and to meet the growing demand for food by the increasing world population. Seed priming technology has been proposed, and it is considered to be a promising strategy for agricultural sciences and food technology. This technology helps to curtail the germination time, increase the seed vigor, improve the seedling establishment, and enhance the stress tolerance, all of which are conducive to improving the crop yield. Meanwhile, it can be used to reduce seed infection for better physiological or phytosanitary quality. Compared to conventional methods, such as the use of water or chemical-based agents, X-rays, gamma rays, electron beams, proton beams, and heavy ion beams have emerged as promising physics strategies for seed priming as they are time-saving, more effective, environmentally friendly, and there is a greater certainty for yield improvement. Ionizing radiation (IR) has certain biological advantages over other seed priming methods since it generates charged ions while penetrating through the target organisms, and it has enough energy to cause biological effects. However, before the wide utilization of ionizing priming methods in agriculture, extensive research is needed to explore their effects on seed priming and to focus on the underlying mechanism of them. Overall, this review aims to highlight the current understanding of ionizing priming methods and their applicability for promoting agroecological resilience and meeting the challenges of food crises nowadays.

Genome and transcriptome-based characterization of high energy carbon-ion beam irradiation induced delayed flower senescence mutant in Lotus japonicus.

BACKGROUND: Flower longevity is closely related to pollen dispersal and reproductive success in all plants, as well as the commercial value of ornamental plants. Mutants that display variation in flower longevity are useful tools for understanding the mechanisms underlying this trait. Heavy-ion beam irradiation has great potential to improve flower shapes and colors; however, few studies are available on the mutation of flower senescence in leguminous plants. RESULTS: A mutant (C416) exhibiting blossom duration eight times longer than that of the wild type (WT) was isolated in Lotus japonicus derived from carbon ion beam irradiation. Genetic assays supported that the delayed flower senescence of C416 was a dominant trait controlled by a single gene, which was located between 4,616,611 Mb and 5,331,876 Mb on chromosome III. By using a sorting strategy of multi-sample parallel genome sequencing, candidate genes were narrowed to the gene CUFF.40834, which exhibited high identity to ethylene receptor 1 in other model plants. A physiological assay demonstrated that C416 was insensitive to ethylene precursor. Furthermore, the dynamic changes of phytohormone regulatory network in petals at different developmental stages was compared by using RNA-seq. In brief, the ethylene, jasmonic acid (JA), and salicylic acid (SA) signaling pathways were negatively regulated in C416, whereas the brassinosteroid (BR) and cytokinin signaling pathways were positively regulated, and auxin exhibited dual effects on flower senescence in Lotus japonicus. The abscisic acid (ABA) signaling pathway is positively regulated in C416. CONCLUSION: So far, C416 might be the first reported mutant carrying a mutation in an endogenous ethylene-related gene in Lotus japonicus, rather than through the introduction of exogenous genes by transgenic techniques. A schematic of the flower senescence of Lotus japonicus from the perspective of the phytohormone regulatory network was provided based on transcriptome profiling of petals at different developmental stages. This study is informative for elucidating the molecular mechanism of delayed flower senescence in C416, and lays a foundation for candidate flower senescence gene identification in Lotus japonicus. It also provides another perspective for the improvement of flower longevity in legume plants by heavy-ion beam.

Response characteristics of the membrane integrity and physiological activities of the mutant strain Y217 under exogenous butanol stress.

Butanol inhibits bacterial activity by destroying the cell membrane of Clostridium acetobutylicum strains and altering functionality. Butanol toxicity also results in destruction of the phosphoenolpyruvate-carbohydrate phosphotransferase system (PTS), thereby preventing glucose transport and phosphorylation and inhibiting transmembrane transport and assimilation of sugars, amino acids, and other nutrients. In this study, based on the addition of exogenous butanol, the tangible macro indicators of changes in the carbon ion beam irradiation-mutant Y217 morphology were observed using scanning electron microscopy (SEM). The mutant has lower microbial adhesion to hydrocarbon (MATH) value than C. acetobutylicum ATCC 824 strain. FDA fluorescence intensity and conductivity studies demonstrated the intrinsically low membrane permeability of the mutant membrane, with membrane potential remaining relatively stable. Monounsaturated FAs (MUFAs) accounted for 35.17% of the mutant membrane, and the saturated fatty acids (SFA)/unsaturated fatty acids (UFA) ratio in the mutant cell membrane was 1.65. In addition, we conducted DNA-level analysis of the mutant strain Y217. Expectedly, through screening, we found gene mutant sites encoding membrane-related functions in the mutant, including ATP-binding cassette (ABC) transporter-related genes, predicted membrane proteins, and the PTS transport system. It is noteworthy that an unreported predicted membrane protein (CAC 3309) may be related to changes in mutant cell membrane properties. KEY POINTS: • Mutant Y217 exhibited better membrane integrity and permeability. • Mutant Y217 was more resistant to butanol toxicity. • Some membrane-related genes of mutant Y217 were mutated.

