Genome-wide identification and characterization of greenbug-inducible NAC transcription factors in sorghum.

BACKGROUND: Sorghum (Sorghum bicolor) is an important cereal crop grown worldwide because of its multipurpose uses such as food, forage, and bioenergy feedstock and its wide range of adaption even in marginal environments. Greenbug can cause severe damage to sorghum plants and yield loss. Plant NAC transcription factors (TFs) have been reported to have diverse functions in plant development and plant defense but has not been studied in sorghum yet. METHODS AND RESULTS: In this study, a comprehensive analysis of the sorghum NAC (SbNAC) gene family was conducted through genome-wide analysis. A total of 112 NAC genes has been identified in the sorghum genome. These SbNAC genes are phylogenetically clustered into 15 distinct subfamilies and unevenly distribute in clusters at the telomeric ends of each chromosome. Twelve pairs of SbNAC genes are possibly involved in the segmental duplication among nine chromosomes except chromosome 10. Structure analysis showed the diverse structures with a highly variable number of exons in the SbNAC genes. Furthermore, most of the SbNAC genes showed specific temporal and spatial expression patterns according to the results of RNA-seq analysis, suggesting their diverse functions during sorghum growth and development. We have also identified nine greenbug-inducible SbNAC genes by comparing the expression profiles between two sorghum genotypes (susceptible BTx623 and resistant PI607900) in response to greenbug infestation. CONCLUSIONS: Our systematic analysis of the NAC gene expression profiles provides both a preliminary survey into their roles in plant defense against insect pests and a useful reference for in-depth characterization of the SbNAC genes and the regulatory network that contributes genetic resistance to aphids.

Spectrum of PHEX Mutations and FGF23 Profiles in a Taiwanese Cohort With X-Linked Hypophosphatemia Including 102 Patients.

BACKGROUND/AIM: X-linked hypophosphatemia (XLH), the most common form of hereditary rickets, results from loss-of-function mutations in the phosphate-regulating PHEX gene. Elevated fibroblast growth factor 23 (FGF23) contributes to hypophosphatemia in XLH. This study aimed to characterize PHEX variants and serum FGF23 profiles in Taiwanese patients with XLH. PATIENTS AND METHODS: We retrospectively reviewed the records of 102 patients clinically suspected of having hypophosphatemic rickets from 2006 to 2022. Serum intact Fibroblast growth factor-23 (iFGF23) levels were measured on clinic visit days. PHEX mutations were identified using Sanger sequencing, and negative cases were analyzed using whole-exome sequencing. RESULTS: The majority (92.1%) of patients exhibited elevated FGF23 compared with normal individuals. Among 102 patients, 44 distinct PHEX mutations were identified. Several mutations recurred in multiple unrelated Taiwanese families. We discovered a high frequency of novel PHEX mutations and identified variants associated with extreme FGF23 elevation and tumorigenesis. CONCLUSION: Our findings revealed the PHEX genotypic variants and FGF23 levels in Taiwanese patients with XLH. These results are crucial given the recent approval of burosumab, a monoclonal FGF23 antibody, for XLH therapy. This study provides key insights into the clinical management of XLH in Taiwan.

Evaluation of Infective Endocarditis in Children: A 19-Year Retrospective Study in Taiwan.

BACKGROUND: Infective endocarditis (IE) is an important cause of morbidity and mortality in pediatric patients with heart disease. Little literature has explored differences in the presentation of endocarditis in children with and without heart disease. This study aimed to compare the clinical outcomes and determine the risk of in-hospital death in the study population. METHODS: Data were retrospectively collected from 2001 to 2019 from the Chang Gung Research Database (CGRD), which is the largest collection of multi-institutional electronic medical records in Taiwan. Children aged 0-20 years with IE were enrolled. We extracted and analyzed the demographic and clinical features, complications, microbiological information, and outcomes of each patient. RESULTS: Of the 208 patients with IE, 114 had heart disease and 94 did not. Compared to those without heart disease, more streptococcal infections (19.3% vs. 2.1%, p < 0.001) and cardiac complications (29.8% vs. 6.4%, p < 0.001) were observed in patients with heart disease. Although patients with heart disease underwent valve surgery more frequently (43.9% vs. 8.5%, p < 0.001) and had longer hospital stays (28.5 vs. 12.5, p = 0.021), their mortality was lower than that of those without heart disease (3.5% vs. 10.6%, p = 0.041). Thrombocytopenia was independent risk factor for in-hospital mortality in pediatric patients with IE (OR = 6.56, 95% CI: 1.43-40.37). CONCLUSION: Among pediatric patients diagnosed with IE, microbiological and clinical features differed between those with and without heart disease. Platelet counts can be used as a risk factor for in-hospital mortality in pediatric patients with IE.

