Nitric oxide mitigates the phytotoxicity of imidazolium-based ionic liquids in Arabidopsis.

Ionic liquids (ILs) have many beneficial properties that are extensively used in various fields. Despite their utility, the phytotoxic aspects of ILs are poorly known. This is especially true at the transcriptomic level and the role of nitric oxide (NO) in this process. Herein, we studied the mechanism by which endogenous NO reduces the toxicity of ILs in Arabidopsis. We examined the effects of two imidazolium-based ILs (IILs) on three Arabidopsis lines, each characterized by distinct endogenous NO levels, using a combination of physiological and transcriptomics methods. IILs impaired seed germination, seedling development, chlorophyll content, and redox homeostasis in Arabidopsis. Notably, 1,3-dibutyl imidazole bromide had greater toxicity than 1-butyl-3-methylimidazolium chloride. Nox1, a mutant with an elevated NO level, had enhanced resistance, while nia1nia2, a mutant with a diminished NO level, had increased susceptibility compared to the wild type. RNA sequencing results suggested that NO mitigates IILs-induced phytotoxicity by modulating the metabolism of chlorophyll and secondary metabolites, and by bolstering the antioxidant defense system. These findings illustrate the complex molecular networks that respond to IIL stress and reveal the potential of endogenous NO as a mitigating factor in plant stress physiology.

Comparative Field Evaluation and Transcriptome Analysis Reveals that Chromosome Doubling Enhances Sheath Blight Resistance in Rice.

Rice sheath blight, caused by Rhizoctonia solani Kihn (R. solani), poses a significant threat to rice production and quality. Autotetraploid rice, developed through chromosome doubling of diploid rice, holds great potential for enhancing biological and yield traits. However, its resistance to sheath blight in the field has remained unclear. In this study, the field resistance of 35 autotetraploid genotypes and corresponding diploids was evaluated across three environments from 2020 to 2021. The booting stage was optimal for inoculating period based on the inoculation and analysis of R. solani at five rice growth stages. We found autotetraploids generally exhibited lower disease scores than diploids, indicating enhanced resistance after chromosome doubling. Among the 35 genotypes, 16 (45.71%) displayed increased resistance, 2 (5.71%) showed decreased resistance, and 17 (48.57%) displayed unstable resistance in different sowing dates. All combinations of the genotype, environment and ploidy, including the genotype-environment-ploidy interaction, contributed significantly to field resistance. Chromosome doubling increased sheath blight resistance in most genotypes, but was also dependent on the genotype-environment interaction. To elucidate the enhanced resistance mechanism, RNA-seq revealed autotetraploid recruited more down-regulated differentially expressed genes (DEGs), additionally, more resistance-related DEGs, were down-regulated at 24 h post inoculation in autotetraploid versus diploid. The ubiquinone/terpenoid quinone and diterpenoid biosynthesis pathways may play key roles in ploidy-specific resistance mechanisms. In summary, our findings shed light on the understanding of sheath blight resistance mechanisms in autotetraploid rice.

Comparative Cytological and Gene Expression Analysis Reveals That a Common Wild Rice Inbred Line Showed Stronger Drought Tolerance Compared with the Cultivar Rice.

Common wild rice (Oryza rufipogon Griff.) is an important germplasm resource containing valuable genes. Our previous analysis reported a stable wild rice inbred line, Huaye3, which derives from the common wild rice of Guangdong Province. However, there was no information about its drought tolerance ability. Here, we assessed the germination characteristics and seedling growth between the Dawennuo and Huaye3 under five concentrations of PEG6000 treatment (0, 5%, 10%, 15%, and 20%). Huaye3 showed a stronger drought tolerance ability, and its seed germination rate still reached more than 52.50% compared with Dawennuo, which was only 25.83% under the 20% PEG6000 treatment. Cytological observations between the Dawennuo and Huaye3 indicated the root tip elongation zone and buds of Huaye3 were less affected by the PEG6000 treatment, resulting in a lower percentage of abnormalities of cortical cells, stele, and shrinkage of epidermal cells. Using the re-sequencing analysis, we detected 13,909 genes that existed in the genetic variation compared with Dawennuo. Of these genes, 39 were annotated as drought stress-related genes and their variance existed in the CDS region. Our study proved the strong drought stress tolerance ability of Huaye3, which provides the theoretical basis for the drought resistance germplasm selection in rice.