Repair characteristics and time-dependent effects in response to heavy-ion beam irradiation in Saccharomyces cerevisiae: a comparison with X-ray irradiation.

Heavy-ion beam (HIB) irradiation has been widely used in microbial mutation breeding. However, a global cellular response to such radiation remains mostly uncharacterised. In this study, we used transcriptomics to analyse the damage repair response in Saccharomyces cerevisiae following a semi-lethal HIB irradiation (80 Gy), which induced a significant number of DNA double-strand breaks. Our analysis of differentially expressed genes (DEGs) from 50 to 150 min post-irradiation revealed that upregulated genes were significantly enriched for gene ontology and Kyoto encyclopaedia of genes and genomes terms related to damage repair response. Based on the number of DEGs, their annotation, and their relative expression, we established that the peak of the damage repair response occurred 75 min post-irradiation. Moreover, we exploited the data from our recent study on X-ray irradiation-induced repair to compare the transcriptional patterns induced by semi-lethal HIB and X-ray irradiations. Although these two radiations have different properties, we found a significant overlap (> 50%) for the DEGs associated with five typical DNA repair pathways and, in both cases, identified homologous recombination repair (HRR) as the predominant repair pathway. Nevertheless, when we compared the relative enrichment of the five DNA repair pathways at the key time point of the repair process, we found that the relative enrichment of HRR was higher after HIB irradiation than after X-ray irradiation. Additionally, the peak stage of HRR following HIB irradiation was ahead of that following X-ray irradiation. Since mutations occur during the DNA repair process, uncovering detailed repair characteristics should further the understanding of the associated mutagenesis features.

Quantitative multi-omics analysis of the effects of mitochondrial dysfunction on lipid metabolism in Saccharomyces cerevisiae.

In this study, combined genome, transcriptome, and metabolome analysis was performed for eight Saccharomyces cerevisiae mitochondrial respiration-deficient mutants. Each mutant exhibited a unique nuclear genome mutation pattern; the nuclear genome mutations, and thus potentially affected genes and metabolic pathways, showed a co-occurrence frequency of ≤ 3 among the eight mutants. For example, only a lipid metabolism-related pathway was likely to be affected by the nuclear genome mutations in one of the mutants. However, large deletions in the mitochondrial genome were the shared characteristic among the eight mutants. At the transcriptomic level, lipid metabolism was the most significantly enriched Kyoto Encyclopaedia of Genes and Genomes (KEGG) pathway for differentially expressed genes (DEGs) co-occurring in both ≥ 4 and ≥ 5 mutants. Any identified DEG enriched in lipid metabolism showed the same up-/down-regulated pattern among nearly all eight mutants. Further, 126 differentially expressed lipid species (DELS) were identified, which also showed the same up-/down-regulated pattern among nearly all investigated mutants. It was conservatively demonstrated that the similar change pattern of lipid metabolism in the entire investigated mutant population was attributed to mitochondrial dysfunction. The change spectrum of lipid species was presented, suggesting that the number and change degree of up-regulated lipid species were higher than those of down-regulated lipid species. Additionally, energy storage lipids increased in content and plasma-membrane phospholipid compositions varied in the relative proposition. The results for the genome, transcriptome, and lipidome were mutually validated, which provides quantitative data revealing the roles of mitochondria from a global cellular perspective.

Smad7 enables STAT3 activation and promotes pluripotency independent of TGF-ß signaling.