α-Pheromone Precursor Protein Foc4-PP1 Is Essential for the Full Virulence of Fusarium oxysporum f. sp. cubense Tropical Race 4.

Fusarium oxysporum f. sp. cubense (Foc), which causes Fusarium wilt of bananas, is considered one of the most destructive fungal pathogens of banana crops worldwide. During infection, Foc secretes many different proteins which promote its colonization of plant tissues. Although F. oxysporum has no sexual cycle, it has been reported to secrete an α-pheromone, which acts as a growth regulator, chemoattractant, and quorum-sensing signaling molecule; and to encode a putative protein with the hallmarks of fungal α-pheromone precursors. In this study, we identified an ortholog of the α-pheromone precursor gene, Foc4-PP1, in Foc tropical race 4 (TR4), and showed that it was necessary for the growth and virulence of Foc TR4. Foc4-PP1 deletion from the Foc TR4 genome resulted in decreased fungal growth, increased sensitivity to oxidative stress and cell-wall-damaging agents, and attenuation of pathogen virulence towards banana plantlets. Subcellular localization analysis revealed that Foc4-PP1 was concentrated in the nuclei and cytoplasm of Nicotiana benthamiana cells, where it could suppress BAX-induced programmed cell death. In conclusion, these findings suggest that Foc4-PP1 contributes to Foc TR4 virulence by promoting hyphal growth and abiotic stress resistance and inhibiting the immune defense responses of host plants.

Potential disturbance of methylphenidate of gonadal hormones or pubescent development in patients with attention-deficit/hyperactivity disorder: A twelve-month follow-up study.

Several animal or case reports have demonstrated that methylphenidate (MPH) disrupts endogenous gonadal hormones and interferes with the pubescent development of children with attention-deficit/hyperactivity disorder (ADHD). Therefore, this prospective study examined the changes in gonadal hormones and pubescent development in children with ADHD undergoing 12-month MPH treatment. We recruited 146 patients with ADHD (mean age: 8.9 years, 76.7% males) and 70 healthy controls (mean age: 9.2 years, 65.7% males). Blood samples were obtained to measure the serum levels of sex hormone-binding globulin (SHBG), follicle stimulating hormone (FSH), luteinizing hormone (LH), estradiol, progesterone, testosterone, free testosterone, and prolactin in each child. The sex maturation of ADHD patients was evaluated using the Tanner Stage. Patients with ADHD (107 received MPH treatment and 39 were under natural observation) were followed up for 12 months, and we re-examined hormone levels and Tanner Stage at the endpoint. During a 12-month follow-up for all ADHD patients, the serum levels of SHBG and progesterone significantly decreased, while LH, FSH, and free-testosterone levels significantly increased. However, the duration, drug formulations, and doses of the MPH treatment did not significantly influence gonadal hormone trends or changes of Tanner Stage. This study provides evidence about gonadal hormone trends and pubescent development in children with ADHD who receive long-term MPH treatment in natural settings. We suggest that MPH treatment at usual doses does not significantly alter gonadal function trends in ADHD patients over the course of one year.

JMJD3 acts in tandem with KLF4 to facilitate reprogramming to pluripotency.

