Single-molecule real-time transcript sequencing identified flowering regulatory genes in Crocus sativus.

BACKGROUND: Saffron crocus (Crocus sativus) is a valuable spice with medicinal uses in gynaecopathia and nervous system diseases. Identify flowering regulatory genes plays a vital role in increasing flower numbers, thereby resulting in high saffron yield. RESULTS: Two full length transcriptome gene sets of flowering and non-flowering saffron crocus were established separately using the single-molecule real-time (SMRT) sequencing method. A total of sixteen SMRT cells generated 22.85 GB data and 75,351 full-length saffron crocus unigenes on the PacBio RS II panel and further obtained 79,028 SSRs, 72,603 lncRNAs and 25,400 alternative splicing (AS) events. Using an Illumina RNA-seq platform, an additional fifteen corms with different flower numbers were sequenced. Many differential expression unigenes (DEGs) were screened separately between flowering and matched non-flowering top buds with cold treatment (1677), flowering top buds of 20 g corms and non-flowering top buds of 6 g corms (1086), and flowering and matched non-flowering lateral buds (267). A total of 62 putative flower-related genes that played important roles in vernalization (VRNs), gibberellins (G3OX, G2OX), photoperiod (PHYB, TEM1, PIF4), autonomous (FCA) and age (SPLs) pathways were identified and a schematic representation of the flowering gene regulatory network in saffron crocus was reported for the first time. After validation by real-time qPCR in 30 samples, two novel genes, PB.20221.2 (p = 0.004, r = 0.52) and PB.38952.1 (p = 0.023, r = 0.41), showed significantly higher expression levels in flowering plants. Tissue distribution showed specifically high expression in flower organs and time course expression analysis suggested that the transcripts increasingly accumulated during the flower development period. CONCLUSIONS: Full-length transcriptomes of flowering and non-flowering saffron crocus were obtained using a combined NGS short-read and SMRT long-read sequencing approach. This report is the first to describe the flowering gene regulatory network of saffron crocus and establishes a reference full-length transcriptome for future studies on saffron crocus and other Iridaceae plants.

Transcriptome analysis revealed the dynamic oil accumulation in Symplocos paniculata fruit.

BACKGROUND: Symplocos paniculata, asiatic sweetleaf or sapphire berry, is a widespread shrub or small tree from Symplocaceae with high oil content and excellent fatty acid composition in fruit. It has been used as feedstocks for biodiesel and cooking oil production in China. Little transcriptome information is available on the regulatory molecular mechanism of oil accumulation at different fruit development stages. RESULTS: The transcriptome at four different stages of fruit development (10, 80,140, and 170 days after flowering) of S. paniculata were analyzed. Approximately 28 million high quality clean reads were generated. These reads were trimmed and assembled into 182,904 non-redundant putative transcripts with a mean length of 592.91 bp and N50 length of 785 bp, respectively. Based on the functional annotation through Basic Local Alignment Search Tool (BLAST) with public protein database, the key enzymes involved in lipid metabolism were identified, and a schematic diagram of the pathway and temporal expression patterns of lipid metabolism was established. About 13,939 differentially expressed unigenes (DEGs) were screened out using differentially expressed sequencing (DESeq) method. The transcriptional regulatory patterns of the identified enzymes were highly related to the dynamic oil accumulation along with the fruit development of S. paniculata. In addition, quantitative real-time PCR (qRT-PCR) of six vital genes was significantly correlated with DESeq data. CONCLUSIONS: The transcriptome sequences obtained and deposited in NCBI would enrich the public database and provide an unprecedented resource for the discovery of the genes associated with lipid metabolism pathway in S. paniculata. Results in this study will lay the foundation for exploring transcriptional regulatory profiles, elucidating molecular regulatory mechanisms, and accelerating genetic engineering process to improve the yield and quality of seed oil of S. paniculata.

Monitoring the environmental effects of CeO2 and ZnO nanoparticles through the life cycle of corn (Zea mays) plants and in situ µ-XRF mapping of nutrients in kernels.

