The benzoylphenylurea derivative BPU17 acts as an inhibitor of prohibitin and exhibits antifibrotic activity.

Inflammation-induced choroidal neovascularization followed by the epithelial-mesenchymal transition (EMT) of retinal pigment epithelial cells (RPEs) is a cause of neovascular age-related macular degeneration (nAMD). RPE-derived myofibroblasts overproduce extracellular matrix, leading to subretinal fibrosis. We already have demonstrated that benzylphenylurea (BPU) derivatives inhibit the function of cancer-associated fibroblasts. Here, we investigated the anti-myofibroblast effects of BPU derivatives and examined such BPU activity on subretinal fibrosis. A BPU derivative, BPU17, exhibits the most potent anti-myofibroblast activity among dozens of BPU derivatives and inhibits subretinal fibrosis in a mouse model of retinal degeneration. Investigations with primary cultured RPEs reveal that BPU17 suppresses cell motility and collagen synthesis in RPE-derived myofibroblasts. These effects depend on repressing the serum response factor (SRF)/CArG-box-dependent transcription. BPU17 inhibits the expression of SRF cofactor, cysteine and glycine-rich protein 2 (CRP2), which activates the SRF function. Proteomics analysis reveals that BPU17 binds to prohibitin 1 (PHB1) and inhibits the PHB1-PHB2 interaction, resulting in mild defects in mitochondrial function. This impairment causes a decrease in the expression of CRP2 and suppresses collagen synthesis. Our findings suggest that BPU17 is a promising agent against nAMD and the close relationship between PHB function and EMT.

Role of CRP2-MRTF interaction in functions of myofibroblasts.

Inflammatory response induces phenotypic modulation of fibroblasts into myofibroblasts. Although transforming growth factor-ßs (TGF-ßs) evoke such transition, the details of the mechanism are still unknown. Here, we report that a LIM domain protein, cysteine-and glycine-rich protein 2 (CSRP2 [CRP2]) plays a vital role in the functional expression profile in myofibroblasts and cancer-associated fibroblasts (CAFs). Knock-down of CRP2 severely inhibits the expression of smooth muscle cell (SMC) genes, cell motility, and CAF-mediated collective invasion of epidermoid carcinoma. We elucidate the following molecular bases: CRP2 directly binds to myocardin-related transcription factors (MRTF-A/B [MRTFs]) and serum response factor (SRF) and stabilizes the MRTF/SRF/CArG-box complex to activate SMC gene expression. Furthermore, a three-dimensional structural analysis of CRP2 identifies the amino acids required for the CRP2-MRTF-A interaction. Polar amino acids in the C-terminal half (serine-152, glutamate-154, serine-155, threonine-156, threonine-157, and threonine-159 in human CRP2) are responsible for direct binding to MRTF-A. On the other hand, hydrophobic amino acids outside the consensus sequence of the LIM domain (tryptophan-139, phenylalanine-144, leucine-153, and leucine-158 in human CRP2) play a role in stabilizing the unique structure of the LIM domain.Key words: CRP2, 3D structure, myocardin-related transcription factor, myofibroblast, cancer-associated fibroblasts.

Development of a Genetically Stable Live Attenuated Influenza Vaccine Strain Using an Engineered High-Fidelity Viral Polymerase.