The interplay between the Yamanaka factors (OCT4, SOX2, KLF4 and c-MYC) and transcriptional/epigenetic co-regulators in somatic cell reprogramming is incompletely understood. Here, we demonstrate that the histone H3 lysine 27 trimethylation (H3K27me3) demethylase JMJD3 plays conflicting roles in mouse reprogramming. On one side, JMJD3 induces the pro-senescence factor Ink4a and degrades the pluripotency regulator PHF20 in a reprogramming factor-independent manner. On the other side, JMJD3 is specifically recruited by KLF4 to reduce H3K27me3 at both enhancers and promoters of epithelial and pluripotency genes. JMJD3 also promotes enhancer-promoter looping through the cohesin loading factor NIPBL and ultimately transcriptional elongation. This competition of forces can be shifted towards improved reprogramming by using early passage fibroblasts or boosting JMJD3's catalytic activity with vitamin C. Our work, thus, establishes a multifaceted role for JMJD3, placing it as a key partner of KLF4 and a scaffold that assists chromatin interactions and activates gene transcription.

ß-Catenin safeguards the ground state of mousepluripotency by strengthening the robustness of the transcriptional apparatus.

Mouse embryonic stem cells cultured with MEK (mitogen-activated protein kinase kinase) and GSK3 (glycogen synthase kinase 3) inhibitors (2i) more closely resemble the inner cell mass of preimplantation blastocysts than those cultured with SL [serum/leukemia inhibitory factor (LIF)]. The transcriptional mechanisms governing this pluripotent ground state are unresolved. Release of promoter-proximal paused RNA polymerase II (Pol2) is a multistep process necessary for pluripotency and cell cycle gene transcription in SL. We show that ß-catenin, stabilized by GSK3 inhibition in medium with 2i, supplies transcriptional coregulators at pluripotency loci. This selectively strengthens pluripotency loci and renders them addicted to transcription initiation for productive gene body elongation in detriment to Pol2 pause release. By contrast, cell cycle genes are not bound by ß-catenin, and proliferation/self-renewal remains tightly controlled by Pol2 pause release under 2i conditions. Our findings explain how pluripotency is reinforced in the ground state and also provide a general model for transcriptional resilience/adaptation upon network perturbation in other contexts.

mTORC1-PGC1 axis regulates mitochondrial remodeling during reprogramming.

Metabolic reprogramming, hallmarked by enhanced glycolysis and reduced mitochondrial activity, is a key event in the early phase of somatic cell reprogramming. Although extensive work has been conducted to identify the mechanisms of mitochondrial remodeling in reprogramming, many questions remain. In this regard, different laboratories have proposed a role in this process for either canonical (ATG5-dependent) autophagy-mediated mitochondrial degradation (mitophagy), noncanonical (ULK1-dependent, ATG5-independent) mitophagy, mitochondrial fission or reduced biogenesis due to mTORC1 suppression. Clarifying these discrepancies is important for providing a comprehensive picture of metabolic changes in reprogramming. Yet, the comparison among these studies is difficult because they use different reprogramming conditions and mitophagy detection/quantification methods. Here, we have systematically explored mitochondrial remodeling in reprogramming using different culture media and reprogramming factor cocktails, together with appropriate quantification methods and thorough statistical analysis. Our experiments show lack of evidence for mitophagy in mitochondrial remodeling in reprogramming, and further confirm that the suppression of the mTORC1-PGC1 pathway drives this process. Our work helps to clarify the complex interplay between metabolic changes and nutrient sensing pathways in reprogramming, which may also shed light on other contexts such as development, aging and cancer.

NCoR/SMRT co-repressors cooperate with c-MYC to create an epigenetic barrier to somatic cell reprogramming.

Somatic cell reprogramming by exogenous factors requires cooperation with transcriptional co-activators and co-repressors to effectively remodel the epigenetic environment. How this interplay is regulated remains poorly understood. Here, we demonstrate that NCoR/SMRT co-repressors bind to pluripotency loci to create a barrier to reprogramming with the four Yamanaka factors (OCT4, SOX2, KLF4 and c-MYC), and consequently, suppressing NCoR/SMRT significantly enhances reprogramming efficiency and kinetics. The core epigenetic subunit of the NCoR/SMRT complex, histone deacetylase 3 (HDAC3), contributes to the effects of NCoR/SMRT by inducing histone deacetylation at pluripotency loci. Among the Yamanaka factors, recruitment of NCoR/SMRT-HDAC3 to genomic loci is mostly facilitated by c-MYC. Hence, we describe how c-MYC is beneficial for the early phase of reprogramming but deleterious later. Overall, we uncover a role for NCoR/SMRT co-repressors in reprogramming and propose a dual function for c-MYC in this process.