Smad7 is a negative feedback product of TGF-ß superfamily signaling and fine tunes a plethora of pleiotropic responses induced by TGF-ß ligands. However, its noncanonical functions independent of TGF-ß signaling remain to be elucidated. Here, we show that Smad7 activates signal transducers and activators of transcription 3 (STAT3) signaling in maintaining mouse embryonic stem cell pluripotency in a manner independent of the TGF-ß receptors, yet dependent on the leukemia inhibitory factor (LIF) coreceptor glycoprotein 130 (gp130). Smad7 directly binds to the intracellular domain of gp130 and disrupts the SHP2-gp130 or SOCS3-gp130 complex, thereby amplifying STAT3 activation. Consequently, Smad7 facilitates LIF-mediated self-renewal of mouse ESCs and is also critical for induced pluripotent stem cell reprogramming. This finding illustrates an uncovered role of the Smad7-STAT3 interplay in maintaining cell pluripotency and also implicates a mechanism involving Smad7 underlying cytokine-dependent regulation of cancer and inflammation.

Respiratory Syncytial Virus Replication Is Promoted by Autophagy-Mediated Inhibition of Apoptosis.

Respiratory syncytial virus (RSV) is the main cause of acute lower respiratory tract infection (ALRI) in children worldwide. Virus-host interactions affect the progression and prognosis of the infection. Autophagy plays important roles in virus-host interactions. Respiratory epithelial cells serve as the front line of host defense during RSV infection, However, it is still unclear how they interact with RSV. In this study, we found that RSV induced autophagy that favored RSV replication and exacerbated lung pathology in vivo Mechanistically, RSV induced complete autophagy flux through reactive oxygen species (ROS) generation and activation of the AMP-activated protein kinase/mammalian target of rapamycin (AMPK-MTOR) signaling pathway in HEp-2 cells. Furthermore, we evaluated the functions of autophagy in RSV replication and found that RSV replication was increased in HEp-2 cells treated with rapamycin but decreased remarkably in cells treated with 3-methylademine (3-MA) or wortmannin. Knockdown key molecules in the autophagy pathway with short hairpinp RNA (shRNA) against autophagy-related gene 5 (ATG5), autophagy-related gene 7 (ATG7), or BECN1/Beclin 1 or treatment with ROS scavenger N-acetyl-l-cysteine (NAC) and AMPK inhibitor (compound C) suppressed RSV replication. 3-MA or shATG5/BECN1 significantly decreased cell viability and increased cell apoptosis at 48 hours postinfection (hpi). Blocking apoptosis with Z-VAD-FMK partially restored virus replication at 48 hpi. Those results provide strong evidence that autophagy may function as a proviral mechanism in a cell-intrinsic manner during RSV infection.IMPORTANCE An understanding of the mechanisms that respiratory syncytial virus utilizes to interact with respiratory epithelial cells is critical to the development of novel antiviral strategies. In this study, we found that RSV induces autophagy through a ROS-AMPK signaling axis, which in turn promotes viral infection. Autophagy favors RSV replication through blocking cell apoptosis at 48 hpi. Mechanistically, RSV induces mitophagy, which maintains mitochondrial homeostasis and therefore decreases cytochrome c release and apoptosis induction. This study provides a novel insight into this virus-host interaction, which may help to exploit new antiviral treatments targeting autophagy processes.

Current strategies and future prospects for enhancing microbial production of citric acid.

Aspergillus niger and Yarrowia lipolytica are highly important in citric acid (CA) production. To further minimize the cost of CA bio-production using A. niger and Y. lipolytica, some strategies (e.g., metabolic engineering, efficient mutagenesis, and optimal fermentation strategies) were developed to enhance CA production and low-cost carbon sources were also utilized to decrease CA bio-production cost. In this review, we summarize the recent significant progresses in CA bio-production, including metabolic engineering, efficient mutagenesis and screening methods, optimal fermentation strategies, and use of low-cost carbon sources, and future prospects in this field are also discussed, which could help in the development of CA production industry.

A genome-wide view of mutations in respiration-deficient mutants of Saccharomyces cerevisiae selected following carbon ion beam irradiation.

