N4-acetylation of cytidine in mRNA plays essential roles in plants.

The biological function of RNA can be modulated by base modifications. Here, we unveiled the occurrence of N4-acetylation of cytidine in plant RNA, including mRNA, by employing LC-MS/MS and acRIP-seq. We identified 325 acetylated transcripts from the leaves of 4-week-old Arabidopsis (Arabidopsis thaliana) plants and determined that 2 partially redundant N-ACETYLTRANSFERASEs FOR CYTIDINE IN RNA (ACYR1 and ACYR2), which are homologous to mammalian NAT10, are required for acetylating RNA in vivo. A double-null mutant was embryo lethal, while eliminating 3 of the 4 ACYR alleles led to defects in leaf development. These phenotypes could be traced back to the reduced acetylation and concomitant destabilization of the transcript of TOUGH, which is required for miRNA processing. These findings indicate that N4-acetylation of cytidine is a modulator of RNA function with a critical role in plant development and likely many other processes.

Pyrimidine catabolism is required to prevent the accumulation of 5-methyluridine in RNA.

5-Methylated cytosine is a frequent modification in eukaryotic RNA and DNA influencing mRNA stability and gene expression. Here we show that free 5-methylcytidine (5mC) and 5-methyl-2'-deoxycytidine are generated from nucleic acid turnover in Arabidopsis thaliana, and elucidate how these cytidines are degraded, which is unclear in eukaryotes. First CYTIDINE DEAMINASE produces 5-methyluridine (5mU) and thymidine which are subsequently hydrolyzed by NUCLEOSIDE HYDROLASE 1 (NSH1) to thymine and ribose or deoxyribose. Interestingly, far more thymine is generated from RNA than from DNA turnover, and most 5mU is directly released from RNA without a 5mC intermediate, since 5-methylated uridine (m5U) is an abundant RNA modification (m5U/U ∼1%) in Arabidopsis. We show that m5U is introduced mainly by tRNA-SPECIFIC METHYLTRANSFERASE 2A and 2B. Genetic disruption of 5mU degradation in the NSH1 mutant causes m5U to occur in mRNA and results in reduced seedling growth, which is aggravated by external 5mU supplementation, also leading to more m5U in all RNA species. Given the similarities between pyrimidine catabolism in plants, mammals and other eukaryotes, we hypothesize that the removal of 5mU is an important function of pyrimidine degradation in many organisms, which in plants serves to protect RNA from stochastic m5U modification.

Visual reading for [18F]Florzolotau Tau PET scans in progressive supranuclear palsy.

PURPOSE: The identification of tau accumulation within living brains holds significant potential in facilitating accurate diagnosis of progressive supranuclear palsy (PSP). While visual assessment is frequently employed, standardized methods for tau positron emission tomography (PET) specifically in PSP are absent. We aimed to develop a visual reading algorithm dedicated to the evaluation of [18F]Florzolotau PET in PSP. METHODS: 148 PSP and 30 healthy volunteers were divided into a development set (for the establishment of the reading rules; n = 89) and a testing set (for the validation of the reading rules; n = 89). For differential diagnosis, 55 α-synucleinopathies were additionally included into the testing set. The visual reading method was established by an experienced assessor (Reader 0) and was then validated by Reader 0 and two additional readers on regional and overall binary manners. A positive binding in both midbrain and globus pallidus/putamen regions was characterized as a PSP-like pattern, whereas any other pattern was classified as non-PSP-like. RESULTS: Reader 1 (94.4%) and Reader 2 (93.8%) showed excellent agreement for the overall binary determination against Reader 0. The regional binary determinations of midbrain and globus pallidus/putamen showed excellent agreement among readers (kappa > 0.80). The overall binary evaluation demonstrated reproducibility of 86.1%, 94.4% and 77.8% for three readers. The visual reading algorithm showed high agreement with regional standardized uptake value ratios and clinical diagnoses. CONCLUSION: Through the application of the suggested visual reading algorithm, [18F]Florzorotau PET imaging demonstrated a robust performance for the imaging diagnosis of PSP.