RNA viruses demonstrate a vast range of variants, called quasispecies, due to error-prone replication by viral RNA-dependent RNA polymerase. Although live attenuated vaccines are effective in preventing RNA virus infection, there is a risk of reversal to virulence after their administration. To test the hypothesis that high-fidelity viral polymerase reduces the diversity of influenza virus quasispecies, resulting in inhibition of reversal of the attenuated phenotype, we first screened for a high-fidelity viral polymerase using serial virus passages under selection with a guanosine analog ribavirin. Consequently, we identified a Leu66-to-Val single amino acid mutation in polymerase basic protein 1 (PB1). The high-fidelity phenotype of PB1-L66V was confirmed using next-generation sequencing analysis and biochemical assays with the purified influenza viral polymerase. As expected, PB1-L66V showed at least two-times-lower mutation rates and decreased misincorporation rates, compared to the wild type (WT). Therefore, we next generated an attenuated PB1-L66V virus with a temperature-sensitive (ts) phenotype based on FluMist, a live attenuated influenza vaccine (LAIV) that can restrict virus propagation by ts mutations, and examined the genetic stability of the attenuated PB1-L66V virus using serial virus passages. The PB1-L66V mutation prevented reversion of the ts phenotype to the WT phenotype, suggesting that the high-fidelity viral polymerase could contribute to generating an LAIV with high genetic stability, which would not revert to the pathogenic virus.IMPORTANCE The LAIV currently in use is prescribed for actively immunizing individuals aged 2 to 49 years. However, it is not approved for infants and elderly individuals, who actually need it the most, because it might prolong virus propagation and cause an apparent infection in these individuals, due to their weak immune systems. Recently, reversion of the ts phenotype of the LAIV strain currently in use to a pathogenic virus was demonstrated in cultured cells. Thus, the generation of mutations associated with enhanced virulence in LAIV should be considered. In this study, we isolated a novel influenza virus strain with a Leu66-to-Val single amino acid mutation in PB1 that displayed a significantly higher fidelity than the WT. We generated a novel LAIV candidate strain harboring this mutation. This strain showed higher genetic stability and no ts phenotype reversion. Thus, our high-fidelity strain might be useful for the development of a safer LAIV.

A novel method for transforming Geobacillus kaustophilus with a chromosomal segment of Bacillus subtilis transferred via pLS20-dependent conjugation.

BACKGROUND: Geobacillus kaustophilus is a thermophilic Gram-positive bacterium. Methods for its transformation are still under development. Earlier studies have demonstrated that pLS20catΔoriT mobilized the resident mobile plasmids from Bacillus subtilis to G. kaustophilus and transferred long segments of chromosome from one cell to another between B. subtilis. RESULTS: In this study, we applied mobilization of the B. subtilis chromosome mediated by pLS20catΔoriT to transform G. kaustophilus. We constructed a gene cassette to be integrated into G. kaustophilus and designed it within the B. subtilis chromosome. The pLS20catΔoriT-mediated conjugation successfully transferred the gene cassette from the B. subtilis chromosome into the G. kaustophilus allowing for the desired genetic transformation. CONCLUSIONS: This transformation approach described here will provide a new tool to facilitate the flexible genetic manipulation of G. kaustophilus.

Clinical Implication of Smoking-Related Aryl-Hydrocarbon Receptor Repressor (AHRR) Hypomethylation in Japanese Adults.

BACKGROUND: Tobacco smoking is a leading preventable cause of morbidity and mortality worldwide; still, the success rate of smoking cessation is low in general. From the viewpoint of public health and clinical care, an objective biomarker of long-term smoking behavior is sought.Methods and Results: This study assessed DNA methylation as a biomarker of smoking in a hospital setting through a combination of molecular approaches including genetic, DNA methylation and mRNA expression analyses. First, in an epigenome-wide association study involving Japanese individuals with chronic cardiovascular disease (n=94), genome-wide significant smoking association was identified at 2 CpG sites on chromosome 5, with the strongest signal at cg05575921 located in intron 3 of the aryl-hydrocarbon receptor repressor (AHRR) gene. Highly significant (P<1×10-27) smoking-cg05575921 association was validated in 2 additional panels (n=339 and n=300). For the relationship of cg05575921 methylation extent with time after smoking cessation and cumulative cigarette consumption among former smokers, smoking-related hypomethylation was found to remain for ≥20 years after smoking cessation and to be affected by multiple factors, such as cis-interaction of genetic variation. There was a significant inverse correlation (P=0.0005) between cg05575921 methylation extent and AHRR mRNA expression. CONCLUSIONS: The present study results support that reversion of AHRR hypomethylation can be a quantifiable biomarker for progress in and observance of smoking cessation, although some methodological points need to be considered.

Multisystem Inflammatory Syndrome in Children　- A New Syndrome Complicated With Acute Heart Failure Following Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) Infection.