Prenatal Dexamethasone Exposure Programs the Development of the Pancreas and the Secretion of Insulin in Rats.

BACKGROUND: There is increasing epidemiological evidence indicating that many chronic diseases originate during early life, even before birth, through what are termed fetal programming effects. Prenatal glucocorticoid is frequently used clinically to accelerate the maturation of the lung, but its long-term effects remain unclear. METHODS: We gave pregnant Sprague-Dawley rats either intraperitoneal dexamethasone (0.1 mg/kg body weight) or vehicle at Gestational Days 14-20 and assessed the effects to pancreas at Postnatal Days 7 and 120. RESULTS: We found fewer pancreatic ß cell fractions (0.31±0.05 % vs. 0.49±0.05 %, p=0.013) and tissues (0.0017±0.0002 % vs. 0.0025±0.0002 %, p=0.042) and decreased secretion of insulin in response to a glucose challenge at Postnatal Day 105 (1.00±0.19 ng/mL vs. 1.57±0.17 ng/mL at the 15-minute time-point, p=0.046) in rats treated prenatally with dexamethasone. At Postnatal Day 7 in rats treated prenatally with dexamethasone, the expression of pancreatic duodenal homeobox gene-1 and V-maf avian musculoaponeurotic fibrosarcoma oncogene homolog A was lower than that in the rats in the Vehicle group (0.22±0.07 vs. 1.00±0.41 fold, p=0.01, 0.20±0.12 vs. 1.00±0.35 fold, p=0.01) while the histone deacetylases activity (54.2±3.7 ng/h/mL vs. 37.6±3.5 ng/h/mL, p=0.012) and 8-hydroxy-2-deoxyguanosine staining (1.34±0.01 vs. 1.00±0.02 fold, p<0.01) were higher. CONCLUSION: Prenatal dexamethasone exposure affects early postnatal gene expression related to pancreas development and may exert an effect on ß-cell development at 120 postnatal days.

[Effects of acupuncture stimulation of different acupoint groups on sleeping latency, serum and hippocampal TNF-α and IL-25 contents in rats with gastric mucosal injury].

OBJECTIVE: To observe the effect of acupuncture intervention on gastric ulcer (GU) and sleeping quality from the viewpoint of brain-gut axis which plays an important role in the regulation of many vital functions in the body. METHODS: Forty male Wistar rats were randomized into normal control, GU model, acupuncture of "Zhongwan"(CV 12)-"Zusanli"(ST 36, gastric function regulating acupoints), acupuncture of "Shenmai" (BL 62)-"Zhaohai" (KI 6, sleep-promotion acupoints), and acupuncture of CV 12-ST 36+ BL 62-KI 6 (combined treatment) groups, with 8 rats in each group. GU model was established by intragastric perfusion of dehydrated alcohol (1 mL/rat), and sleep model established by intraperitoneal injection of pentobarbital sodium (40 mg/kg) after the last treatment. The abovementioned acupoints were punctured with filiform needles and stimulated by manipulating the needle for about 30 s, once every 5 min during 20 min of needle retention. The treatment was conducted once daily for five days. The contents of tumor necrosis factor-alpha (TNF-α) and interleukin-25(IL-25) in the serum and hippocampal tissues were detected by ELISA. RESULTS: Compared with the normal control group, the gastric ulcer index score, barbiturate-induced sleeping time, and TNF-α and IL-25 contents in both serum and hippocampus were significantly increased in the model group (P < 0.01). Following acupuncture treatment, in comparison with the model group, the gastric ulcer index score, barbiturate-induced sleeping time, and TNF-α and IL-25 contents in both serum and hippocampus were significantly down-regulated in the CV 12-ST 36, BL 62-KI 6 and combined treatment groups (P < 0.01, P < 0.05). The effects of the CV 12-ST 36 and combined treatment groups were remarkably superior to those of the BL 62-KI 6 group in down-regulating ulcer index score, serum IL-25, and hippocampal TNF-α and IL-25 contents (P < 0.01, P < 0.05). In addition, the effects of the BL 62-KI 6 and combined treatment groups was considerably better than that of the CV 12-ST 36 group in shortening barbiturate-induced sleeping time (P < 0.01, P < 0.05). The effect of the combined treatment group was markedly better than that of the CV 12-ST 36 and BL 62-KI 6 groups in lowering serum TNF-α content (P < 0.05). CONCLUSION: Acupuncture stimulation of CV 12, ST 36, KI 6 and BL 62 can relieve the gastric mucosal lesion, and shorten barbiturate-induced sleeping time in gastric ulcer rats, which may be related to its effects in reducing TNF-α and IL-25 contents in the serum and hippocampus tissues, suggesting a correlation between the gastrointestinal disorder and sleeping.