Physically based simulation of focusing schlieren imaging for a hypersonic boundary layer flow.

Focusing schlieren systems are more advantageous than conventional schlieren systems in providing a schlieren image with certain spatial discrimination along the light path. The present work employed a hybrid of the optical-transfer matrix and ray-tracing method to faithfully replicate complete physical imaging processes throughout a focusing schlieren optic system. A direct numerical simulation of a hypersonic boundary layer flow was employed to synthesize focusing schlieren images. The influence of various configuration parameters on the properties of focusing schlieren image such as local schlieren structure, brightness, sensitivity, and depth of field were systematically explored. In addition, an approximation method was proposed as a simplified means to facilitate the simulation of a focusing schlieren image.

Psychometric properties of the Chinese version of the Hypertension Belief Assessment Tool.

BACKGROUND: Hypertension is prevalent in China. Hypertensive patients suffer from many health problems in life. Hypertension is a common chronic disease with long-term and lifelong characteristics. In the long run, the existence of chronic diseases will affect the patient's own health beliefs. However, people's health beliefs about Hypertension are not explicit. Therefore, it is vital to find a suitable instrument to comprehend and improve the health beliefs of hypertensive patients, thus, better control of blood pressure and improvement of patient's quality of life are now crucial issues. This study aimed to translate the Hypertension Belief Assessment Tool (HBAT) into Chinese and examine the psychometric properties of the Chinese version of the Hypertension Belief Assessment Tool in hypertensive patients. METHODS: This is a cross-sectional study. We translated the HBAT into Chinese and tested the reliability and validity of the Chinese version among 325 hypertensive patients. RESULTS: The Chinese version of the scale contains 21 items. The Exploratory Factor Analysis (EFA) revealed six factors and explained 77.898% of the total variation. A six-factor model eventually showed acceptable fit indices in the Confirmatory Factor Analysis (CFA). With modified Confirmatory Factor Analysis, the fit indices were Chi-square/Degree of Freedom (CMIN/DF) = 2.491, Comparative Fit Index (CFI) = 0.952, Incremental Fit Index (IFI) = 0.952, Root-mean-square Error of Approximation (RMSEA) = 0.068, Tucker Lewis Index (TLI) = 0.941. The HBAT exhibits high internal consistency reliability (0.803), and the scale has good discriminant validity. CONCLUSION: The results suggest that the HBAT is a reliable and valid instrument for assessing the beliefs of Chinese hypertensive patients.

A Kinase and a Glycosylase Catabolize Pseudouridine in the Peroxisome to Prevent Toxic Pseudouridine Monophosphate Accumulation.

Pseudouridine (Ψ) is a frequent nucleoside modification that occurs in both noncoding RNAs and mRNAs. In pseudouridine, C5 of uracil is attached to the Rib via an unusual C-glycosidic bond. This RNA modification is introduced on the RNA by site-specific transglycosylation of uridine (U), a process mediated by pseudouridine synthases. RNA is subject to constant turnover, releasing free pseudouridine, but the metabolic fate of pseudouridine in eukaryotes is unclear. Here, we show that in Arabidopsis (Arabidopsis thaliana), pseudouridine is catabolized in the peroxisome by (1) a pseudouridine kinase (PUKI) from the PfkB family that generates 5'-pseudouridine monophosphate (5'-ΨMP) and (2) a ΨMP glycosylase (PUMY) that hydrolyzes ΨMP to uracil and ribose-5-phosphate. Compromising pseudouridine catabolism leads to strong pseudouridine accumulation and increased ΨMP content. ΨMP is toxic, causing delayed germination and growth inhibition, but compromising pseudouridine catabolism does not affect the Ψ/U ratios in RNA. The bipartite peroxisomal PUKI and PUMY are conserved in plants and algae, whereas some fungi and most animals (except mammals) possess a PUMY-PUKI fusion protein, likely in mitochondria. We propose that vacuolar turnover of ribosomal RNA produces most of the pseudouridine pool via 3'-ΨMP, which is imported through the cytosol into the peroxisomes for degradation by PUKI and PUMY, a process involving a toxic 5'-ΨMP intermediate.

