Impact of extended lung protection during mechanical ventilation on lung recovery in patients with COVID-19 ARDS: a phase II randomized controlled trial.

BACKGROUND: Protective ventilation seems crucial during early Acute Respiratory Distress Syndrome (ARDS), but the optimal duration of lung protection remains undefined. High driving pressures (ΔP) and excessive patient ventilatory drive may hinder lung recovery, resulting in self-inflicted lung injury. The hidden nature of the ΔP generated by patient effort complicates the situation further. Our study aimed to assess the feasibility of an extended lung protection strategy that includes a stepwise protocol to control the patient ventilatory drive, assessing its impact on lung recovery. METHODS: We conducted a single-center randomized study on patients with moderate/severe COVID-19-ARDS with low respiratory system compliance (CRS < 0.6 (mL/Kg)/cmH2O). The intervention group received a ventilation strategy guided by Electrical Impedance Tomography aimed at minimizing ΔP and patient ventilatory drive. The control group received the ARDSNet low-PEEP strategy. The primary outcome was the modified lung injury score (mLIS), a composite measure that integrated daily measurements of CRS, along with oxygen requirements, oxygenation, and X-rays up to day 28. The mLIS score was also hierarchically adjusted for survival and extubation rates. RESULTS: The study ended prematurely after three consecutive months without patient enrollment, attributed to the pandemic subsiding. The intention-to-treat analysis included 76 patients, with 37 randomized to the intervention group. The average mLIS score up to 28 days was not different between groups (P = 0.95, primary outcome). However, the intervention group showed a faster improvement in the mLIS (1.4 vs. 7.2 days to reach 63% of maximum improvement; P < 0.001), driven by oxygenation and sustained improvement of X-ray (P = 0.001). The intervention group demonstrated a sustained increase in CRS up to day 28 (P = 0.009) and also experienced a shorter time from randomization to room-air breathing (P = 0.02). Survival at 28 days and time until liberation from the ventilator were not different between groups. CONCLUSIONS: The implementation of an individualized PEEP strategy alongside extended lung protection appears viable. Promising secondary outcomes suggested a faster lung recovery, endorsing further examination of this strategy in a larger trial. Clinical trial registration This trial was registered with ClinicalTrials.gov (number NCT04497454) on August 04, 2020.

The association between subjective anti-doping knowledge and objective knowledge among Japanese university athletes: a cross-sectional study.

Introduction: This study aimed to assess the association between subjective anti-doping knowledge (subjective ADK) and objective anti-doping knowledge (objective ADK) among Japanese university athletes, framed within the context of the Theory of Planned Behavior (TPB). Methods: Eligible participants were 486 university athletes [320 men (65.8%), 166 women; mean age of 18.9 ± 1.0 years]. The participants categorized themselves in terms of the quality of their anti-doping knowledge. This assessment resulted in an independent variable coded as "(1) substantial lack of adequate knowledge," "(2) some lack of adequate knowledge," "(3) fair amount of knowledge" or "(4) good amount of knowledge." Objective ADK was assessed using the Athlete Learning Program about Health and Anti-Doping (ALPHA) test, a set of questions derived from the ALPHA-a former World Anti-Doping Agency e-learning program. The test comprises 12 questions (four choices each; passing index: ≧10 points or 80% correct answer rate). ANCOVA was conducted using subjective ADK as an independent variable and ALPHA scores as a dependent variable, adjusting for confounding factors (anti-doping experience). Results: The ALPHA corrected answer rate across subjective ADK levels for the group were 73.10% for "(1) substantial lack of adequate knowledge," 71.97% for "(2) some lack of adequate knowledge," 75.18% for "(3) fair amount of knowledge" and 72.86% for "(4) good amount of knowledge." Comparison between different levels of subjective ADK revealed no significant differences in ALPHA score considering the main effects or any of their interactions. Discussion: The present results revealed that Japanese university athletes' subjective ADK did not match their objective ADK. In the context of the TPB, there may be limitations in the perceived behavioral control in anti-doping knowledge. Even if athletes view doping as a wrongful act and have formed attitudes and subjective norms to comply with the rules, the results suggest that errors may occur in the composition of behavioral intentions due to a lack of knowledge. This could lead to the possibility of facing the risk of unintentional anti-doping rule violations. It highlights the need for targeted educational interventions to align subjective ADK of athletes with their objective ADK.