Autophagy and mTORC1 regulate the stochastic phase of somatic cell reprogramming.

We describe robust induction of autophagy during the reprogramming of mouse fibroblasts to induced pluripotent stem cells by four reprogramming factors (Sox2, Oct4, Klf4 and c-Myc), henceforth 4F. This process occurs independently of p53 activation, and is mediated by the synergistic downregulation of mechanistic target of rapamycin complex 1 (mTORC1) and the induction of autophagy-related genes. The 4F coordinately repress mTORC1, but bifurcate in their regulation of autophagy-related genes, with Klf4 and c-Myc inducing them but Sox2 and Oct4 inhibiting them. On one hand, inhibition of mTORC1 facilitates reprogramming by promoting cell reshaping (mitochondrial remodelling and cell size reduction). On the other hand, mTORC1 paradoxically impairs reprogramming by triggering autophagy. Autophagy does not participate in cell reshaping in reprogramming but instead degrades p62, whose accumulation in autophagy-deficient cells facilitates reprogramming. Our results thus reveal a complex signalling network involving mTORC1 inhibition and autophagy induction in the early phase of reprogramming, whose delicate balance ultimately determines reprogramming efficiency.

Generation of human induced pluripotent stem cells from urine samples.

Human induced pluripotent stem cells (iPSCs) have been generated with varied efficiencies from multiple tissues. Yet, acquiring donor cells is, in most instances, an invasive procedure that requires laborious isolation. Here we present a detailed protocol for generating human iPSCs from exfoliated renal epithelial cells present in urine. This method is advantageous in many circumstances, as the isolation of urinary cells is simple (30 ml of urine are sufficient), cost-effective and universal (can be applied to any age, gender and race). Moreover, the entire procedure is reasonably quick--around 2 weeks for the urinary cell culture and 3-4 weeks for the reprogramming--and the yield of iPSC colonies is generally high--up to 4% using retroviral delivery of exogenous factors. Urinary iPSCs (UiPSCs) also show excellent differentiation potential, and thus represent a good choice for producing pluripotent cells from normal individuals or patients with genetic diseases, including those affecting the kidney.

Variation of taxane content in needles of Taxus x media cultivars with different growth characteristics.

Needles from 17 different Taxus x media cultivars, belonging to 4 groups showing different growth characteristics, were analyzed using high performance liquid chromatography for their content of 10-deacetylbaccatin III, baccatin III, cephalomannine and paclitaxel (Taxol). The 4 Taxus x media cultivar groups were: 1.) medium to fast growing and upright form; 2.) slow growing and upright form; 3.) fast growing and spreading form; and 4.) slow growing and spreading form. The purpose of this study was to identify yew cultivars of fast growth rate, upright growth and high taxane content in their needles. The highest content of paclitaxel was found in 'Coleana' of group 1 (378 microg/g of the extracted dry weight). Three cultivars in group 1, 'Coleana', 'Stovekenii' and 'Hicksii', make good candidates for taxane extraction because of their high paclitaxel and 10-deacetylbaccatin III content, fast biomass accumulation and upright growing form. They are also good starting materials to develop alternative methods for the production of paclitaxel and its analogous compounds through modern biotechnology approaches.