Cytological Observation and RNA-Seq Analyses Reveal miR9564 and Its Target Associated with Pollen Sterility in Autotetraploid Rice.

Understanding the regulation of autotetraploid sterility is essential for harnessing the strong advantages in genomic buffer capacity, biodiversity, and heterosis of autotetraploid rice. miRNAs play crucial roles in fertility regulation, yet information about their reproductive roles and target genes in tetraploid rice remains limited. Here, we used three tetraploid lines, H1 (fertile), HF (fertile), and LF (sterile), to investigate cytological features and identify factors associated with autotetraploid sterility. LF showed abnormal meiosis, resulting in low pollen fertility and viability, ultimately leading to scarce fertilization and a low-seed setting compared to H1 and HF. RNA-seq revealed 30 miRNA-candidate target pairs related to autotetraploid pollen sterility. These pairs showed opposite expression patterns, with differential expression between fertile lines (H1 and HF) and the sterile line (LF). qRT-PCR confirmed that miR9564, miR528, and miR27874 were highly expressed in the anthers of H1 and HF but not in LF, while opposite results were obtained in their targets (ARPS, M2T, and OsRPC53). Haplotype and expression pattern analyses revealed that ARPS was specifically expressed in lines with the same haplotype of MIR9564 (the precursor of miR9564) as LF. Furthermore, the Dual-GFP assay verified that miR9564 inhibited the fluorescence signal of ARPS-GFP. The over-expression of ARPS significantly decreased the seed setting rate (59.10%) and pollen fertility (50.44%) of neo-tetraploid rice, suggesting that ARPS plays important roles in autotetraploid pollen sterility. This study provides insights into the cytological characteristic and miRNA expression profiles of tetraploid lines with different fertility, shedding light on the role of miRNAs in polyploid rice.

Seed development-related genes contribute to high yield heterosis in integrated utilization of elite autotetraploid and neo-tetraploid rice.

Introduction: Autotetraploid rice holds high resistance to abiotic stress and substantial promise for yield increase, but it could not be commercially used because of low fertility. Thus, our team developed neo-tetraploid rice with high fertility and hybrid vigor when crossed with indica autotetraploid rice. Despite these advances, the molecular mechanisms underlying this heterosis remain poorly understood. Methods: An elite indica autotetraploid rice line (HD11) was used to cross with neo-tetraploid rice, and 34 hybrids were obtained to evaluate agronomic traits related to yield. WE-CLSM, RNA-seq, and CRISPR/Cas9 were employed to observe endosperm structure and identify candidate genes from two represent hybrids. Results and discussion: These hybrids showed high seed setting and an approximately 55% increase in 1000-grain weight, some of which achieved grain yields comparable to those of the diploid rice variety. The endosperm observations indicated that the starch grains in the hybrids were more compact than those in paternal lines. A total of 119 seed heterosis related genes (SHRGs) with different expressions were identified, which might contribute to high 1000-grain weight heterosis in neo-tetraploid hybrids. Among them, 12 genes had been found to regulate grain weight formation, including OsFl3, ONAC023, OsNAC024, ONAC025, ONAC026, RAG2, FLO4, FLO11, OsISA1, OsNF-YB1, NF-YC12, and OsYUC9. Haplotype analyses of these 12 genes revealed the various effects on grain weight among different haplotypes. The hybrids could polymerize more dominant haplotypes of above grain weight regulators than any homozygous cultivar. Moreover, two SHRGs (OsFl3 and SHRG2) mutants displayed a significant reduction in 1000-grain weight and an increase in grain chalkiness, indicating that OsFl3 and SHRG2 positively regulate grain weight. Our research has identified a valuable indica autotetraploid germplasm for generating strong yield heterosis in combination with neo-tetraploid lines and gaining molecular insights into the regulatory processes of heterosis in tetraploid rice.