BACKGROUND: Multisystem inflammatory syndrome in children (MIS-C) is a rare syndrome temporally related to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). MIS-C shares similarities with Kawasaki disease, but left ventricular dysfunction is more common in MIS-C.Methods and Results:This study reports the case of a 16-year-old Japanese male patient with MIS-C. Although the initial presentation was severe with circulatory and respiratory failure, the patient recovered completely. Endomyocardial biopsy showed active myocarditis with fibrosis. Immunoglobulin treatment was useful for recovery. CONCLUSIONS: This is the first reported case of MIS-C in Japan. Cardiologists should be aware of MIS-C, a new disease, occurring during the global SARS-CoV-2 pandemic.

Case Report of Left Ventricular Noncompaction Cardiomyopathy Characterized by Undulating Phenotypes in Adult Patients.

Left ventricular noncompaction cardiomyopathy (LVNC) is a heart muscle disorder morphologically characterized by reticulated trabeculations and intertrabecular recesses in the left ventricular (LV) cavity. LVNC is a genetically and phenotypically heterogeneous condition, which has been increasingly recognized with the accumulation of evidence provided by genotype-phenotype correlation analyses. Here, we report 2 sporadic adult cases of LVNC; both developed acute heart failure as an initial clinical manifestation and harbored causal sarcomere gene mutations. One case was a 57-year-old male with digenic heterozygote mutations, p.R1344Q in myosin heavy chain 7 (MYH7) and p.R144W in troponin T2, cardiac type (TNNT2), who showed morphological characteristics of LVNC in the lateral to apical regions of the LV together with a comorbidity of non-transmural myocardial infarction, resulting from a coronary artery stenosis. After the removal of ischemic insult and standard heart failure treatment, LVNC became less clear, and LV function gradually improved. The other case was a 36-year-old male with a heterozygote mutation, p.E334K in myosin binding protein C3 (MYBPC3), who exhibited cardiogenic shock on admission with morphological characteristics of LVNC being most prominent in the apical segment of the LV. The dosage of beta-blocker was deliberately increased in an outpatient clinic over 6 months following hospitalization, which remarkably improved the LV ejection fraction from 21% to 54.3%. Via a combination of imaging and histopathological and genetic tests, we have found that these cases are not compatible with a persistent phenotype of primary cardiomyopathy, but their morphological features are changeable in response to treatment. Thus, we point out phenotypic plasticity or undulation as a noticeable element of LVNC in this case report.

Tyr82 Amino Acid Mutation in PB1 Polymerase Induces an Influenza Virus Mutator Phenotype.

In various positive-sense single-stranded RNA viruses, a low-fidelity viral RNA-dependent RNA polymerase (RdRp) confers attenuated phenotypes by increasing the mutation frequency. We report a negative-sense single-stranded RNA virus RdRp mutant strain with a mutator phenotype. Based on structural data of RdRp, rational targeting of key residues, and screening of fidelity variants, we isolated a novel low-fidelity mutator strain of influenza virus that harbors a Tyr82-to-Cys (Y82C) single-amino-acid substitution in the PB1 polymerase subunit. The purified PB1-Y82C polymerase indeed showed an increased frequency of misincorporation compared with the wild-type PB1 in an in vitro biochemical assay. To further investigate the effects of position 82 on PB1 polymerase fidelity, we substituted various amino acids at this position. As a result, we isolated various novel mutators other than PB1-Y82C with higher mutation frequencies. The structural model of influenza virus polymerase complex suggested that the Tyr82 residue, which is located at the nucleoside triphosphate entrance tunnel, may influence a fidelity checkpoint. Interestingly, although the PB1-Y82C variant replicated with wild-type PB1-like kinetics in tissue culture, the 50% lethal dose of the PB1-Y82C mutant was 10 times lower than that of wild-type PB1 in embryonated chicken eggs. In conclusion, our data indicate that the Tyr82 residue of PB1 has a crucial role in regulating polymerase fidelity of influenza virus and is closely related to attenuated pathogenic phenotypes in vivoIMPORTANCE Influenza A virus rapidly acquires antigenic changes and antiviral drug resistance, which limit the effectiveness of vaccines and drug treatments, primarily owing to its high rate of evolution. Virus populations formed by quasispecies can contain resistance mutations even before a selective pressure is applied. To study the effects of the viral mutation spectrum and quasispecies, high- and low-fidelity variants have been isolated for several RNA viruses. Here, we report the discovery of a low-fidelity RdRp variant of influenza A virus that contains a substitution at Tyr82 in PB1. Viruses containing the PB1-Y82C substitution showed growth kinetics and viral RNA synthesis levels similar to those of the wild-type virus in cell culture; however, they had significantly attenuated phenotypes in a chicken egg infection experiment. These data demonstrated that decreased RdRp fidelity attenuates influenza A virus in vivo, which is a desirable feature for the development of safer live attenuated vaccine candidates.