Information about changes in physiological and agronomic parameters through the life cycle of plants exposed to engineered nanoparticles (NPs) is scarce. In this study, corn (Zea mays) plants were cultivated to full maturity in soil amended with either nCeO2 or nZnO at 0, 400, and 800 mg/kg. Gas exchange was monitored every 10 days, and at harvest, bioaccumulation of Ce and Zn in tissues was determined by ICP-OES/MS. The effects of NPs exposure on nutrient concentration and distribution in ears were also evaluated by ICP-OES and µ-XRF. Results showed that nCeO2 at both concentrations did not impact gas exchange in leaves at any growth stage, while nZnO at 800 mg/kg reduced net photosynthesis by 12%, stomatal conductance by 15%, and relative chlorophyll content by 10% at day 20. Yield was reduced by 38% with nCeO2 and by 49% with nZnO. Importantly, µ-XRF mapping showed that nCeO2 changed the allocation of calcium in kernels, compared to controls. In nCeO2 treated plants, Cu, K, Mn, and Zn were mainly localized at the insertion of kernels into cobs, but Ca and Fe were distributed in other parts of the kernels. Results showed that nCeO2 and nZnO reduced corn yield and altered quality of corn.

Trp53 regulates Notch 4 signaling through Mdm2.

Notch receptors and their ligands have crucial roles in development and tumorigenesis. We present evidence demonstrating the existence of an antagonistic relationship between Notch 4 and Trp53, which is controlled by the Mdm2-dependent ubiquitylation and degradation of the Notch receptor. We show that this signal-controlling mechanism is mediated by physical interactions between Mdm2 and Notch 4 and suggest the existence of a trimeric complex between Trp53, Notch 4 and Mdm2, which ultimately regulates Notch activity. Functional studies indicate that Trp53 can suppress NICD4-induced anchorage-independent growth in mammary epithelial cells and present evidence showing that Trp53 has a pivotal role in the suppression of Notch-associated tumorigenesis in the mammary gland.

Functional physical training improves fitness and cognitive development in 4~5 years old children.

Background: Development of physical and cognitive function is very critical in 4~5 years children. It has been addressed in this research if the 18 weeks of specific functional training with or without cognitive training can be effective on improving fitness and cognitive development in 4~5 years preschool children. Methods: A total of 126 preschool children in the 4~5 age range were selected as participants and randomly assigned to one of four groups: the control group (C), the functional physical training group (P), the cognitive training group (CT), and the functional physical training combined with cognitive training group (PCT). Results: The results revealed significant pre/post differences in body height and weight among all four groups of children. Furthermore, there was no significant difference in physical fitness between the C group and the CT group after the intervention. However, the children in the P group and the PCT group showed significant improvements in three indicators including standing long jump, continuous jump, and 10-meter shuttle running. Additionally, the children in P group, CT group, and the PCT group demonstrated significant improvement in simple reaction time, attention, and spatial memory. No significant cognitive improvement was found in C group. Conclusion: Functional physical training with or without cognitive intervention can promote both physical fitness and cognitive development in children aged 4~5 years. Cognitive training alone can significantly improve cognitive function but not physical. Therefore, functional physical training can be used alone to improve the physical and cognitive abilities for aged 4~5 years old children.

Comparative effectiveness of school-based exercise interventions on physical fitness in children and adolescents: a systematic review and network meta-analysis.