18 F-Florzolotau Positron Emission Tomography Imaging of Tau Pathology in the Living Brains of Patients with Corticobasal Syndrome.

BACKGROUND: Recent development in tau-sensitive tracers has sparkled significant interest in tracking tauopathies using positron emission tomography (PET) biomarkers. However, the ability of 18 F-florzolotau PET imaging to topographically characterize tau pathology in corticobasal syndrome (CBS) remains unclear. Further, the question as to whether disease-level differences exist with other neurodegenerative tauopathies is still unanswered. OBJECTIVE: To analyze the topographical patterns of tau pathology in the living brains of patients with CBS using 18 F-florzolotau PET imaging and to examine whether differences with other tauopathies exist. METHODS: 18 F-florzolotau PET imaging was performed in 20 consecutive patients with CBS, 20 cognitively healthy controls (HCs), 20 patients with Alzheimer's disease (AD), and 16 patients with progressive supranuclear palsy-Richardson's syndrome (PSP-RS). Cerebrospinal fluid (CSF) levels of ß-amyloid biomarkers were quantified in all patients with CBS. 18 F-florzolotau uptake was quantitatively assessed using standardized uptake value ratios. RESULTS: Of the 20 patients with CBS, 19 (95%) were negative for CSF biomarkers of amyloid pathology; of them, three had negative 18 F-florzolotau PET findings. Compared with HCs, patients with CBS showed increased 18 F-florzolotau signals in both cortical and subcortical regions. In addition, patients with CBS were characterized by higher tracer retentions in subcortical regions compared with those with AD and showed a trend toward higher signals in cortical areas compared with PSP-RS. An asymmetric pattern of 18 F-florzolotau uptake was associated with an asymmetry of motor severity in patients with CBS. CONCLUSIONS: In vivo 18 F-florzolotau PET imaging holds promise for distinguishing CBS in the spectrum of neurodegenerative tauopathies. © 2023 International Parkinson and Movement Disorder Society.

The relationship between psychological resilience and quality of life among the Chinese diabetes patients: the mediating role of stigma and the moderating role of empowerment.

BACKGROUND: Although some factors, such as stigma and empowerment, influence the complex relationship between psychological resilience and quality of life, few studies have explored similar psychological mechanisms among patients with diabetes. Therefore, this study explored the mediating role of stigma and the moderating role of empowerment in the psychological mechanisms by which psychological resilience affects quality of life. METHODS: From June to September 2022, data were collected by multi-stage stratified sampling and random number table method. Firstly, six tertiary hospitals in Wuhu were numbered and then selected using the random number table method, resulting in the First Affiliated Hospital of Wannan Medical College being selected. Secondly, two departments were randomly selected from this hospital: endocrinology and geriatrics. Thirdly, survey points were set up in each department, and T2DM patients were randomly selected for questionnaire surveys. In addition, we used the Connor-Davidson Elasticity Scale (CD-RISC) to measure the psychological resilience of patients, and used the Stigma Scale for Chronic Illness (SSCI) to measure stigma. Empowerment was measured by the Diabetes Empowerment Scale (DES). Quality of Life was assessed by the Diabetes Quality of Life Scale (DQoL). We used SPSS (version 21) and PROCESS (version 4.1) for data analysis. RESULTS: (1) Psychological resilience was negatively correlated with stigma and quality of life, and positively correlated with empowerment. Stigma was positively associated with empowerment and quality of life. Empowerment was negatively correlated with quality of life. (2) The mediation analysis showed that psychological resilience had a direct predictive effect on the quality of life, and stigma partially mediated the relationship; Empowerment moderates the first half of "PR → stigma → quality of life"; Empowerment moderates the latter part of "PR → stigma → quality of life." CONCLUSIONS: Under the mediating effect of stigma, psychological resilience can improve quality of life. Empowerment has a moderating effect on the relationship between psychological resilience and stigma, and it also has a moderating effect on the relationship between stigma and quality of life. These results facilitate the understanding of the relationship mechanisms between psychological resilience and quality of life.