Serum Concentrations of Infliximab and IL-6 for Predicting One-Year Discontinuation of Infliximab Treatment Owing to Secondary Non-response in Patients with Rheumatoid Arthritis.

Secondary non-response to infliximab (IFX) occurs in some patients with rheumatoid arthritis (RA). Although therapeutic drug monitoring (TDM) is a useful tool to optimize IFX therapy, it is unclear whether it can help to identify the risk of secondary non-response. This study aimed to explore the utility of serum levels of IFX or other biomarkers to predict IFX discontinuation owing to secondary non-response. A single-center, retrospective study was conducted using the Kyoto University Rheumatoid Arthritis Management Alliance cohort database between 2011 and 2020. Serum IFX levels were measured using liquid chromatography-tandem mass spectrometry. An electrochemiluminescence assay was used to quantify serum levels of tumor necrosis factor-α and interleukin-6 and detect anti-drug antibodies. Eighty-four out of 310 patients were eligible for this study. The cutoff levels of biomarkers were determined by receiver operating characteristic analysis. IFX persistence was similar between groups stratified using IFX levels, tumor necrosis factor-α levels, interleukin-6 levels, and anti-drug antibodies positivity. The group with lower IFX and higher interleukin-6 levels had the worst therapy persistence (p = 0.017) and the most frequent disease worsening (90.0%, p < 0.001). Evaluating both interleukin-6 and IFX levels, not just IFX alone, enabled us to identify patients at risk of discontinuing IFX treatment. These findings support the utility of measuring IFX and interleukin-6 levels for successful maintenance therapy for RA.

Differential effects of acute exercise on emotional memory in men and women.

Exercise may change emotional memory, which is associated with the induction of mental disorders such as depression and anxiety. This effect of exercise may be influenced by exercise-induced cortisol release. Depending on sex, cortisol exerts differential effects on emotional memory consolidation. However, whether acute exercise and exercise-induced cortisol release have sex-dependent effects on emotional memory has not been established. Therefore, first, we aimed to determine the effects of acute exercise on emotional memory, separately for men and women, in a within-subjects design. Second, we aimed to examine whether the effects of acute exercise on emotional memory are related to the effects of exercise-induced cortisol release, separately for men and women. Sixteen healthy men and 15 healthy women were presented with positive and negative emotional images, followed by either rest or a vigorous-intensity cycling exercise condition using a within-subjects design on separate days. Salivary cortisol was measured before presenting the emotional images presentation and 20 min after each intervention. Emotional memory was assessed two days later. Vigorous-intensity exercise decreased emotional memory in women, whereas there was no change in men after rest or exercise. Cortisol levels increased after exercise intervention in both men and women, although there was no association between cortisol levels and emotional memory. These findings demonstrate that the effect of a single bout of vigorous-intensity exercise on emotional memory differs between men and women and is associated with decreased emotional memory in women.

Self-organization, quality control, and preclinical studies of human iPSC-derived retinal sheets for tissue-transplantation therapy.

Three-dimensional retinal organoids (3D-retinas) are a promising graft source for transplantation therapy. We previously developed self-organizing culture for 3D-retina generation from human pluripotent stem cells (hPSCs). Here we present a quality control method and preclinical studies for tissue-sheet transplantation. Self-organizing hPSCs differentiated into both retinal and off-target tissues. Gene expression analyses identified the major off-target tissues as eye-related, cortex-like, and spinal cord-like tissues. For quality control, we developed a qPCR-based test in which each hPSC-derived neuroepithelium was dissected into two tissue-sheets: inner-central sheet for transplantation and outer-peripheral sheet for qPCR to ensure retinal tissue selection. During qPCR, tissue-sheets were stored for 3-4 days using a newly developed preservation method. In a rat tumorigenicity study, no transplant-related adverse events were observed. In retinal degeneration model rats, retinal transplants differentiated into mature photoreceptors and exhibited light responses in electrophysiology assays. These results demonstrate our rationale toward self-organizing retinal sheet transplantation therapy.