Background: Schools provide a favorable setting for health education, however, the most effective school-based exercise mode for improving physical fitness remains unclear. This network meta-analysis was designed to assess and rank the comparative efficacy of six exercise modalities on physical fitness indicators in a school-based setting. Methods: An online search of the Web of Science, PubMed, SPORTDiscus, and Scopus databases was conducted. Randomized and quasi-randomized controlled trials were considered. Outcomes included measures of anthropometry and body composition, muscular fitness, and cardiorespiratory fitness. Data were pooled with a random effects model using the frequentist framework. Results: A total of 66 studies with 8,578 participants (48% girls) were included. High-intensity interval training was the most effective intervention reducing body mass index (mean difference (MD) = -0.60 kg·m-2, 95% confidence interval (95%CI) = -1.04 to -0.15, p = 0.009), elevating VO2max (MD = 3.59 mL·kg-1·min-1, 95% CI = 2.45 to 4.74, p < 0.001), and 20-meter sprint performance (MD = -0.35 s, 95% CI = -0.55 to -0.14, p = 0.001). Aerobic training had the highest probability of reducing waist circumference (standardized mean difference (SMD) = -0.60, 95% CI = -0.88 to -0.32, p < 0.001). Active video games emerged as a promising modality for improving countermovement jump (MD = 2.43 cm, 95% CI = 0.06 to 4.80, p = 0.041) and shuttle running performance (SMD = 0.86, 95% CI = 0.29 to 1.43, p = 0.003). Strength training was the best exercise mode for improving standing long jump performance (SMD = 1.03, 95% CI = 0.07 to 1.98, p = 0.035) while combined training was rated the first for decreasing body fat percent (MD = -2.56%, 95% CI = -4.73 to -0.40, p = 0.022) and increasing push-up repetitions (SMD = 3.59, 95% CI = 0.81 to 6.37, p = 0.012). Conclusion: School-based exercise interventions have multiple effects on physical fitness. The findings of this study will help to inform physical education teachers and coaches how best to deliver exercise programs in a school setting. Since the study was limited by the original research, the conclusions will require further verification using high-quality randomized controlled trials. Systematic Review Registration: PROSPERO, Identifier: CRD42023401963.

Effects of Water Availability on Leaf Trichome Density and Plant Growth and Development of Shepherdia ×utahensis.

Many arid lands across the globe are experiencing more frequent and extreme droughts due to warmer temperatures resulting from climate change, less predictable precipitation patterns, and decreased soil moisture. Approximately 60-90% of household water is used for urban landscape irrigation in the western United States, necessitating the establishment of landscapes using drought-tolerant plants that conserve water. Shepherdia ×utahensis (hybrid buffaloberry) is a drought-tolerant plant with dense leaf trichomes (epidermal appendages) that may limit excessive water loss by transpiration. However, little is known about how S. ×utahensis regulates leaf heat balance when transpirational cooling is limited. The objective of this research was to investigate the effects of substrate water availability on plant growth and development and trichome density of S. ×utahensis. Ninety-six clonally propagated plants were grown using an automated irrigation system, and their substrate volumetric water contents were controlled at 0.05-0.40 m3·m-3 for 2 months. Results showed that water stress impaired plant growth and increased the proportion of visibly wilted leaves. Shepherdia ×utahensis acclimates to drought by reducing cell dehydration and canopy overheating, which may be accomplished through decreased stomatal conductance, smaller leaf development, leaf curling, increased leaf thickness, and greater root-to-shoot ratio. Leaf trichome density increased when stem water potential decreased, resulting in greater leaf reflectance of visible light. Cell and leaf expansion were restricted under water stress, and negative correlations were exhibited between epidermal cell size and trichome density. According to our results, plasticity in leaves and roots aids plants in tolerating abiotic stresses associated with drought. Acclimation of S. ×utahensis to water stress was associated with increased trichome density due to plasticity in cell size. Dense trichomes on leaves reflected more lights which appeared to facilitate leaf temperature regulation.

Copper oxide (CuO) nanoparticles affect yield, nutritional quality, and auxin associated gene expression in weedy and cultivated rice (Oryza sativa L.) grains.

Weedy rice grows competitively with cultivated rice and significantly diminishes rice grain production worldwide. The different effects of Cu-based nanomaterials on the production of weedy and cultivated rice, especially the grain qualities are not known. Grains were collected from weedy and cultivated rice grown for four months in field soil amended with nanoscale CuO (nCuO), bulk CuO (bCuO), and copper sulfate (CuSO4) at 0, 75, 150, 300, and 600 mg Cu/kg soil. Cu translocation, essential element accumulation, yield, sugar, starch, protein content, and the expression of auxin associated genes in grains were determined. The grains of weedy and cultivated rice were differentially impacted by CuO-based compounds. At ≥300 mg/kg, nCuO and bCuO treated rice had no grain production. Treatment at 75 mg/kg significantly decreased grain yield as compared to control with the order: bCuO (by 88.7%) > CuSO4 (by 47.2%) ~ nCuO (by 38.3% only in cultivated rice); at the same dose, the Cu grain content was: nCuO ~ CuSO4 > bCuO > control. In weedy grains, K, Mg, Zn, and Ca contents were decreased by 75 and 150 mg/kg nCuO by up to 47.4%, 34.3%, 37.6%, and 60.0%, but no such decreases were noted in cultivated rice, and Fe content was increased by up to 88.6%, and 53.2%. In rice spikes, nCuO increased Mg, Ca, Fe, and Zn levels by up to 118.1%, 202.6%, 133.8%, and 103.9%, respectively. Nanoscale CuO at 75 and 150 mg/kg upregulated the transcription of an auxin associated gene by 5.22- and 1.38-fold, respectively, in grains of weedy and cultivated rice. The biodistribution of Cu-based compounds in harvested grain was determined by two-photon microscopy. These findings demonstrate a cultivar-specific and concentration-dependent response of rice to nCuO. A potential use of nCuO at 75 and 150 mg/kg in cultivar-dependent delivery system was suggested based on enhanced grain nutritional quality, although the yield was compromised. This knowledge, at the physiological and molecular level, provides valuable information for the future use of Cu-based nanomaterials in sustainable agriculture.