Lectin Receptor-Like Protein Kinase OsNRFG6 is Required for Embryo Sac Development and Fertilization in Neo-Tetraploid Rice.

Great yield-enhancing prospects of autotetraploid rice was restricted by various polyploidy-induced reproductive dysfunction. To surmount these challenges, our group has generated a series of valuable fertile tetraploid lines (denoted as neo-tetraploid rice) through 20-year efforts. With this context, a G-type lectin receptor-like kinase, OsNRFG6, was identified as a pivotal factor associated with reproductive regulation in neo-tetraploid rice. Nevertheless, it is still elusive about a comprehensive understanding of its precise functional roles and underlying molecular mechanisms during reproduction of neo-tetraploid rice. Here, we demonstrated that OsNRFG6 executed a constitutive expression pattern and encoded proteins localizing in perinucleus and endoplasmic reticulum. Subsequently, four independent mutant lines of OsNRFG6 within neo-tetraploid rice background were further identified, all displaying low seed-setting rate due to abortive embryo sacs and defective double fertilization. RNA-seq and RT-qPCR revealed a significant down-regulation of OsNRFG6 and female reproductive genes such as OsMEL1 and LOG in ovaries prior to and post-fertilization, attributing this effect to OsNRFG6 mutation. Furthermore, through yeast-two hybrids, bimolecular fluorescence complementation assays, and luciferase complementation imaging assays, it was determined that OsNRFG6 could interact with itself and two female reproductive proteins (LOG and OsDES1) to form protein complexes. These results elucidate the reproductive functions and molecular pathway governed by OsNRFG6 in regulating fertility of neo-tetraploid rice, offering insights into molecular understanding of fertility improvement in polyploid rice.

HEMAsNet: A Hemisphere Asymmetry Network Inspired by the Brain for Depression Recognition From Electroencephalogram Signals.

Depression is a prevalent mental disorder that affects a significant portion of the global population. Despite recent advancements in EEG-based depression recognition models rooted in machine learning and deep learning approaches, many lack comprehensive consideration of depression's pathogenesis, leading to limited neuroscientific interpretability. To address these issues, we propose a hemisphere asymmetry network (HEMAsNet) inspired by the brain for depression recognition from EEG signals. HEMAsNet employs a combination of multi-scale Convolutional Neural Network (CNN) and Long Short-Term Memory (LSTM) blocks to extract temporal features from both hemispheres of the brain. Moreover, the model introduces a unique 'Callosum-like' block, inspired by the corpus callosum's pivotal role in facilitating inter-hemispheric information transfer within the brain. This block enhances information exchange between hemispheres, potentially improving depression recognition accuracy. To validate the performance of HEMAsNet, we first confirmed the asymmetric features of frontal lobe EEG in the MODMA dataset. Subsequently, our method achieved a depression recognition accuracy of 0.8067, indicating its effectiveness in increasing classification performance. Furthermore, we conducted a comprehensive investigation from spatial and frequency perspectives, demonstrating HEMAsNet's innovation in explaining model decisions. The advantages of HEMAsNet lie in its ability to achieve more accurate and interpretable recognition of depression through the simulation of physiological processes, integration of spatial information, and incorporation of the Callosum-like block.

Genome-wide and pan-genomic analysis reveals rich variants of NBS-LRR genes in a newly developed wild rice line from Oryza alta Swallen.

Introduction: Oryza alta Swallen is an allotetraploid perennial wild rice and contains CCDD genome, which may harbor favorable genes for the enrichment of genetic resource. Methods: A new wild rice line, Huaye 5, was developed from Oryza alta Swallen in our lab. Whole genome re-sequencing and pan-genomic analysis were employed to analyze its genomic variations and novel genes. Results and Discussion: More than ten million genomic variations were detected when compared with Asian cultivar. Among the variational genes, 724, 197 and 710 genes coded protein kinase, synthetase and transcription factor, respectively. A total of 353, 131 and 135 variational genes were associated with morphological trait, physiological trait, resistance or tolerance, respectively. A total of 62 were NBS-LRR genes were detected, in which 11 NBS-LRR genes expressed in sheath and mature stem, and 26 expressed in young and mature roots expressed. The pan-genome sequences of wild rice species with CCDD genome were constructed by integrating 8 Oryza alta (OA), 2 Oryza grandiglumis (OG) and 18 Oryza latifolia (OL) accessions. A total of 28 non-reference NBS-LRR genes were revealed, and 7 of which were mainly expressed in mature roots. This research demonstrated rich DNA variation in the Oryza alta Swallen that may provide a new germplasm for rice resistance breeding.