Red-Light-Responsive Metallopolymer Nanocarriers with Conjugated and Encapsulated Drugs for Phototherapy Against Multidrug-Resistant Tumors.

It is challenging to treat multidrug-resistant tumors because such tumors are resistant to a broad spectrum of structurally and functionally unrelated drugs. Herein, treatment of multidrug-resistant tumors using red-light-responsive metallopolymer nanocarriers that are conjugated with the anticancer drug chlorambucil (CHL) and encapsulated with the anticancer drug doxorubicin (DOX) is reported. An amphiphilic metallopolymer PolyRuCHL that contains a poly(ethylene glycol) (PEG) block and a red-light-responsive ruthenium (Ru)-containing block is synthesized. Chlorambucil is covalently conjugated to the Ru moieties of PolyRuCHL. Encapsulation of DOX into PolyRuCHL in an aqueous solution results in DOX@PolyRuCHL micelles. The DOX@PolyRuCHL micelles are efficiently taken up by the multidrug-resistant breast cancer cell line MCF-7R and which carries DOX into the cells. Free DOX, without the nanocarriers, is not taken up by MCF-7R or pumped out of MCF-7R via P-glycoproteins. Red light irradiation of DOX@PolyRuCHL micelles triggers the release of chlorambucil-conjugated Ru moieties and DOX. Both act synergistically to inhibit the growth of multidrug-resistant cancer cells. Furthermore, the inhibition of the growth of multidrug-resistant tumors in a mouse model using DOX@PolyRuCHL micelles is demonstrated. The design of red-light-responsive metallopolymer nanocarriers with both conjugated and encapsulated drugs opens up an avenue for photoactivated chemotherapy against multidrug-resistant tumors.

18 F-Florzolotau Tau Positron Emission Tomography Imaging in Patients with Multiple System Atrophy-Parkinsonian Subtype.

BACKGROUND: Anecdotal evidence suggests that patients diagnosed with the parkinsonian subtype of multiple system atrophy (MSA-P) may show uptake of the second-generation tau positron emission tomography (PET) tracer 18 F-Florzolotau (previously known as 18 F-APN-1607) in the putamen. OBJECTIVES: This study systematically investigated the localization and magnitude of 18 F-Florzolotau uptake in a relatively large cohort of patients with MSA-P. METHODS: 18 F-Florzolotau PET imaging was performed in 31 patients with MSA-P, 24 patients with Parkinson's disease (PD), and 20 age-matched healthy controls. 18 F-Florzolotau signal in the striatum was analyzed by visual inspection and classified as either positive or negative. Regional 18 F-Florzolotau binding was also expressed as standardized uptake value ratio (SUVR) to assess whether it was associated with core symptoms of MSA-P after adjustment for potential confounders. RESULTS: By visual inspection and semiquantitative SUVR comparisons, patients with MSA-P showed elevated 18 F-Florzolotau uptake in the putamen, globus pallidus, and dentate-a finding that was not observed in PD. This increased signal was significantly associated with the core symptoms of MSA-P. In addition, patients with MSA-P with cerebellar ataxia showed an elevated 18 F-Florzolotau uptake in the cerebellar dentate. CONCLUSIONS: 18 F-Florzolotau tau PET imaging findings may reflect the clinical severity of MSA-P and can potentially discriminate between this condition and PD. © 2022 International Parkinson and Movement Disorder Society.

The mechanism of antigen-presentation of avian bone marrowed dendritic cells suppressed by infectious bronchitis virus.

Dendritic cells are first guard to defend avian infectious bronchitis virus (IBV) infection and invasion. While IBV always suppress dendritic cells and escape the degradation and presentation, which might help viruses to transfer and migrant. Initially, we compared two IBV's function in activating avian bone marrow dendritic cells (BMDCs) and found that both IBV (QX and M41) did not significantly increase surface marker of avian BMDCs. Moreover, a significant decrease of m6A modification level in mRNA, but an increased in the ut RNA were observed in avian BMDCs upon the prevalent IBV (QX) infection. Further study found that both non-structural protein 7 (NSP7) and NSP16 inhibited the maturation and cytokines secretion of BMDCs, as well as their antigen-presentation ability. Lastly, we found that gga-miR21, induced by both NSP7 and NSP16, inhibited the antigen presentation of avian BMDCs. Taken together, our results illustrated how IBV inhibited the antigen-presentation of avian DCs.

m6A RNA Degradation Products Are Catabolized by an Evolutionarily Conserved N6-Methyl-AMP Deaminase in Plant and Mammalian Cells.