Effects of Aging Process on the Damping Performance of ZK60 Magnesium Alloys Prepared by Large Strain Rolling.

In this study, the effects of an aging treatment (T5) and a solution + aging treatment (T6) on the microstructure and damping properties of a ZK60 magnesium alloy prepared by large strain rolling (LSR) were studied by an optical microscope (OM), scanning electron microscopy (SEM), X-ray diffraction (XRD), and dynamic thermomechanical analysis (DMA). The results showed that both the T5 and T6 processes had a great impact on the microstructure and damping properties of the ZK60 magnesium alloy. With the increase in aging time, the grain size was basically unchanged, and the amount of the second phase increased, resulting in a gradual decrease in the damping performance. However, compared with the damping performance of the un-aged ZK60 magnesium alloy, the damping performance of the 4 h-aged ZK60 magnesium alloy was enhanced. At the same aging time, the increase in the aging temperature promoted the precipitation of the second phase, thereby reducing the damping performance of the ZK60 magnesium alloy. It was found that the T6-treated ZK60 magnesium alloy had a larger grain size, which led to a better damping performance; in addition, the T6-treated ZK60 magnesium alloy exhibited a damping plateau, which was determined by the distribution and amount of the second phase.

The effects of ectomycorrhizal fungi on heavy metals' transport in Pinus massoniana and bacteria community in rhizosphere soil in mine tailing area.

Pinus massoniana is one of the potential tree species of afforestation in barren mine tailing area, and ectomycorrhizal fungi contributes remarkably to its survival. However, how ectomycorrhizal fungi interacts with Pinus massoniana under heavy metals' stress is unknown. Two systems (Pinus massoniana inoculated with and without ectomycorrhizal fungi) were designed, and each system contained rhizosphere and non-rhizosphere, while bulk soil was sampled as control. The results showed that treatment of ectomycorrhizal fungi inoculation could obviously improved the absorption of soil moisture, total carbon/total nitrogen and nutrients, while reduced the bulk density and heavy metals of soil when compared with control (p＜0.05). The heavy metals accumulating in plants' roots with ectomycorrhizal fungi were greater than that without ectomycorrhizal fungi. Conversely, they were lower in shoots with ectomycorrhizal fungi. The bacterial community were affiliated with 23 bacterial phyla, 70 classes, 115 orders, 201 families, and 363 genera. Constrained Principal Coordinate Analysis and redundancy analysis demonstrated that bacterial communities structure in the soil inoculated with or without ectomycorrhizal fungi and bulk soil were distinguishable, but no difference between rhizosphere and non-rhizosphere. The LEfSe analysis showed Acidobacteria, Actinobacteria, and Proteobacteria were the dominant phyla that contributed to the difference among treatments.

Effect of the Strain Rate on the Damping and Mechanical Properties of a ZK60 Magnesium Alloy.