Nomogram to predict prognosis of head and neck rhabdomyosarcoma patients in children and adolescents.

Purpose: This study aims to explore the prognostic factors of head and neck rhabdomyosarcoma (HNRMS) in children and adolescents and construct a simple but reliable nomogram model for estimating overall survival (OS) of patients. Methods: Data of all HNRMS patients during 2004-2018 were identified from the Surveillance, Epidemiology, and End Result database. Kaplan-Meier method was performed to calculate OS stratified by subgroups and comparison between subgroups was completed by log-rank test. Univariate and multivariate Cox regressions analysis were employed for identifying independent predictors, which subsequently were used for a predictive model by R software, and the efficacy of the model was evaluated by applying receiver operating curve (ROC), calibration and decision curve analysis (DCA). Results: A total of 446 patients were included in the study. The 1-, 3-, and 5-year OS rate of the whole cohort was 90.6%, 80.0%, and 75.5%, respectively. The results of univariate and multivariate Cox regression analysis indicated that the primary site in parameningeal region, alveolar RMS histology, M1 stage, IRS stage 4, surgery, and chemotherapy were significant prognostic factors (all P<0.05). The performance of nomogram model was validated by discrimination and calibration, with AUC values of 1, 3, and 5 years OS of 0.843, 0.851, and 0.890, respectively. Conclusion: We constructed a prognostic nomogram model for predicting the OS in HNRMS patients in children and adolescents and this model presented practical and applicable clinical value to predict survival when choosing treatment strategies.

OsRH52A, a DEAD-box protein, regulates functional megaspore specification and is required for embryo sac development in rice.

The development of the embryo sac is an important factor that affects seed setting in rice. Numerous genes associated with embryo sac (ES) development have been identified in plants; however, the function of the DEAD-box RNA helicase family genes is poorly known in rice. Here, we characterized a rice DEAD-box protein, RH52A, which is localized in the nucleus and cytoplasm and highly expressed in the floral organs. The knockout mutant rh52a displayed partial ES sterility, including degeneration of the ES (21%) and the presence of a double-female-gametophyte (DFG) structure (11.8%). The DFG developed from two functional megaspores near the chalazal end in one ovule, and 3.4% of DFGs were able to fertilize via the sac near the micropylar pole in rh52a. RH52A was found to interact with MFS1 and ZIP4, both of which play a role in homologous recombination in rice meiosis. RNA-sequencing identified 234 down-regulated differentially expressed genes associated with reproductive development, including two, MSP1 and HSA1b, required for female germline cell specification. Taken together, our study demonstrates that RH52A is essential for the development of the rice embryo sac and provides cytological details regarding the formation of DFGs.

OsmiR5519 regulates grain size and weight and down-regulates sucrose synthase gene RSUS2 in rice (Oryza sativa L.).