N6-methylated adenine (m6A) is the most frequent posttranscriptional modification in eukaryotic mRNA. Turnover of RNA generates N6-methylated AMP (N6-mAMP), which has an unclear metabolic fate. We show that Arabidopsis thaliana and human cells require an N6-mAMP deaminase (ADAL, renamed MAPDA) to catabolize N6-mAMP to inosine monophosphate in vivo by hydrolytically removing the aminomethyl group. A phylogenetic, structural, and biochemical analysis revealed that many fungi partially or fully lack MAPDA, which coincides with a minor role of N6A-RNA methylation in these organisms. MAPDA likely protects RNA from m6A misincorporation. This is required because eukaryotic RNA polymerase can use N6-mATP as a substrate. Upon abrogation of MAPDA, root growth is slightly reduced, and the N6-methyladenosine, N6-mAMP, and N6-mATP concentrations are increased in Arabidopsis. Although this will potentially lead to m6A misincorporation into RNA, we show that the frequency is too low to be reliably detected in vivo. Since N6-mAMP was severalfold more abundant than N6-mATP in MAPDA mutants, we speculate that additional molecular filters suppress the generation of N6-mATP. Enzyme kinetic data indicate that adenylate kinases represent such filters being highly selective for AMP versus N6-mAMP phosphorylation. We conclude that a multilayer molecular protection system is in place preventing N6-mAMP accumulation and salvage.

[The glymphatic system: concept, function and research progresses].

The glymphatic system is a cerebrospinal fluid-interstitial fluid exchange system dependent on the water channel aquaporin-4 polarized on astrocyte endfeet, which is proposed to account for the clearance of abnormal proteins (e.g. ß-amyloid) and metabolites (e.g. lactate) from the brain. Accumulating studies have revealed that glymphatic activity during sleep and general anesthesia is dramatically enhanced, while its function is significantly damaged during aging, traumatic brain injury, Alzheimer's disease, stroke, and diabetes. The glymphatic hypothesis is a breakthrough in the field of neuroscience recently, which would considerably enhance our comprehension on the cerebrospinal fluid circulation and its role in the maintenance of brain homeostasis. In this review, we briefly introduced the conceptualization of glymphatic system, summarized the recent progresses, and prospected its future investigation and potential clinical application.

Of the Nine Cytidine Deaminase-Like Genes in Arabidopsis, Eight Are Pseudogenes and Only One Is Required to Maintain Pyrimidine Homeostasis in Vivo.

CYTIDINE DEAMINASE (CDA) catalyzes the deamination of cytidine to uridine and ammonia in the catabolic route of C nucleotides. The Arabidopsis (Arabidopsis thaliana) CDA gene family comprises nine members, one of which (AtCDA) was shown previously in vitro to encode an active CDA. A possible role in C-to-U RNA editing or in antiviral defense has been discussed for other members. A comprehensive bioinformatic analysis of plant CDA sequences, combined with biochemical functionality tests, strongly suggests that all Arabidopsis CDA family members except AtCDA are pseudogenes and that most plants only require a single CDA gene. Soybean (Glycine max) possesses three CDA genes, but only two encode functional enzymes and just one has very high catalytic efficiency. AtCDA and soybean CDAs are located in the cytosol. The functionality of AtCDA in vivo was demonstrated with loss-of-function mutants accumulating high amounts of cytidine but also CMP, cytosine, and some uridine in seeds. Cytidine hydrolysis in cda mutants is likely caused by NUCLEOSIDE HYDROLASE1 (NSH1) because cytosine accumulation is strongly reduced in a cda nsh1 double mutant. Altered responses of the cda mutants to fluorocytidine and fluorouridine indicate that a dual specific nucleoside kinase is involved in cytidine as well as uridine salvage. CDA mutants display a reduction in rosette size and have fewer leaves compared with the wild type, which is probably not caused by defective pyrimidine catabolism but by the accumulation of pyrimidine catabolism intermediates reaching toxic concentrations.