High strain rate rolling (HRSS) of a ZK60 magnesium alloy at 300 °C with a strain rate from 5 s-1 to 25 s-1 was used to research the effect of the rate on the mechanical properties and damping capacity of the ZK60 alloy. The results show that as the strain rate increases, the tensile strength decreases from 355 MPa at 25 s-1 to 310 MPa at 5 s-1. Two damping peaks (P1 and P2) are detected in the high strain rate rolled ZK60 alloys at different strain rates. The P1 peak appears at low temperatures and is caused by grain boundaries sliding. The P2 peak appears at high temperatures and is caused by recrystallization. As the strain rate increases from 5 to 20 s-1, the dynamic recrystallization (DRX) volume percent rises and the dislocation density decreases, both of which cause the P1 peak to become more and more obvious, and activation energy rises. At the same time, the dislocation density decreases and leads to a decrease in the storage energy, which reduces the recrystallization driving force and shifts the P2 peak to high temperatures. When the strain rate reaches 20 and 25 s-1, DRX occurs fully in the sheet, so the activation energy of the P1 peak and the temperature where the P2 peak appears are basically equal.

Improvement of nutrient elements and allicin content in green onion (Allium fistulosum) plants exposed to CuO nanoparticles.

With the exponential growth of nanomaterial production in the last years, nano copper (Cu)-based compounds are gaining more consideration in agriculture since they can work as pesticides or fertilizers. Chinese scallions (Allium fistulosum), which are characterized by their high content of the antioxidant allicin, were the chosen plants for this study. Spectroscopic and microscopic techniques were used to evaluate the nutrient element, allicin content, and enzyme antioxidant properties of scallion plants. Plants were harvested after growing for 80 days at greenhouse conditions in soil amended with CuO particles [nano (nCuO) and bulk (bCuO)] and CuSO4 at 75-600 mg/kg]. Two-photon microscopy images demonstrated the particulate Cu uptake in nCuO and bCuO treated roots. In plants exposed to 150 mg/kg of the Cu-based compounds, root Cu content was higher in plants treated with nCuO compared with bCuO, CuSO4, and control (p ≤ 0.05). At 150 mg/kg, nCuO increased root Ca (86%), root Fe (71%), bulb Ca (74%), and bulb Mg (108%) content, compared with control (p ≤ 0.05). At the same concentration, bCuO reduced root Ca (67%) and root Mg (33%), compared with control (p ≤ 0.05). At all concentrations, nCuO and CuSO4 increased leaf allicin (56-187% and 42-90%, respectively), compared with control (p ≤ 0.05). The antioxidant enzymes were differentially affected by the Cu-based treatments. Overall, the data showed that nCuO enhances nutrient and allicin contents in scallion, which suggests they might be used as a nanofertilizer for onion production.

Bok choy (Brassica rapa) grown in copper oxide nanoparticles-amended soils exhibits toxicity in a phenotype-dependent manner: Translocation, biodistribution and nutritional disturbance.

The comparative toxicity of nano/bulk cupric oxide (CuO) and ionic copper (Cu) in Rosie and Green bok choy (Brassica rapa) varieties, with higher and lower anthocyanin contents, respectively, was investigated. Both phenotypes were cultivated for 70 days in natural soil amended with nano CuO (nCuO), bulk CuO (bCuO), and Cu chloride (CuCl2) at 75, 150, 300, and 600 mg Cu/kg soil. Essential elements in tissues, agronomical parameters, chlorophyll content, and Cu distribution in leaf were determined. In both varieties, nCuO treatments significantly increased Cu uptake in roots, compared with bCuO and CuCl2 (p ≤ 0.05). At all treatment concentrations, Rosie variety had more Cu than Green. More physiological impairments such as chlorophyll and leaf biomass reduction were observed in treated-Rosie varieties, compared to Green plants. The adverse effects were higher in nCuO-treated plants than their bCuO- or ionic Cu-exposed counterparts. Different distribution patterns of the translocated Cu in leaf midrib and parenchyma depended on particle size and plant phenotype, as demonstrated by two-photon microscopy. The different effects of CuO-based compounds in Rosie and Green varieties may be related to the anthocyanin content. These findings help to understand the factors involved in nanoparticles uptake and translocation to plant edible parts.

Smad2 functions as a co-activator of canonical Wnt/beta-catenin signaling pathway independent of Smad4 through histone acetyltransferase activity of p300.