Relationship between the level of willingness to learn about anti-doping and objective knowledge among Japanese university athletes: A cross-sectional study.

Previous studies have indicated that athletes' anti-doping knowledge is inadequate. Athletes' willingness to learn about anti-doping (willingness to learn) may influence their anti-doping knowledge, but the actual situation is unclear. This study aimed to determine the relationship between athletes' willingness to learn about anti-doping and their objective measurement knowledge and explore directions for educational interventions. The eligible participants were 971 male and 802 female university athletes. We used the ALPHA test (12 questions/four choices; passing index: ≥10 points/80% correct answer rate) to assess objective anti-doping knowledge. The willingness to learn question was, "Would you like to learn more about anti-doping?" Responses were given on a 4-point scale ranging from 1: strongly disagree to 4: strongly agree. An ANCOVA was conducted with four levels of willingness to learn as the independent variable and ALPHA correct answer rate as the dependent variable, adjusting for confounding factors (years of athletic experience and anti-doping education experience). The percentage of athletes (%) and each ALPHA correct answer rate (%) by the level of willingness to learn was 1: strongly disagree, n = 1.64%, 61.78%; 2: somewhat disagree, n = 13.14%, 62.38%; 3: somewhat agree, n = 62.94%, 64.08%; 4: strongly agree, n = 22.28%, 67.11%. The ALPHA correct answer rates showed significant differences in the main effect by the level of willingness to learn [F (3, 1767) = 2.873, p < 0.05, η2 = 0.01], although the effect size was small, and multiple comparisons showed no significant differences between the levels. The results indicated that the ALPHA correct answer rate did not reach 80% even for the "strongly agree" level of willingness to learn, suggesting that information on anti-doping may be inadequate. The need to provide sufficient educational content to improve knowledge was evident.

Latent changes in the pharyngeal stage of swallowing in non-aspirating older adults.

PURPOSE: The characteristic changes in the swallowing mechanism with aging are collectively termed presbyphagia. Although several studies have investigated presbyphagia in older adults, few have assessed oldest-old adults. We aimed to characterize the latent changes of swallowing function in oldest-old adults and to consider risk ages for presbyphagia. METHODS: We analyzed the records of 85 individuals (44 males and 41 females, aged 25-101 years) who underwent videofluoroscopic swallowing studies. The included participants had penetration and aspiration scores of ≤ 2 and no history of aspiration, pneumonia, or diseases that affect swallowing. They were divided into four age groups: 25-64 years (non-older), 65-74 years (young-old), 75-84 years (middle-old), and ≥ 85 years (oldest-old). We analyzed and compared the pharyngeal delay time (PDT), duration of tongue base and posterior pharyngeal wall contact, duration and dimension of upper esophageal sphincter opening (UES-O), and maximal hyoid bone displacement between the age groups. RESULTS: Among the older groups, the oldest-old showed significantly longer PDT than younger-old adults, and the UES-O tended to be wider in the former. However, no other remarkable differences were found between the oldest-old and other old groups. Statistical comparisons between the < 75 and ≥ 75-year age groups revealed significant age-related changes in the PDT and duration and dimension of UES-O. CONCLUSION: On videofluoroscopic evaluation, physiological changes with aging affected few parameters of swallowing in our cohort. These findings indicate that in non-aspirating oldest-old adults, any deterioration may be adjusted for by compensatory changes to maintain swallowing function.

Potential application of measuring serum infliximab levels in rheumatoid arthritis management: A retrospective study based on KURAMA cohort data.