MAIN CONCLUSION: The up-regulation of OsmiR5519 results in the decrease of grain size, weight and seed setting rate. OsmiR5519 plays important roles in the process of grain filling and down-regulates sucrose synthase gene RSUS2. MicroRNAs (miRNAs) are one class of small non-coding RNAs that act as crucial regulators of plant growth and development. In rice, the conserved miRNAs were revealed to regulate the yield components, but the function of rice-specific miRNAs has been rarely studied. The rice-specific OsmiR5519 was found to be abundantly expressed during reproductive development, but its biological roles remain unknown. In this study, the function of rice-specific OsmiR5519 was characterized with the miR5519-overexpressing line (miR5519-OE) and miR5519-silenced line (STTM5519). At seedling stage, the content of sucrose, glucose and fructose was obviously lower in the leaves of miR5519-OE lines than those of wild-type (WT) line. The grain size and weight were decreased significantly in miR5519-OE lines, compared to those of WT rice. The cell width of hull in miR5519-OE was smaller than that in WT. The seed setting rate was notably reduced in miR5519-OE lines, but not in STTM5519 lines. Cytological observation demonstrated that the inadequate grain filling was the main reason for the decline of seed setting rate in miR5519-OE lines. The percentage of the defects of grain amounted to 40% in miR5519-OE lines, which almost equaled to the decreased value of seed setting rate. Furthermore, the sucrose synthase gene RSUS2 was identified as a target of OsmiR5519 via RNA ligase-mediated 3'-amplification of cDNA ends (3'-RLM-RACE), dual luciferase assays and transient expression assays. In summary, our results suggest that OsmiR5519 regulates grain size and weight and down-regulates RSUS2 in rice.

MORE FLORET1 controls anther development by negatively regulating key tapetal genes in both diploid and tetraploid rice.

Polyploid hybrid rice (Oryza sativa) has great potential for increasing yields. However, hybrid rice depends on male fertility and its regulation, which is less well studied in polyploid rice than in diploid rice. We previously identified an MYB transcription factor, MORE FLORET1 (MOF1), whose mutation causes male sterility in neo-tetraploid rice. MOF1 expression in anthers peaks at anther Stage 7 (S7) and progressively decreases to low levels at S10. However, it remains unclear how the dynamics of MOF1 expression contribute to male fertility. Here, we carefully examined anther development in both diploid and tetraploid mof1 rice mutants, as well as lines ectopically expressing MOF1 in a temporal manner. MOF1 mutations caused delayed degeneration of the tapetum and middle layer of anthers and aberrant pollen wall organization. Ectopic MOF1 expression at later stages of anther development led to retarded cytoplasmic reorganization of tapetal cells. In both cases, pollen grains were aborted and seed production was abolished, indicating that precise control of MOF1 expression is essential for male reproduction. We demonstrated that 5 key tapetal genes, CYP703A3 (CYTOCHROME P450 HYDROXYLASE 703A3), OsABCG26 (O. sativa ATP BINDING CASSETTE G26), PTC1 (PERSISTENT TAPETAL CELL1), PKS2 (POLYKETIDE SYNTHASE 2), and OsABCG15 (O. sativa ATP BINDING CASSETTE G15), exhibit expression patterns opposite to those of MOF1 and are negatively regulated by MOF1. Moreover, DNA affinity purification sequencing (DAP-seq), luciferase activity assays, and electrophoretic mobility shift assays indicated that MOF1 binds directly to the PKS2 promoter for transcriptional repression. Our results provide a mechanistic basis for the regulation of male reproduction by MOF1 in both diploid and tetraploid rice. This study will facilitate the development of polyploid male sterile lines, which are useful for breeding of polyploid hybrid rice.

Experimental research on compressive strength deterioration of coal seam floor sandstone under the action of acidic mine drainage.

In sulphur-coal symbiotic coal seams, after the mining of sulphide iron ore, when the coal resources are mined, the mine water accumulated in the roadway mining area will have a certain impact on the stability of the surrounding rock of the coal seam roadway. Taking the floor sandstone of sulfur coal symbiotic coal seam as the research object, the roof fissure water with pH values of 7.48, 4.81 and 2.62 was used as the experimental solution. 10 experimental schemes were designed to measure the compressive strength of the samples under the action of AMD, and the hydrochemical analysis of AMD was conducted. The pore structures of the samples before and after the action of AMD were analyzed. Based on the hydrochemistry and pore structure, the deterioration mechanism of compressive strength of the coal seam floor sandstone under the action of AMD was explained. The results indicated that the compressive strength of the samples decreased with the increasing action time of AMD. The compressive strength decreased with the increment of the porosity. The concentration of H+ ion in AMD was relatively small. Na2O in albite dissolved and reacted with water, leading to an increase in the concentration of Na+ ion. Soluble substances such as MgCl2 and CaSO4 in the pore structure dissolved, leading to an increase in the concentration of Ca2+ and Mg2+ ions. The dissolution of soluble substances and the physical-chemical reactions between solutions and minerals were the essential causes of the continuous deterioration of the compressive strength of the coal seam floor sandstone. The results of this study can provide a theoretical basis for the deterioration of the mechanical properties of the peripheral rock in the roadway of the sulphur coal seam, and can also provide a certain engineering reference for the sulphur coal seam roadway.