Molecular monitoring of Neoseiulus californicus released from sheltered slow-release sachets for spider mite control in a Japanese pear greenhouse.

A novel system for spider mite control was developed with a slow-release sachet containing Neoseiulus californicus (McGregor) (Acari: Phytoseiidae) protected by a waterproof shelter. Monitoring the efficacy of the predator release system for spider mite control at a Japanese pear greenhouse requires discrimination of N. californicus from other indigenous phytoseiid mite species inhabiting the study site and subsequent identification of the released N. californicus. The report of our earlier study described a PCR-based method for discrimination of N. californicus species. For the present study, we first examined phytoseiid mite species composition in the greenhouse. Subsequently, we developed microsatellite markers to identify the released N. californicus. Finally, we installed the predator release system in the greenhouse and conducted a population survey of phytoseiid and spider mites. Results demonstrated that approximately 1 month is necessary for distribution of the released N. californicus on the leaves.

Generation of a Genetically Stable High-Fidelity Influenza Vaccine Strain.

Vaccination is considered the most effective preventive means for influenza control. The development of a master virus with high growth and genetic stability, which may be used for the preparation of vaccine viruses by gene reassortment, is crucial for the enhancement of vaccine performance and efficiency of production. Here, we describe the generation of a high-fidelity and high-growth influenza vaccine master virus strain with a single V43I amino acid change in the PB1 polymerase of the high-growth A/Puerto Rico/8/1934 (PR8) master virus. The PB1-V43I mutation was introduced to increase replication fidelity in order to design an H1N1 vaccine strain with a low error rate. The PR8-PB1-V43I virus exhibited good replication compared with that of the parent PR8 virus. In order to compare the efficiency of egg adaptation and the occurrence of gene mutations leading to antigenic alterations, we constructed 6:2 genetic reassortant viruses between the A(H1N1)pdm09 and the PR8-PB1-V43I viruses; hemagglutinin (HA) and neuraminidase (NA) were from the A(H1N1)pdm09 virus, and the other genes were from the PR8 virus. Mutations responsible for egg adaptation mutations occurred in the HA of the PB1-V43I reassortant virus during serial egg passages; however, in contrast, antigenic mutations were introduced into the HA gene of the 6:2 reassortant virus possessing the wild-type PB1. This study shows that the mutant PR8 virus possessing the PB1 polymerase with the V43I substitution may be utilized as a master virus for the generation of high-growth vaccine viruses with high polymerase fidelity, low error rates of gene replication, and reduced antigenic diversity during virus propagation in eggs for vaccine production.IMPORTANCE Vaccination represents the most effective prophylactic option against influenza. The threat of emergence of influenza pandemics necessitates the ability to generate vaccine viruses rapidly. However, as the influenza virus exhibits a high mutation rate, vaccines must be updated to ensure a good match of the HA and NA antigens between the vaccine and the circulating strain. Here, we generated a genetically stable master virus of the A/Puerto Rico/8/1934 (H1N1) backbone encoding an engineered high-fidelity viral polymerase. Importantly, following the application of the high-fidelity PR8 backbone, no mutation resulting in antigenic change was introduced into the HA gene during propagation of the A(H1N1)pdm09 candidate vaccine virus. The low error rate of the present vaccine virus should decrease the risk of generating mutant viruses with increased virulence. Therefore, our findings are expected to be useful for the development of prepandemic vaccines and live attenuated vaccines with higher safety than that of the present candidate vaccines.