Infliximab (IFX) therapy has considerably improved the treatment of rheumatoid arthritis (RA). However, some patients still do not respond adequately to IFX therapy, or the efficacy of the treatment diminishes over time. Although previous studies have reported a relationship between serum IFX levels and therapeutic efficacy, the potential applications of IFX therapeutic drug monitoring (TDM) in clinical practice remain unclear. The purpose of this study was to investigate the potential applications of IFX TDM by analyzing a Japanese cohort database. Data were collected retrospectively from the Kyoto University Rheumatoid Arthritis Management Alliance cohort between January 1, 2011, and December 31, 2018. Serum IFX levels were measured using a liquid chromatography-tandem mass spectrometer. Out of the 311 RA patients that used IFX, 41 were eligible for the analysis. Serum IFX levels were significantly higher in responders than in non-responders. An optimal cut-off value was determined to be 0.32 µg/mL based on a receiver operating characteristic curve. At the IFX measurement point, a better therapeutic response was observed in the high IFX group (n = 32) than in the low IFX group (n = 9). Conversely, at the maximum effect point, when DAS28-ESR was the lowest between IFX introduction and measurement points, there were no differences in responder proportions between the low and high IFX groups. IFX primary ineffectiveness could be avoided with appropriate dose escalation without blood concentration measurement in clinical practice. In conclusion, IFX TDM could facilitate the identification of secondary non-responders and in turn, proper IFX use.

Polycomb Repressive Complex 2-mediated histone modification H3K27me3 is associated with embryogenic potential in Norway spruce.

Epigenetic reprogramming during germ cell formation is essential to gain pluripotency and thus embryogenic potential. The histone modification H3K27me3, which is catalysed by the Polycomb repressive complex 2 (PRC2), regulates important developmental processes in both plants and animals, and defects in PRC2 components cause pleiotropic developmental abnormalities. Nevertheless, the role of H3K27me3 in determining embryogenic potential in gymnosperms is still elusive. To address this, we generated H3K27me3 profiles of Norway spruce (Picea abies) embryonic callus and non-embryogenic callus using CUT&RUN, which is a powerful method for chromatin profiling. Here, we show that H3K27me3 mainly accumulated in genic regions in the Norway spruce genome, similarly to what is observed in other plant species. Interestingly, H3K27me3 levels in embryonic callus were much lower than those in the other examined tissues, but markedly increased upon embryo induction. These results show that H3K27me3 levels are associated with the embryogenic potential of a given tissue, and that the early phase of somatic embryogenesis is accompanied by changes in H3K27me3 levels. Thus, our study provides novel insights into the role of this epigenetic mark in spruce embryogenesis and reinforces the importance of PRC2 as a key regulator of cell fate determination across different plant species.

Tissue-specific transposon-associated small RNAs in the gymnosperm tree, Norway spruce.

BACKGROUND: Small RNAs (sRNAs) are regulatory molecules impacting on gene expression and transposon activity. MicroRNAs (miRNAs) are responsible for tissue-specific and environmentally-induced gene repression. Short interfering RNAs (siRNA) are constitutively involved in transposon silencing across different type of tissues. The male gametophyte in angiosperms has a unique set of sRNAs compared to vegetative tissues, including phased siRNAs from intergenic or genic regions, or epigenetically activated siRNAs. This is contrasted by a lack of knowledge about the sRNA profile of the male gametophyte of gymnosperms. RESULTS: Here, we isolated mature pollen from male cones of Norway spruce and investigated its sRNA profiles. While 21-nt sRNAs is the major size class of sRNAs in needles, in pollen 21-nt and 24-nt sRNAs are the most abundant size classes. Although the 24-nt sRNAs were exclusively derived from TEs in pollen, both 21-nt and 24-nt sRNAs were associated with TEs. We also investigated sRNAs from somatic embryonic callus, which has been reported to contain 24-nt sRNAs. Our data show that the 24-nt sRNA profiles are tissue-specific and differ between pollen and cell culture. CONCLUSION: Our data reveal that gymnosperm pollen, like angiosperm pollen, has a unique sRNA profile, differing from vegetative leaf tissue. Thus, our results reveal that angiosperm and gymnosperm pollen produce new size classes not present in vegetative tissues; while in angiosperm pollen 21-nt sRNAs are generated, in the gymnosperm Norway spruce 24-nt sRNAs are generated. The tissue-specific production of distinct TE-derived sRNAs in angiosperms and gymnosperms provides insights into the diversification process of sRNAs in TE silencing pathways between the two groups of seed plants.

Association of Decreased Handgrip Strength with Reduced Cortical Thickness in Japanese Female Patients with Type 2 Diabetes Mellitus.