Both canonical Wnt/beta-catenin and TGFbeta/Smad signaling pathways coordinately regulate pattern formation during embryogenesis as well as tumor progression. Evidence of cross-talk between these two pathways has been reported. Here we demonstrated that the Activin-like kinase 4 (Alk4)/Smad2 pathway facilitates the transcriptional activity of the oncogenic Wnt/beta-catenin/Tcf4 pathway through a novel Smad4-independent mechanism. Upon activation, Smad2 physically interacted with Tcf4, beta-catenin and the co-activator p300 to enhance transcriptional activity of beta-catenin/Tcf4 through the histone acetyltransferase activity of p300. Transactivation by Smad2 was independent of a Smad-binding element (SBE) and Smad4. Indeed, the enhancement of beta-catenin/Tcf4 transcriptional activity by activated Smad2 was negatively regulated by the presence of Smad4. Moreover, a tumor-derived missense mutant of Smad2, lacking the ability to bind to Smad4 was still able to enhance the Tcf4 transcriptional reporter in the presence of beta-catenin and Tcf4. Our findings suggest that Smad2 may function as an activator of canonical Wnt/beta-catenin/Tcf4 signaling through a SBE/Smad4-independent pathway.

Cripto-1 activates nodal- and ALK4-dependent and -independent signaling pathways in mammary epithelial Cells.

Cripto-1 (CR-1), an epidermal growth factor-CFC (EGF-CFC) family member, has a demonstrated role in embryogenesis and mammary gland development and is overexpressed in several human tumors. Recently, EGF-CFC proteins were implicated as essential signaling cofactors for Nodal, a transforming growth factor beta family member whose expression has previously been defined as embryo specific. To identify a receptor for CR-1, a human brain cDNA phage display library was screened using CR-1 protein as bait. Phage inserts with identity to ALK4, a type I serine/threonine kinase receptor for Activin, were identified. CR-1 binds to cell surface ALK4 expressed on mammalian epithelial cells in fluorescence-activated cell sorter analysis, as well as by coimmunoprecipitation. Nodal is coexpressed with mouse Cr-1 in the mammary gland, and CR-1 can phosphorylate the transcription factor Smad-2 in EpH-4 mammary epithelial cells only in the presence of Nodal and ALK4. In contrast, CR-1 stimulation of mitogen-activated protein kinase and AKT in these cells is independent of Nodal and ALK4, suggesting that CR-1 may modulate different signaling pathways to mediate its different functional roles.

Notch4 intracellular domain binding to Smad3 and inhibition of the TGF-beta signaling.

We present evidence that Notch4ICD attenuates TGF-beta signaling. Cells expressing the activated form of the Notch4 receptor (ICD4) were resistant to the growth-inhibitory effects of TGF-beta. Notch4ICD was found to bind to Smad2, Smad3 and Smad4 but with higher affinity to Smad3. Deletion analysis showed that binding of Smad3 to ICD4 was mediated by its MH2 domain and was not dependent on the presence of the RAM23 region in ICD4. Using two TGF-beta/Activin reporter luciferase assays, RT-PCR and Western blot analysis, we demonstrate that ICD4 and ICD4 deltaRAM23 inhibit Smad-binding element and 3TP luciferase reporter activity and PAI-1 gene expression. MCF-7 human breast cancer cells express Notch4ICD (ICD4) and are resistant to the growth-inhibitory effects of TGF-beta. Blockage of Notch4 processing to ICD4 by gamma-secretase inhibitor renders MCF-7 cells sensitive to growth inhibition by TGF-beta. The interplay between these two signaling pathways may be a significant determinant during mammary tumorigenesis.

Human Cripto-1 overexpression in the mouse mammary gland results in the development of hyperplasia and adenocarcinoma.

Human Cripto-1 (CR-1) is overexpressed in approximately 80% of human breast, colon and lung carcinomas. Mouse Cr-1 upregulation is also observed in a number of transgenic (Tg) mouse mammary tumors. To determine whether CR-1 can alter mammary gland development and/or may contribute to tumorigenesis in vivo, we have generated Tg mouse lines that express human CR-1 under the transcriptional control of the mouse mammary tumor virus (MMTV). Stable Tg MMTV/CR-1 FVB/N lines expressing different levels of CR-1 were analysed. Virgin female MMTV/CR-1 Tg mice exhibited enhanced ductal branching, dilated ducts, intraductal hyperplasia, hyperplastic alveolar nodules and condensation of the mammary stroma. Virgin aged MMTV/CR-1 Tg mice also possessed persistent end buds. In aged multiparous MMTV/CR-1 mice, the hyperplastic phenotype was most pronounced with multifocal hyperplasias. In the highest CR-1-expressing subline, G4, 38% (12/31) of the multiparous animals aged 12-20 months developed hyperplasias and approximately 33% (11/31) developed papillary adenocarcinomas. The long latency period suggests that additional genetic alterations are required to facilitate mammary tumor formation in conjunction with CR-1. This is the first in vivo study that shows hyperplasia and tumor growth in CR-1-overexpressing animals.