Role of sagittal and oblique smiling profiles in evaluating facial esthetics.

INTRODUCTION: Surgeons and orthodontists used to use a conventional set of facial photographs, composed of front, front smiling, and profile images to evaluate facial esthetics, whereas sagittal and oblique smiling profile images have been largely neglected in practice. The aim of this study was to explore the importance of sagittal and oblique smiling profiles in evaluating facial esthetics. METHODS: Photographs from 80 patients, of whom 40 underwent orthognathic surgery and 40 underwent orthodontic treatment, including front, front smiling, profile, sagittal profile smiling, and oblique profile smiling images before and after treatment, were collected and synthesized into 6 categories. Thirty judges gave scores to these photographs based on their own esthetic conception with a 1-week interval for each category. RESULTS: The results demonstrated that the mean score change of evaluating facial attractiveness of patients who underwent orthognathic surgery was lower when adding sagittal or oblique smiling profiles before the treatment, whereas it was higher after the treatment, which were opposite to the orthodontic treatment group with a higher score before the treatment and a lower score after the treatment when sagittal or oblique smiling profiles were added. The changes have a significant difference in adding both sagittal smiling profiles (P < 0.05) and oblique smiling profiles (P < 0.05) before and after treatment. CONCLUSIONS: Along with oblique smiling profile, sagittal smiling profile is crucial in evaluating facial esthetics for orthodontic treatment and orthognathic surgery. Both of them suggested to be integrated in routine photographic assessment of facial attractiveness evaluation before and after treatment, especially in orthognathic surgery for facial esthetic evaluation.

A novel elicitor identified from Magnaporthe oryzae triggers defense responses in tobacco and rice.

KEY MESSAGE: Our studies indicate a potential important elicitor candidate which can aid in the fight against a worldwide disease, rice blast. In this study, we report the purification, identification, characterization, and gene cloning of a novel hypersensitive response-inducing protein elicitor (MoHrip2) secreted from an important pathogenic fungus, Magnaporthe oryzae. The protein fraction was isolated from the culture filtrate of M. oryzae and identified by de novo sequencing. The elicitor-encoding gene mohrip2 was cloned following sequence comparison and PCR amplification. This 459-bp gene encodes a 152-residue polypeptide that contains an 18-residue signal peptide and exhibits a pI of 4.72 and an apparent molecular mass of 16 kDa. The hypothetical protein, MoHrip2, was expressed in Escherichia coli, and both the recombinant and the endogenous protein caused necrotic lesions in tobacco leaves. In addition to phenolic compound deposition and alkalization of the extracellular medium, MoHrip2 also induced hydrogen peroxide production and nitric oxide accumulation in tobacco cells. Moreover, rice seedlings treated with MoHrip2 exhibited pronounced resistance to M. oryzae compared with control seedlings.

Endogenous γ-Aminobutyric Acid Accumulation Enhances Salinity Tolerance in Rice.

Rice is an important food crop worldwide but is usually susceptible to saline stress. When grown on soil with excessive salt, rice plants experience osmotic, ionic, and oxidative stresses that adversely affect growth performance. γ-Aminobutyric acid (GABA) is a nonproteinogenic amino acid that plays an important role in the metabolic activities of organisms. Glutamate decarboxylase (GAD) is the rate-limiting enzyme in GABA metabolism. Here, we genetically modified rice GAD by overexpression or CRISPR-mediated genome editing. These lines, named gad3-ox1 and gad3-ox2 or gad1/3-ko, were used to explore the effects of endogenous GABA accumulation on salt tolerance in rice. Both the gad3-ox1 and gad3-ox2 lines exhibited significant accumulation of the GABA content, whereas the gad1/3-ko line presented a reduced GABA content in vivo. Notably, the two overexpression lines were markedly resistant to salt stress compared with the wild-type and knockout lines. Furthermore, our results demonstrated that endogenous GABA accumulation in the gad3-ox1 and gad3-ox2 lines increased the contents of antioxidant substances and osmotic regulators, decreased the content of membrane lipid peroxidation products and the Na+ content, and resulted in strong tolerance to salt stress. Together, these data provide a theoretical basis for cultivating rice varieties with strong salt tolerance.