LD-100, a quantitative ultrasonic device, allows us to measure cortical thickness (CoTh). Patients with type 2 diabetes mellitus (T2DM) show high prevalence of sarcopenia. This study aimed to clarify the association of handgrip strength (HGS) with cortical porosis, a major risk for fracture of DM. CoTh and trabecular bone mineral density (TrBMD) at the 5.5% distal radius were assessed in T2DM female patients (n = 122) and non-DM female controls (n = 704) by LD-100. T2DM patients aged older 40 years showed significantly lower HGS and CoTh, but not TrBMD, than non-DM counterparts. Although HGS was significantly and positively correlated with CoTh and TrBMD in T2DM patients, multivariate analysis revealed HGS as an independent factor positively associated with CoTh, but not TrBMD, in T2DM patients, suggesting the preferential association of HGS with cortical, but not trabecular, bone component in T2DM female patients. In conclusion, the present study demonstrated an early decline of HGS in T2DM female patients as compared with non-DM healthy controls after the age of 40 years, which is independently associated with thinner CoTh, but not TrBMD in T2DM patients, and thus suggested that reduced muscle strength associated with DM might be a major factor for cortical porosis development in DM patients.

FACT complex is required for DNA demethylation at heterochromatin during reproduction in Arabidopsis.

The DEMETER (DME) DNA glycosylase catalyzes genome-wide DNA demethylation and is required for endosperm genomic imprinting and embryo viability. Targets of DME-mediated DNA demethylation reside in small, euchromatic, AT-rich transposons and at the boundaries of large transposons, but how DME interacts with these diverse chromatin states is unknown. The STRUCTURE SPECIFIC RECOGNITION PROTEIN 1 (SSRP1) subunit of the chromatin remodeler FACT (facilitates chromatin transactions), was previously shown to be involved in the DME-dependent regulation of genomic imprinting in Arabidopsis endosperm. Therefore, to investigate the interaction between DME and chromatin, we focused on the activity of the two FACT subunits, SSRP1 and SUPPRESSOR of TY16 (SPT16), during reproduction in Arabidopsis We found that FACT colocalizes with nuclear DME in vivo, and that DME has two classes of target sites, the first being euchromatic and accessible to DME, but the second, representing over half of DME targets, requiring the action of FACT for DME-mediated DNA demethylation genome-wide. Our results show that the FACT-dependent DME targets are GC-rich heterochromatin domains with high nucleosome occupancy enriched with H3K9me2 and H3K27me1. Further, we demonstrate that heterochromatin-associated linker histone H1 specifically mediates the requirement for FACT at a subset of DME-target loci. Overall, our results demonstrate that FACT is required for DME targeting by facilitating its access to heterochromatin.

Correlation analysis of monocyte subsets and insulin resistance considering fetuin-A involvement in patients with type 2 diabetes.

BACKGROUND: Fetuin-A is a multifunctional circulating glycoprotein that can induce insulin resistance. Lately, adipose tissue has gained prominence as an effector site of fetuin-A. Although fetuin-A-induced proinflammatory polarization and migration of macrophages plays a crucial role, it remains obscure whether monocyte subsets in circulation could simulate characteristics of macrophages in adipose tissues. This study aims to investigate the correlation between monocyte subsets with fetuin-A and its relevant insulin resistance. RESULTS: We evaluated serum fetuin-A levels in 107 patients with type 2 diabetes (T2D). Using flow cytometry, we classified monocyte subsets into three subtypes: (a) classical, CD14++CD16-; (b) intermediate, CD14++CD16+, the most proinflammatory one; (c) and nonclassical, CD14+CD16++. We assessed the insulin resistance by the homeostasis model assessment for insulin resistance (HOMA-IR) in 68 patients without insulin injections. We observed no correlation between fetuin-A levels and classical (ρ = - 0.005; P = 0.959), intermediate (ρ = 0.022; P = 0.826), and nonclassical monocyte counts (ρ = 0.063; P = 0.516), respectively. In addition, no significant correlation was found between log (HOMA-IR) and classical (ρ = 0.052; P = 0.688), intermediate (ρ = 0.054; P = 0.676), and nonclassical monocyte counts (ρ = 0.012; P = 0.353), respectively. However, serum fetuin-A levels showed positive correlation with log (HOMA-IR) (ρ = 0.340; P = 0.007). Multiple regression analyses revealed a significant relationship between fetuin-A and log (HOMA-IR) (ß = 0.313; P = 0.016), but not with monocyte subsets. CONCLUSIONS: Monocyte subsets in circulation, including proinflammatory intermediate monocytes, were not associated with fetuin-A and insulin resistance.