A novel method for transforming the thermophilic bacterium Geobacillus kaustophilus.

BACKGROUND: Bacterial strains of the genus Geobacillus grow at high temperatures of 50-75 °C and could thus be useful for biotechnological applications. However, genetic manipulation of these species is difficult because the current techniques for transforming Geobacillus species are not efficient. In this study, we developed an easy and efficient method for transforming Geobacillus kaustophilus using the conjugative plasmid pLS20cat. RESULTS: We constructed a transformation system comprising (i) a mobilizable Bacillus subtilis-G. kaustophilus shuttle plasmid named pGK1 that carries the elements for selection and replication in Geobacillus, and (ii) a pLS20cat-harboring B. subtilis donor strain expressing the dam methylase gene of Escherichia coli and the conjugation-stimulating rapLS20 gene of pLS20cat. This system can be used to efficiently introduce pGK1 into G. kaustophilus by mobilization in a pLS20cat-dependent way. Whereas the thermostable kanamycin marker and Geobacillus replication origin of pGK1 as well as expression of dam methylase in the donor were indispensable for mobilization, ectopic expression of rapLS20 increased its efficiency. In addition, the conditions of the recipient influenced mobilization efficiency: the highest mobilization efficiencies were obtained using recipient cells that were in the exponential growth phase. Furthermore, elimination of the origin of transfer from pLS20cat enhanced the mobilization. CONCLUSIONS: We describe a novel method of plasmid mobilization into G. kaustophilus recipient from B. subtilis donor depending on the helper function of pLS20cat, which enables simple, rapid, and easy transformation of the thermophilic Gram-positive bacterium.

Variation in nesting behavior of eight species of spider mites, Stigmaeopsis having sociality.

Nesting behavior is considered to be an important element of social living in animals. The spider mites belonging to the genus Stigmaeopsis spend their lives within nests produced from silk threads. Several of these species show cooperative sociality, while the others are subsocial. In order to identify the origins of this social behavior, comparisons of nest sizes, nesting behaviors (making nests continuously or separately), and their associated traits (fecal deposition patterns) were made for eight cogeneric Stigmaeopsis species showing various levels of social development. All of these species inhabit bamboo plants (Poaceae). We initially addressed the proximate factor of nest size variation. The variation in nest size of the eight species corresponded well with the variation in dorsal seta sc1 length, suggesting that nest size variation among species may have a genetic basis. The time spent within a nest (nest duration) increased with nest size on the respective host plants. Nest arrangement patterns varied among species showing different sized nests: Large nest builders continuously extended their nests, while middle and small nest-building species built new separate nests, which resulted in different social interaction times among species, and is thought to be closely related to social development. Fecal deposition behaviors also varied among Stigmaeopsis species, suggesting diversity in anti-predatory adaptations. Finally, we discuss how the variation in sociality observed within this genus is likely the result of nest size variation that initially evolved as anti-predator strategies.

Quantitative structure-activity relationship of 2,6-dimethoxy-N-(3-(4-substituted phenyl)isoxazol-5-yl)benzamide for the inhibition of chitin synthesis.

Previously, we found that 5-(2,6-dimethoxybenzoylamino)-3-phenylisoxazoles (IOXs) inhibit chitin synthesis in the cultured integument of Chilo suppressalis. In this study, IOXs with various substituents at the para-position of the 3-phenyl ring were synthesized, and the concentrations required to inhibit chitin synthesis to 50% (IC50) were determined for all compounds. The introduction of halogens-such as F, Cl, and Br-and small alkyls-such as Me, Et, Pr, and n-Bu-at the 3-phenyl ring slightly enhanced the activity. However, the activity decreased drastically with the introduction of NO2, CF3, and t-Bu. The quantitative analysis of the substituent effect at the 3-phenyl ring on chitin-synthesis inhibition using the Hansch-Fujita method showed that the hydrophobic substituent with the optimum value was favored for the activity, but the bulky substituent in terms of E s was detrimental to the activity.

Novel ability of diflubenzuron as an inhibitor of mitochondrial function.