Establishment of callus and cell suspension cultures of Corydalis saxicola Bunting, a rare medicinal plant.

An efficient procedure has been developed for callus induction and cell suspension cultures of C. saxicola for the first time. Explant selection was carried out among leaf, stem and root to select a suitable type of explants capable of higher callus formation. Leaf explants thus selected showed maximum response to callus induction (67.1%). Modified B5 medium supplemented with 0.5 mg l(-1) 2,4-D plus 2 mg l(-1) BA was the most favorable medium for callus formation with the highest induction rate (94.8%) and greatest fresh weight of callus (1.7 g per explant). Cell suspension cultures were established by transferring 2-8 g fresh callus to 80 ml liquid B5 medium. An inoculum size of 8 g produced the greatest biomass accumulation, dehydrocavidine and berberine productions, which was 13.1 g l(-1), 8.0 mg l(-1) and 4.1 mg l(-1), respectively. In response to various sucrose concentrations from 10 g l(-1) to 80 g l(-1), cultures with 60 g sucrose l(-1) not only produced the highest dry biomass (18.5 g l(-1)) but also the highest formation of dehydrocavidine (11.6 mg l(-1)) and berberine (7.6 mg l(-1)). These prepared cell suspension cultures provided a useful material for further regulation of alkaloid biosynthesis and for enhanced production of valuable alkaloids on a large scale.

A Nodal- and ALK4-independent signaling pathway activated by Cripto-1 through Glypican-1 and c-Src.

Human Cripto-1 (CR-1) is a member of the epidermal growth factor-Cripto FRL1 Cryptic family that has been shown to function as a coreceptor with the type I Activin serine-threonine kinase receptor ALK4 for the transforming growth factor beta-related peptide Nodal. However, CR-1 can also activate the mitogen-activated protein kinase and Akt pathways independently of Nodal and ALK4 by an unknown mechanism. Here, we demonstrate that CR-1 specifically binds to Glypican-1, a membrane-associated heparan sulfate proteoglycan, and activates the tyrosine kinase c-Src, triggering the mitogen-activated protein kinase and Akt signaling pathways. Finally, an active Src kinase is necessary for CR-1 to induce in vitro transformation and migration in mouse mammary epithelial cells.

Advantages of Binomial Likelihood Maximization for Analyzing and Modeling Cell Survival Curves.

Mathematical analysis of cell survival allows parameter estimation for radiobiological models and selection of an appropriate model. To our knowledge, no rigorous comparisons on the accuracy of various methods used for such analysis have been performed. In this study we compared three methods: 1. maximization of binomial log-likelihood (BLL); 2. minimization of sum of squares (SS); and 3. method 2 using log-transformed data (SSlog). Analysis of Monte Carlo simulated data (A) generated from the linear-quadratic (LQ) model showed that model parameter estimates from the BLL method were more accurate and less affected by "noise" than those from other methods. Analysis of actual breast cancer cell data showed substantial differences among LQ parameters estimated by the three methods (B). To select among radiobiological models, we used: 1. Sample size-corrected Akaike information criterion (AICc), calculated from BLL method-generated log-likelihood values; and 2. Adjusted coefficient of determination (R(2)), calculated from SS/SSlog method-generated SS values. Analysis of data simulated from the repair-misrepair (RMR) formalism (C) showed that the first approach outperformed the second approach at identifying the true data-generating model. Examples of how the first approach discriminates between several models were explored using actual mouse (H2AX-proficient and -deficient) and human [DNA-dependent protein kinase (DNA-PK)-proficient and -deficient] cell data (D). Based on this work, we concluded that BLL maximization combined with AICc-based model selection constitutes an effective method for analyzing cell survival data.