Inheritance of vernalization memory at FLOWERING LOCUS C during plant regeneration.

Specific gene states can be transmitted to subsequent cell generations through mitosis involving particular chromatin (epigenetic) states. During reproduction of plants and animals, however, most epigenetic states are reset to allow development to start anew. Flowering is one of the critical developmental steps by which plants acquire their reproductive capacity. This phase transition is controlled by environmental signals and autonomous regulation. The FLOWERING LOCUS C (FLC) gene is a flowering repressor that is epigenetically silenced after long-term exposure to cold, ensuring flowering in the spring season. In Arabidopsis thaliana, epigenetically silenced FLC expression is reset during sexual reproduction. Plants have a remarkable potential to regenerate from somatic cells. However, little is known about whether the regeneration process is similar to sexual reproduction in terms of affecting chromatin states. Here, we tested whether FLC silencing is reset during in vitro regeneration. Transcriptional repression and high H3K27me3 at FLC were both stably transmitted, resulting in early flowering in regenerated shoots. Thus, the silenced epigenetic state of FLC is reset only during sexual reproduction and not during in vitro regeneration. In contrast, the active epigenetic state of FLC was only partially maintained through in vitro reproduction, suggesting that regeneration causes stochastic FLC silencing.

H3K23me1 is an evolutionarily conserved histone modification associated with CG DNA methylation in Arabidopsis.

Amino-terminal tails of histones are targets for diverse post-translational modifications whose combinatorial action may constitute a code that will be read and interpreted by cellular proteins to define particular transcriptional states. Here, we describe monomethylation of histone H3 lysine 23 (H3K23me1) as a histone modification not previously described in plants. H3K23me1 is an evolutionarily conserved mark in diverse species of flowering plants. Chromatin immunoprecipitation followed by high-throughput sequencing in Arabidopsis thaliana showed that H3K23me1 was highly enriched in pericentromeric regions and depleted from chromosome arms. In transposable elements it co-localized with CG, CHG and CHH DNA methylation as well as with the heterochromatic histone mark H3K9me2. Transposable elements are often rich in H3K23me1 but different families vary in their enrichment: LTR-Gypsy elements are most enriched and RC/Helitron elements are least enriched. The histone methyltransferase KRYPTONITE and normal DNA methylation were required for normal levels of H3K23me1 on transposable elements. Immunostaining experiments confirmed the pericentromeric localization and also showed mild enrichment in less condensed regions. Accordingly, gene bodies of protein-coding genes had intermediate H3K23me1 levels, which coexisted with CG DNA methylation. Enrichment of H3K23me1 along gene bodies did not correlate with transcription levels. Together, this work establishes H3K23me1 as a so far undescribed component of the plant histone code.

The Association between Monocyte Surface CD163 and Insulin Resistance in Patients with Type 2 Diabetes.

AIM: To investigate the association between monocyte CD163 and insulin resistance in patients with type 2 diabetes. METHODS: One hundred sixty-six patients with type 2 diabetes without inflammatory or chronic kidney disease were recruited. The monocyte CD163 levels were measured by flow cytometry and soluble CD163 (sCD163) by ELISA. Insulin resistance was evaluated by the index of the homeostasis model assessment (HOMA-R). RESULTS: The median sCD163 and monocyte CD163 expression levels were 582.9 (472.4-720.0) ng/ml and 6061 (4486-7876) mean fluorescent intensity (MFI), respectively. In a simple regression analysis, monocyte CD163 was inversely correlated with log [HOMA-R] (r = -0.257, p = 0.010), and sCD163 was positively correlated with log [HOMA-R] (r = 0.198, p = 0.042). In multiple regression analyses, monocyte CD163 was an independent contributor to log [HOMA-R] (ß = -0.220, p = 0.020) even after adjustment of various clinical factors for HOMA-R (R2 = 0.281, p = 0.001), whereas sCD163 was not. CONCLUSIONS: Monocyte surface CD163 expression levels were more significantly associated with insulin resistance than sCD163 in patients with type 2 diabetes, suggesting a novel pathophysiological role of CD163.