OsmiRNA5488 Regulates the Development of Embryo Sacs and Targets OsARF25 in Rice (Oryza sativa L.).

Small RNAs are a class of non-coding RNAs that typically range from 20 to 24 nucleotides in length. Among them, microRNAs (miRNAs) are particularly important regulators for plant development. The biological function of the conserved miRNAs has been studied extensively in plants, while that of the species-specific miRNAs has been studied in-depth. In this study, the regulatory role of a rice-specific OsmiRNA5488 (OsmiR5488) was characterized with the miR5488-overexpressed line (miR5488-OE) and miR5488-silenced line (STTM-5488). The seed-setting rate was notably reduced in miR5488-OE lines, but not in STTM-5488 lines. Cytological observation demonstrated the different types of abnormal mature embryo sacs, including the degeneration of embryo sacs and other variant types, in miR5488-OE lines. The percentage of the abnormal mature embryo sacs accounted for the reduced value of the seed-setting rate. Furthermore, OsARF25 was identified as a target of OsmiR5488 via RNA ligase-mediated 3'-amplifification of cDNA ends, dual luciferase assays, and transient expression assays. The primary root length was decreased with the increases in auxin concentrations in miR5488-OE lines compared to wild-type rice. Summarily, our results suggested that OsmiR5488 regulates the seed-setting rate and down-regulates the targeted gene OsARF25.

Autotetraploid rice: challenges and opportunities.

Autotetraploid rice is a type of germplasm developed from the whole genome duplication of diploid rice, leading to large grains, high nutrient content, and resistance. However, its low fertility has reduced yield and hampered commercialization. To address this issue, a new type of high fertility tetraploid rice was developed, which may serve as a useful germplasm for polyploid rice breeding. In this review, we summarize the progress made in understanding the cellular and molecular genetic mechanisms underlying the low fertility of autotetraploid rice and its F1 hybrid, as well as the main types of new tetraploid rice with high fertility. Lastly, the idea of utilizing the multi-generation heterosis of neo-tetraploid rice in the future is proposed as a reference for polyploid rice breeding.

Comparative cytological and transcriptome analyses of ny2 mutant delayed degeneration of tapetal cells and promotes abnormal microspore development in neo-tetraploid rice.

We aimed to investigate the genetic defects related to pollen development and infertility in NY2, a novel tetraploid rice germplasm known as Neo-tetraploid rice. This rice variety was created through the crossbreeding and selective breeding of various autotetraploid rice lines and has previously shown high fertility. Our previous research has revealed that the NY2 gene, encoding a eukaryotic translation initiation factor 3 subunit E, regulates pollen fertility. However, the underlying mechanism behind this fertility is yet to be understood. To shed light on this matter, we performed a combined cytological and transcriptome analysis of the NY2 gene. Cytological analysis indicated that ny2 underwent abnormal tapetal cells, microspore, and middle layer development, which led to pollen abortion and ultimately to male sterility. Genetic analysis revealed that the F1 plants showed normal fertility and an obvious advantage for seed setting compared to ny2. Global gene expression analysis in ny2 revealed a total of 7545 genes were detected at the meiosis stage, and 3925 and 3620 displayed upregulation and downregulation, respectively. The genes were significantly enriched for the gene ontology (GO) term "carbohydrate metabolic process. Moreover, 9 genes related to tapetum or pollen fertility showed down-regulation, such as OsABCG26 (ATP Binding Cassette G26), TMS9-1 (Thermosensitive Male Sterility), EAT1 (Programmed cell death regulatory), KIN14M (Kinesin Motor), OsMT1a (Metallothionein), and OsSTRL2 (Atypical strictosidine synthase), which were validated by qRT-PCR. Further analyses of DEGs identified nine down-regulated transcription factor genes related to pollen development. NY2 is an important regulator of the development of tapetum and microspore. The regulatory gene network described in this study may offer important understandings into the molecular processes that underlie fertility control in tetraploid rice.