Compounds classified as benzoylphenylurea (BPU), such as diflubenzuron (DFB), are used as insecticides. Although BPU disrupts molting by inhibiting chitin biosynthesis and exhibits insecticidal activity, their exact mode of action remains unknown. Since epidermal cells proliferate and morphologically change from squamous to columnar cells during the early stages of insect molting, we speculate that a transition similar to that from epithelium to mesenchyme occurs and that BPU may inhibit this transition. Here, we addressed this possibility. We found that DFB decreases actin expression in insect cells (the tissue cultures of insect integument). Detailed analysis in Schneider S2 cells reveals that DFB inhibits the expression of actin isoforms (Act5C and Act42A) and the Drosophila ortholog of myocardin-related transcription factor (Mrtf), leading to cell growth suppression. Proteomics identified the Drosophila ortholog of prohibitin (Phb1D and Phb2E) as one of the DFB-binding proteins. DFB inhibits the interaction between Phb1D and Phb2E and induces mitochondrial dysfunction. The knock-down of Phb2E suppresses the expression of Act5C, Act42A, and Mrtf, leading to cell growth inhibition. Thus, the disruption of Phb function is a possible novel target of DFB.

Outcomes of critically ill patients with liver failure who require mechanical ventilation: A retrospective, single-center study.

Background and Aims: Critically ill patients with liver failure have high mortality. Besides the management of organ-specific complications, liver transplantation constitutes a definitive treatment. However, clinicians may hesitate to introduce mechanical ventilation for patients on liver transplantation waitlists because of poor prognosis. This study investigated the outcomes of intensive care and ventilation support therapy effects in patients with liver failure. Methods: This single-center study retrospectively enrolled 32 consecutive patients with liver failure who were admitted to the intensive care unit from January 2014 to December 2020. The medical records were reviewed and analyzed retrospectively for Acute Physiologic and Chronic Health Evaluation (APACHE)-II. The model for end-stage liver disease scores, 90-day mortality, and survival was assessed using the Kaplan-Meier method. Results: The average patient age was 45.5 ± 20.1 years, and 53% of patients were women. On intensive care unit admission, APACHE-II and model for end-stage liver disease scores were 20 and 28, respectively. Among 13 patients considered for liver transplantation, 4 received transplants. Thirteen patients (40.6%) were intubated and mechanically ventilated in the intensive care unit. The 90-day mortality rate of patients with and without mechanical ventilation in the intensive care unit (13, 61.5% vs. 19, 47.4%, p = 0.4905) was similar. APACHE-II score >21 was an independent predictor of mechanical ventilation requirement in patients with liver failure during intensive care unit stay. Conclusion: Although critically ill patients with liver failure are at risk of multiorgan failure with poor outcomes, mechanical ventilation did not negatively affect the 90-day mortality or performance rates of liver transplantation. Clinicians should consider mechanical ventilation-based life support in critically ill patients with liver failure who are awaiting liver transplantation.

Ferroelectric mesocrystals of bismuth sodium titanate: formation mechanism, nanostructure, and application to piezoelectric materials.

Ferroelectric mesocrystals of Bi0.5Na0.5TiO3 (BNT) with [100]-crystal-axis orientation were successfully prepared using a topotactic structural transformation process from a layered titanate H1.07Ti1.73O4·nH2O (HTO). The formation reactions of BNT mesocrystals in HTO-Bi2O3-Na2CO3 and HTO-TiO2-Bi2O3-Na2CO3 reaction systems and their nanostructures were studied by XRD, FE-SEM, TEM, SAED, and EDS, and the reaction mechanisms were given. The BNT mesocrystals are formed by a topotactic structural transformation mechanism in the HTO-Bi2O3-Na2CO3 reaction system and by a combination mechanism of the topotactic structural transformation and epitaxial crystal growth in the HTO-TiO2-Bi2O3-Na2CO3 reaction system, respectively. The BNT mesocrystals prepared by these methods are constructed from [100]-oriented BNT nanocrystals. Furthermore, these reaction systems were successfully applied to the fabrication of [100]-oriented BNT ferroelectric ceramic materials. A BNT ceramic material with a high degree of orientation, high relative density, and small grain size was achieved.