The H3 chaperone function of NASP is conserved in Arabidopsis.

Histones are abundant cellular proteins but, if not incorporated into chromatin, they are usually bound by histone chaperones. Here, we identify Arabidopsis NASP as a chaperone for histones H3.1 and H3.3. NASP interacts in vitro with monomeric H3.1 and H3.3 as well as with histone H3.1-H4 and H3.3-H4 dimers. However, NASP does not bind to monomeric H4. NASP shifts the equilibrium between histone dimers and tetramers towards tetramers but does not interact with tetramers in vitro. Arabidopsis NASP promotes [H3-H4]2 tetrasome formation, possibly by providing preassembled histone tetramers. However, NASP does not promote disassembly of in vitro preassembled tetrasomes. In contrast to its mammalian homolog, Arabidopsis NASP is a predominantly nuclear protein. In vivo, NASP binds mainly monomeric H3.1 and H3.3. Pulldown experiments indicated that NASP may also interact with the histone chaperone MSI1 and a HSC70 heat shock protein.

H3K36ac Is an Evolutionary Conserved Plant Histone Modification That Marks Active Genes.

In eukaryotic cells, histones are subject to a large number of posttranslational modifications whose sequential or combinatorial action affects chromatin structure and genome function. We identified acetylation at Lys-36 in histone H3 (H3K36ac) as a new chromatin modification in plants. The H3K36ac modification is evolutionary conserved in seed plants, including the gymnosperm Norway spruce (Picea abies) and the angiosperms rice (Oryza sativa), tobacco (Nicotiana tabacum), and Arabidopsis (Arabidopsis thaliana). In Arabidopsis, H3K36ac is highly enriched in euchromatin but not in heterochromatin. Genome-wide chromatin immunoprecipitation sequencing experiments revealed that H3K36ac peaks at the 5' end of genes, mainly on the two nucleosomes immediately distal to the transcription start site, independently of gene length. H3K36ac overlaps with H3K4me3 and the H2A.Z histone variant. The histone acetyl transferase GCN5 and the histone deacetylase HDA19 are required for H3K36ac homeostasis. H3K36ac and H3K36me3 show negative crosstalk, which is mediated by GCN5 and the histone methyl transferase SDG8. Although H3K36ac is associated with gene activity, we did not find a linear relationship between H3K36ac and transcript levels, suggesting that H3K36ac is a binary indicator of transcription.

Epigenetic role for the conserved Fe-S cluster biogenesis protein AtDRE2 in Arabidopsis thaliana.

On fertilization in Arabidopsis thaliana, one maternal gamete, the central cell, forms a placenta-like tissue, the endosperm. The DNA glycosylase DEMETER (DME) excises 5-methylcytosine via the base excision repair pathway in the central cell before fertilization, creating patterns of asymmetric DNA methylation and maternal gene expression across DNA replications in the endosperm lineage (EDL). Active DNA demethylation in the central cell is essential for transcriptional activity in the EDL of a set of genes, including FLOWERING WAGENINGEN (FWA). A DME-binding motif for iron-sulfur (Fe-S) cluster cofactors is indispensable for its catalytic activity. We used an FWA-GFP reporter to find mutants defective in maternal activation of FWA-GFP in the EDL, and isolated an allele of the yeast Dre2/human antiapoptotic factor CIAPIN1 homolog, encoding an enzyme previously implicated in the cytosolic Fe-S biogenesis pathway (CIA), which we named atdre2-2. We found that AtDRE2 acts in the central cell to regulate genes maternally activated in the EDL by DME. Furthermore, the FWA-GFP expression defect in atdre2-2 was partially suppressed genetically by a mutation in the maintenance DNA methyltransferase MET1; the DNA methylation levels at four DME targets increased in atdre2-2 seeds relative to WT. Although atdre2-2 shares zygotic seed defects with CIA mutants, it also uniquely manifests dme phenotypic hallmarks. These results demonstrate a previously unidentified epigenetic function of AtDRE2 that may be separate from the CIA pathway.