Multi-Omics Analyses Uncover the Mechanism Underlying Polyploidization-Enhanced Steviol Glycosides Biosynthesis in Stevia rebaudiana.

Stevia rebaudiana (Bertoni) is a valuable sweetener plant whose sweetness primarily derives from steviol glycosides (SGs), especially rebaudioside A (RA). Polyploidization has the potential to enhance the content of active ingredients in medicinal plants, making this strategy a promising avenue for genetic improvement. However, the underlying regulatory mechanisms that contribute to the fluctuating SGs content between autotetraploid and diploid stevia remain unclear. In this study, we employed metabolic analysis to identify 916 differentially accumulated metabolites (DAMs), with the majority, specifically terpenoids, flavonoids, and lipids, exhibiting upregulation due to polyploidization. Notably, the content of stevia's signature metabolite SGs (including RA, steviolbioside, and rebaudioside C), along with their precursor steviol, increased significantly after polyploidization. Furthermore, a comprehensive analysis of the transcriptome and metabolome revealed that the majority of differentially expressed genes (DEGs) involved in the SG-synthesis pathway (ent-KAH, ent-KS1, UGT73E1, UGT74G1, UGT76G1, UGT85C2, and UGT91D2) were upregulated in autotetraploid stevia, and these DEGs exhibited a positive correlation with the polyploidization-enhanced SGs. Additionally, multi-omics network analysis indicated that several transcription factor families (such as five NACs, four WRKYs, three MYBs, eight bHLHs, and three AP2/ERFs), various transporter genes (four ABC transporters, three triose-phosphate transporters, and two sugar efflux transporters for intercellular exchange), as well as microorganisms (including Ceratobasidium and Flavobacterium) were positively correlated with the accumulation of RA and steviol. Overall, our results indicate the presence of a regulatory circuit orchestrated by polyploidization, which recruits beneficial rhizosphere microbes and modulates the expression of genes associated with SG biosynthesis, ultimately enhancing the SG content in stevia. This finding will provide new insights for promoting the propagation and industrial development of stevia.

Genome-Wide Identification of the Soybean AlkB Homologue Gene Family and Functional Characterization of GmALKBH10Bs as RNA m6A Demethylases and Expression Patterns under Abiotic Stress.

Soybean (Glycine max (L.) Merr) is one of the most important crops worldwide, but its yield is vulnerable to abiotic stresses. In Arabidopsis, the AlkB homologue (ALKBH) family genes plays a crucial role in plant development and stress response. However, the identification and functions of its homologous genes in soybean remain obscured. Here, we identified a total of 22 ALKBH genes in soybean and classified them into seven subfamilies according to phylogenetic analysis. Gene duplication events among the family members and gene structure, conserved domains, and motifs of all candidate genes were analyzed. By comparing the changes in the m6A levels on mRNA from hair roots between soybean seedlings harboring the empty vector and those harboring the GmALKBH10B protein, we demonstrated that all four GmALKBH10B proteins are bona fide m6A RNA demethylases in vivo. Subcellular localization and expression patterns of the GmALKBH10B revealed that they might be functionally redundant. Furthermore, an analysis of cis-elements coupled with gene expression data demonstrated that GmALKBH10B subfamily genes, including GmALKBH10B1, GmALKBH10B2, GmALKBH10B3, and GmALKBH10B4, are likely involved in the cis-elements' response to various environmental stimuli. In summary, our study is the first to report the genome-wide identification of GmALKBH family genes in soybean and to determine the function of GmALKBH10B proteins as m6A RNA demethylases, providing insights into GmALKBH10B genes in response to abiotic stresses.