The m6A reader ECT8 is an abiotic stress sensor that accelerates mRNA decay in Arabidopsis.

N 6-methyladenosine (m6A) is the most abundant mRNA modification and plays diverse roles in eukaryotes, including plants. It regulates various processes, including plant growth, development, and responses to external or internal stress responses. However, the mechanisms underlying how m6A is related to environmental stresses in both mammals and plants remain elusive. Here, we identified EVOLUTIONARILY CONSERVED C-TERMINAL REGION 8 (ECT8) as an m6A reader protein and showed that its m6A-binding capability is required for salt stress responses in Arabidopsis (Arabidopsis thaliana). ECT8 accelerates the degradation of its target transcripts through direct interaction with the decapping protein DECAPPING 5 within processing bodies. We observed a significant increase in the ECT8 expression level under various environmental stresses. Using salt stress as a representative stressor, we found that the transcript and protein levels of ECT8 rise in response to salt stress. The increased abundance of ECT8 protein results in the enhanced binding capability to m6A-modified mRNAs, thereby accelerating their degradation, especially those of negative regulators of salt stress responses. Our results demonstrated that ECT8 acts as an abiotic stress sensor, facilitating mRNA decay, which is vital for maintaining transcriptome homeostasis and enhancing stress tolerance in plants. Our findings not only advance the understanding of epitranscriptomic gene regulation but also offer potential applications for breeding more resilient crops in the face of rapidly changing environmental conditions.

The m6A reader SlYTH2 negatively regulates tomato fruit aroma by impeding the translation process.

N6-methyladenosine (m6A) is a fundamentally important RNA modification for gene regulation, whose function is achieved through m6A readers. However, whether and how m6A readers play regulatory roles during fruit ripening and quality formation remains unclear. Here, we characterized SlYTH2 as a tomato m6A reader protein and profiled the binding sites of SlYTH2 at the transcriptome-wide level. SlYTH2 undergoes liquid-liquid phase separation and promotes RNA-protein condensate formation. The target mRNAs of SlYTH2, namely m6A-modified SlHPL and SlCCD1B associated with volatile synthesis, are enriched in SlYTH2-induced condensates. Through polysome profiling assays and proteomic analysis, we demonstrate that knockout of SlYTH2 expedites the translation process of SlHPL and SlCCD1B, resulting in augmented production of aroma-associated volatiles. This aroma enrichment significantly increased consumer preferences for CRISPR-edited fruit over wild type. These findings shed light on the underlying mechanisms of m6A in plant RNA metabolism and provided a promising strategy to generate fruits that are more attractive to consumers.

Principles, functions, and biological implications of m6A in plants.

Over the past decade, N 6-methyladenosine (m6A) has emerged as a prevalent and dynamically regulated modification across the transcriptome; it has been reversibly installed, removed, and interpreted by specific binding proteins, and has played crucial roles in molecular and biological processes. Within this scope, we consolidate recent advancements of m6A research in plants regarding gene expression regulation, diverse physiologic and pathogenic processes, as well as crop trial implications, to guide discussions on challenges associated with and leveraging epitranscriptome editing for crop improvement.

OsALKBH9-mediated m6A demethylation regulates tapetal PCD and pollen exine accumulation in rice.

The N6-methyladenosine (m6A) mRNA modification is crucial for plant development and stress responses. In rice, the male sterility resulting from the deficiency of OsFIP37, a core component of m6A methyltransferase complex, emphasizes the significant role of m6A in male fertility. m6A is reversible and can be removed by m6A demethylases. However, whether mRNA m6A demethylase regulates male fertility in rice has remained unknown. Here, we identify the mRNA m6A demethylase OsALKBH9 and demonstrate its involvement in male fertility regulation. Knockout of OsALKBH9 causes male sterility, dependent on its m6A demethylation activity. Cytological analysis reveals defective tapetal programmed cell death (PCD) and excessive accumulation of microspores exine in Osalkbh9-1. Transcriptome analysis of anthers shows up-regulation of genes involved in tapetum development, sporopollenin synthesis, and transport pathways in Osalkbh9-1. Additionally, we demonstrate that OsALKBH9 demethylates the m6A modification in TDR and GAMYB transcripts, which affects the stability of these mRNAs and ultimately leads to excessive accumulation of pollen exine. Our findings highlight the precise control of mRNA m6A modification and reveal the pivotal roles played by OsALKBH9-mediated m6A demethylation in tapetal PCD and pollen exine accumulation in rice.

Methyltransferase ATMETTL5 writes m6A on 18S ribosomal RNA to regulate translation in Arabidopsis.

Aberrant RNA modifications can lead to dysregulated gene expression and impeded growth in plants. Ribosomal RNA (rRNA) constitutes a substantial portion of total RNA, while the precise functions and molecular mechanisms underlying rRNA modifications in plants remain largely elusive. Here, we elucidated the exclusive occurrence of the canonical RNA modification N6-methyladenosine (m6A) solely 18S rRNA, but not 25S rRNA. We identified a completely uncharacterized protein, ATMETTL5, as an Arabidopsis m6A methyltransferase responsible for installing m6A methylation at the 1771 site of the 18S rRNA. ATMETTL5 is ubiquitously expressed and localized in both nucleus and cytoplasm, mediating rRNA m6A methylation. Mechanistically, the loss of ATMETTL5-mediated methylation results in attenuated translation. Furthermore, we uncovered the role of ATMETTL5-mediated methylation in coordinating blue light-mediated hypocotyl growth by regulating the translation of blue light-related messenger RNAs (mRNAs), specifically HYH and PRR9. Our findings provide mechanistic insights into how rRNA modification regulates ribosome function in mRNA translation and the response to blue light, thereby advancing our understanding of the role of epigenetic modifications in precisely regulating mRNA translation in plants.