In Vitro Induction of Interspecific Hybrid and Polyploidy Derived from Oryza officinalis Wall.

Oryza officinalis Wall is a potential genetic resource for rice breeding; however, its distant genome limits its crossing ability with cultivated rice. The interspecific hybridization of O. officinalis and cultivated rice, establishment of its tissue culture, and induction of polyploidy are ways to improve O. officinalis's poor crossability. We developed an interspecific hybrid and studied its reproductive pollen development process in this work, and the results showed that abortive pollens (81.94%) and embryo sac abnormalities (91.04%) were the key causes of its high sterility. In order to induce callus formation in interspecific hybrid explants, two different culture media, namely Chu's N-6 medium (N6) and 1/2 Murashig and Skoog medium (1/2 MS), were employed. Additionally, two plant growth regulators (PGRs), namely 2,4-dichlorophenoxyacetic acid (2,4-D) and 6-benzylaminopurine (BA), along with L-proline (Pro) and acid hydrolyzed casein, were utilized in the experiment. The optimal N6 medium, supplemented with 2.0 mg·L-1 2,4-D, produced the highest induction rate (80.56 ± 5.44)%. For callus differentiation and proliferation, the MS medium supplemented with 2.0 mg·L-1 BA + 0.2 mg·L-1 NAA produced the highest differentiation rate (75.00 ± 4.97)% and seedling emergence ratio (28.97 ± 4.67)%. The optimal combination for seedling rooting was the 1/2 MS medium supplemented with 2.0 mg L-1 NAA and 0.2 mg L-1 BA, which produced an average of 13.95 roots per plant. For polyploidy induction in the interspecific hybrid, the concentration of colchicine treatment at 400 mg·L-1 for three days is an ideal protocol. We devised tissue culture and interspecific hybrid polyploidy induction to overcome O. officinalis' poor crossability and introduce its favorable features into cultivated rice.

Inhibiting HIF-1 signaling alleviates HTRA1-induced RPE senescence in retinal degeneration.

BACKGROUND: Age-related macular degeneration (AMD), characterized by the degeneration of retinal pigment epithelium (RPE) and photoreceptors, is the leading cause of irreversible vision impairment among the elderly. RPE senescence is an important contributor to AMD and has become a potential target for AMD therapy. HTRA1 is one of the most significant susceptibility genes in AMD, however, the correlation between HTRA1 and RPE senescence hasn't been investigated in the pathogenesis of AMD. METHODS: Western blotting and immunohistochemistry were used to detect HTRA1 expression in WT and transgenic mice overexpressing human HTRA1 (hHTRA1-Tg mice). RT-qPCR was used to detect the SASP in hHTRA1-Tg mice and ARPE-19 cells infected with HTRA1. TEM, SA-ß-gal was used to detect the mitochondria and senescence in RPE. Retinal degeneration of mice was investigated by fundus photography, FFA, SD-OCT and ERG. The RNA-Seq dataset of ARPE-19 cells treated with adv-HTRA1 versus adv-NC were analyzed. Mitochondrial respiration and glycolytic capacity in ARPE-19 cells were measured using OCR and ECAR. Hypoxia of ARPE-19 cells was detected using EF5 Hypoxia Detection Kit. KC7F2 was used to reduce the HIF1α expression both in vitro and in vivo. RESULTS: In our study, we found that RPE senescence was facilitated in hHTRA1-Tg mice. And hHTRA1-Tg mice became more susceptible to NaIO3 in the development of oxidative stress-induced retinal degeneration. Similarly, overexpression of HTRA1 in ARPE-19 cells accelerated cellular senescence. Our RNA-seq revealed an overlap between HTRA1-induced differentially expressed genes associated with aging and those involved in mitochondrial function and hypoxia response in ARPE-19 cells. HTRA1 overexpression in ARPE-19 cells impaired mitochondrial function and augmented glycolytic capacity. Importantly, upregulation of HTRA1 remarkably activated HIF-1 signaling, shown as promoting HIF1α expression which mainly located in the nucleus. HIF1α translation inhibitor KC7F2 significantly prevented HTRA1-induced cellular senescence in ARPE-19 cells, as well as improved the visual function in hHTRA1-Tg mice treated with NaIO3. CONCLUSIONS: Our study showed elevated HTRA1 contributes to the pathogenesis of AMD by promoting cellular senescence in RPE through damaging mitochondrial function and activating HIF-1 signaling. It also pointed out that inhibition of HIF-1 signaling might serve as a potential therapeutic strategy for AMD. Video Abstract.