Transition from positive to neutral in mutation fixation along with continuing rising fitness in thermal adaptive evolution.

It remains to be determined experimentally whether increasing fitness is related to positive selection, while stationary fitness is related to neutral evolution. Long-term laboratory evolution in Escherichia coli was performed under conditions of thermal stress under defined laboratory conditions. The complete cell growth data showed common continuous fitness recovery to every 2°C or 4°C stepwise temperature upshift, finally resulting in an evolved E. coli strain with an improved upper temperature limit as high as 45.9°C after 523 days of serial transfer, equivalent to 7,560 generations, in minimal medium. Two-phase fitness dynamics, a rapid growth recovery phase followed by a gradual increasing growth phase, was clearly observed at diverse temperatures throughout the entire evolutionary process. Whole-genome sequence analysis revealed the transition from positive to neutral in mutation fixation, accompanied with a considerable escalation of spontaneous substitution rate in the late fitness recovery phase. It suggested that continually increasing fitness not always resulted in the reduction of genetic diversity due to the sequential takeovers by fit mutants, but caused the accumulation of a considerable number of mutations that facilitated the neutral evolution.

Genetic basis of woven nest size in subsocial spider mites.

The variation in nest size of social spider mites of the genus Stigmaeopsis is assumed to correspond to their anti-predator strategy and to be a key aspect of their social organization and speciation. It is known that the length of the dorsal setae (sc1, 2nd propodosomal setae) correlates with the nest size. We conducted interspecies cross experiments to determine the heredity of sc1 length and nest size using two closely related species that build different sized nests, Stigmaeopsis saharai Saito et Mori and Stigmaeopsis takahashii Saito et Mori. A cross between a S. saharai female and a S. takahashii male produced several viable F1 females. We measured sc1 length and the nest size of the F1 females and then compared these values with those of their parent species. The sc1 length of F1 females and the nest size constructed by these mites were intermediate with respect to the values of the two parent species. Therefore, the length of the sc1 and nest size are heritable. This result sheds light on the importance of considering the genetic basis for the variations in social organization.

Quantitative evaluation of the biological activity of various brassinosteroids using spiral root induction in rice seeds.

Spiral roots are induced in germinated rice seeds through treatment with nanomolar brassinosteroids (BRs) but not with other plant hormones. Here, we determined the minimum effective concentration (MEC) of various BRs to induce spiral roots in germinated rice seeds. The reciprocal logarithm of MEC, pMEC, was used as the BL-like activity index, which was linearly correlated with the reciprocal logarithm of a 50% effective dose (pED50) as evaluated in the rice lamina inclination assay. Furthermore, a ligand-receptor docking simulation was performed against the BL receptor complex, Arabidopsis thaliana BRI1/SERK1, and the binding free energy (ΔGbind) was calculated for the tested BRs. The ΔGbind calculation was performed using the molecular mechanics/generalized Born surface area method on an ensemble of uncorrelated snapshots collected via molecular dynamics to predict biological activity.

Gene-regulation modules in nonalcoholic fatty liver disease revealed by single-nucleus ATAC-seq.

We investigated the progression of nonalcoholic fatty liver disease from fatty liver to steatohepatitis using single-nucleus and bulk ATAC-seq on the livers of rats fed a high-fat diet (HFD). Rats fed HFD for 4 wk developed fatty liver, and those fed HFD for 8 wk further progressed to steatohepatitis. We observed an increase in the proportion of inflammatory macrophages, consistent with the pathological progression. Utilizing machine learning, we divided global gene regulation into modules, wherein transcription factors within a module could regulate genes within the same module, reaffirming known regulatory relationships between transcription factors and biological processes. We identified core genes-central to co-expression and protein-protein interaction-for the biological processes discovered. Notably, a large part of the core genes overlapped with genes previously implicated in nonalcoholic fatty liver disease. Single-nucleus ATAC-seq, combined with data-driven statistical analysis, offers insight into in vivo global gene regulation as a combination of modules and assists in identifying core genes of relevant biological processes.