Mitochondrial dysfunction in Saccharomyces cerevisiae was selected as a marker of ion penetration following carbon ion beam (CIB) irradiation. Respiration-deficient mutants were screened. Following confirmation of negligible spontaneous mutation, eight genetically stable S. cerevisiae respiration-deficient mutant strains and a control strain were resequenced with ~ 200-fold read depth. Strategies were established to identify and validate the particular mutations induced by CIB irradiation. In the nuclear genome, CIB irradiation mainly caused base substitutions and some small (< 100 bp) insertions/deletions (indels), which were widely distributed across the chromosomes. Although mitochondrial dysfunction was selected as a screening marker, variants in the nuclear genome were detected at a high frequency (10-7) relative to spontaneous mutations (10-9). The transition to transversion ratio for base substitutions was 0.746, which was less than that of spontaneous mutations. In the mitochondrial genome, there were very large deletions including substantial gene areas, resulting in extremely low read coverage. Meanwhile, every mutant possessed a distinctive mutation pattern, for both the nuclear and the mitochondrial genome. Nuclear genomes contained scanty mitochondrial respiration-related genes that were potentially affected by verified mutations, suggesting that variants in the mitochondrial genome may be the main drivers of respiratory deficiencies. Our study confirmed the previous finding that heavy ion beam (HIB) irradiation mainly induces substantial base substitutions and some small indels but also yielded some novel findings, in particular, novel structural variants in the mitochondrial genomes. These data will enhance the understanding of HIB-induced damage and mutations and aid in the HIB-based microbial mutation breeding.

CpG in Combination with an Inhibitor of Notch Signaling Suppresses Formalin-Inactivated Respiratory Syncytial Virus-Enhanced Airway Hyperresponsiveness and Inflammation by Inhibiting Th17 Memory Responses and Promoting Tissue-Resident Memory Cells in Lungs.

Respiratory syncytial virus (RSV) is the leading cause of childhood hospitalizations. The formalin-inactivated RSV (FI-RSV) vaccine-enhanced respiratory disease (ERD) has been an obstacle to the development of a safe and effective killed RSV vaccine. Agonists of Toll-like receptor (TLR) have been shown to regulate immune responses induced by FI-RSV. Notch signaling plays critical roles during the differentiation and effector function phases of innate and adaptive immune responses. Cross talk between TLR and Notch signaling pathways results in fine-tuning of TLR-triggered innate inflammatory responses. We evaluated the impact of TLR and Notch signaling on ERD in a murine model by administering CpG, an agonist of TLR9, in combination with L685,458, an inhibitor of Notch signaling during FI-RSV immunization. Activation with CpG or deficiency of MyD88-dependent TLR signaling did not alleviate airway inflammation in FI-RSV-immunized mice. Activation or inhibition of Notch signaling with Dll4, one of the Notch ligands, or L685,458 did not suppress FI-RSV-enhanced airway inflammation either. However, the CpG together with L685,458 markedly inhibited FI-RSV-enhanced airway hyperresponsiveness, weight loss, and lung inflammation. Interestingly, CpG plus L685,458 completely inhibited FI-RSV-associated Th17 and Th17-associated proinflammatory chemokine responses in lungs following RSV challenge but not Th1 or Th2, memory responses. In addition, FI-RSV plus CpG plus L685,458 promoted protective CD8+ lung tissue-resident memory (TRM) cells. These results indicate that activation of TLR signaling combined with inhibition of Notch signaling prevent FI-RSV ERD, and the mechanism appears to involve suppressing proinflammatory Th17 memory responses and promoting protective TRM in lungs.IMPORTANCE RSV is the most important cause of lower respiratory tract infections in infants. The FI-RSV-enhanced respiratory disease (ERD) is a major impediment to the development of a safe and effective killed RSV vaccine. Using adjuvants to regulate innate and adaptive immune responses could be an effective method to prevent ERD. We evaluated the impact of TLR and Notch signaling on ERD by administering CpG, an agonist of TLR9, in combination with L685,458, an inhibitor of Notch signaling, during FI-RSV immunization. The data showed that treatment of TLR or Notch signaling alone did not suppress FI-RSV-enhanced airway inflammation, while CpG plus L685,458 markedly inhibited ERD. The mechanism appears to involve suppressing Th17 memory responses and promoting tissue-resident memory cells. Moreover, these results suggest that regulation of lung immune memory with adjuvant compounds containing more than one immune-stimulatory molecule may be a good strategy to prevent FI-RSV ERD.