Natural allelic variation in GRAIN SIZE AND WEIGHT 3 of wild rice regulates the grain size and weight.

Grain size is important for yield in rice (Oryza sativa L.). Although many genes involved in grain size have been isolated, few can be used in breeding due to their interactions and phenotypic effects. Here, we describe natural variation in the granule-type quantitative trait locus GRAIN SIZE AND WEIGHT 3 (GSW3) located on chromosome 3 in wild rice (Oryza rufipogon Griff.) that encodes a GTPase-regulated protein and negatively regulates grain length, grain width, and 1,000-grain weight. The insertion of a 232-bp fragment of the genomic sequence in the wild rice, a natural allelic variant gene (GSW3), increased the expression levels and reduced the grain length and width and 1,000-grain weight. Knockout of GSW3 in the wild rice inbred line Huaye 3 increased the grain length and width and 1,000-grain weight. Introducing GSW3Huaye3 into cultivated rice line KJ01 and overexpressing GSW3Huaye3 in Huaye 3 resulted in reduced grain length and width and 1,000-grain weight, and grain size and 1,000-grain weight changes were closely related to GSW3 expression levels. GSW3 regulated the grain length and width simultaneously by promoting grain glume cell division and longitudinal and transverse cell growth. GSW3 was also involved in regulating the gibberellic acid signaling pathway and negatively regulated plant growth. Furthermore, a critical SNP in the GSW3 coding region was obviously correlated with grain size variation in a core collection of cultivated rice. This SNP resulted in an amino acid substitution from Gln to Arg at position 161 in GSW3, which reduced the grain size. Our study shows that GSW3 negatively regulates the grain shape, which could explain different grain shapes in modern cultivars and wild rice. GSW3 may also be used for breeding rice varieties with improved grain shapes and higher yield.

Polyploidy and zinc oxide nanoparticles alleviated Cd toxicity in rice by modulating oxidative stress and expression levels of sucrose and metal-transporter genes.

The Cd toxicity causes severe perturbations to the plant's growth and development. Here, polyploid and diploid rice lines were treated with zinc-oxide nanoparticles (ZnO-NPs) and Cd, and physiological, cytological and molecular changes were observed. The Cd toxicity significantly reduced plant's growth attributes (such as shoot length, biological yield, dry matter, and chlorophyll contents, which decreased by 19%, 18%, 16%, 19% in polyploid and 35%, 43%, 45% and 43% in diploid rice, respectively), and disturbed the sugar level through the production of electrolytes, hydrogen peroxide, and malondialdehyde. The application of ZnO-NPs significantly alleviated the Cd toxicity in both lines by improving the antioxidant enzymes activities and physiochemical attributes. Semi-thin sections and transmission electron microscope revealed more and different types of abnormalities in diploid rice compared to polyploid rice under Cd stress. Moreover, RNA-seq analysis identified several differentially expressed genes between polyploid and diploid rice, especially metal and sucrose transporter genes. The GO, COG, and KEGG analyses revealed ploidy-specific pathways associated with plant growth and development. In conclusion, ZnO-NPs application to both rice lines significantly improved plant growth and decreased Cd accumulation in plants. We inferred that polyploid rice is more resistant to Cd stress than diploid rice.