Dual-recognition colorimetric platform based on porous Au@Pt nanozymes for highly sensitive washing-free detection of Staphylococcus aureus.

A dual-recognition strategy is reported to construct a one-step washing and highly efficient signal-transduction tag system for high-sensitivity colorimetric detection of Staphylococcus aureus (S. aureus). The porous (gold core)@(platinum shell) nanozymes (Au@PtNEs) as the signal labels show highly efficient peroxidase mimetic activity and are robust. For the sake of simplicity the detection involved the use of a vancomycin-immobilized magnetic bead (MB) and aptamer-functionalized Au@PtNEs for dual-recognition detection in the presence of S. aureus. In addition, we designed a magnetic plate to fit the 96-well microplate to ensure consistent magnetic properties of each well, which can quickly remove unreacted Au@PtNEs and sample matrix while avoiding tedious washing steps. Subsequently, Au@PtNEs catalyze hydrogen peroxide (H2O2) to oxidize 3,3',5,5'-tetramethylbenzidine (TMB) generating a color signal. Finally, the developed Au@PtNEs-based dual-recognition washing-free colorimetric assay displayed a response in the range of S. aureus of 5 × 101-5 × 105 CFU/mL, and the detection limit was 40 CFU/mL within 1.5 h. In addition, S. aureus-fortified samples were analyzed to further evaluate the performance of the proposed method, which yielded average recoveries ranging from 93.66 to 112.44% and coefficients of variation (CVs) within the range 2.72-9.01%. These results furnish a novel horizon for the exploitation of a different mode of recognition and inexpensive enzyme-free assay platforms as an alternative to traditional enzyme-based immunoassays for the detection of other Gram-positive pathogenic bacteria.

RNA methylation in mammalian development and cancer.

Similar to epigenetic DNA and histone modifications, epitranscriptomic modifications (RNA modifications) have emerged as crucial regulators in temporal and spatial gene expression during eukaryotic development. To date, over 170 diverse types of chemical modifications have been identified upon RNA nucleobases. Some of these post-synthesized modifications can be reversibly installed, removed, and decoded by their specific cellular components and play critical roles in different biological processes. Accordingly, dysregulation of RNA modification effectors is tightly orchestrated with developmental processes. Here, we particularly focus on three well-studied RNA modifications, including N6-methyladenosine (m6A), 5-methylcytosine (m5C), and N1-methyladenosine (m1A), and summarize recent knowledge of underlying mechanisms and critical roles of these RNA modifications in stem cell fate determination, embryonic development, and cancer progression, providing a better understanding of the whole association between epitranscriptomic regulation and mammalian development.

An Improved Helferich Method for the α/ß-Stereoselective Synthesis of 4-Methylumbelliferyl Glycosides for the Detection of Microorganisms.

An improved Helferich method is presented. It involves the glycosylation of 4-methyl-umbelliferone with glycosyl acetates in the presence of boron trifluoride etherate combined with triethylamine, pyridine, or 4-dimethylaminopyridine under mild conditions, followed by deprotection to give fluorogenic 4-methylumbelliferyl glycoside substrates. Due to the use of base, the glycosylation reaction proceeds more easily, is uncommonly α- or ß-stereoselective, and affords the corresponding products in moderate to excellent yields (51%-94%) under appropriate conditions.

m6A readers ECT2/ECT3/ECT4 enhance mRNA stability through direct recruitment of the poly(A) binding proteins in Arabidopsis.

BACKGROUND: RNA N6-methyladenosine (m6A) modification is critical for plant growth and crop yield. m6A reader proteins can recognize m6A modifications to facilitate the functions of m6A in gene regulation. ECT2, ECT3, and ECT4 are m6A readers that are known to redundantly regulate trichome branching and leaf growth, but their molecular functions remain unclear. RESULTS: Here, we show that ECT2, ECT3, and ECT4 directly interact with each other in the cytoplasm and perform genetically redundant functions in abscisic acid (ABA) response regulation during seed germination and post-germination growth. We reveal that ECT2/ECT3/ECT4 promote the stabilization of their targeted m6A-modified mRNAs, but have no function in alternative polyadenylation and translation. We find that ECT2 directly interacts with the poly(A) binding proteins, PAB2 and PAB4, and maintains the stabilization of m6A-modified mRNAs. Disruption of ECT2/ECT3/ECT4 destabilizes mRNAs of ABA signaling-related genes, thereby promoting the accumulation of ABI5 and leading to ABA hypersensitivity. CONCLUSION: Our study reveals a unified functional model of m6A mediated by m6A readers in plants. In this model, ECT2/ECT3/ECT4 promote stabilization of their target mRNAs in the cytoplasm.

Cronobacter spp. Isolated from Quick-Frozen Foods in China: Incidence, Genetic Characteristics, and Antibiotic Resistance.

Cronobacter spp. are emerging foodborne pathogens that cause severe diseases. However, information on Cronobacter contamination in quick-frozen foods in China is limited. Therefore, we studied the prevalence, molecular characterization, and antimicrobial susceptibility of Cronobacter in 576 quick-frozen food samples collected from 39 cities in China. Cronobacter spp. were found in 18.75% (108/576) of the samples, and the contamination degree of the total positive samples was 5.82 MPN/g. The contamination level of frozen flour product samples was high (44.34%). Among 154 isolates, 109 were C. sakazakii, and the main serotype was C. sakazakii O1 (44/154). Additionally, 11 serotypes existed among four species. Eighty-five sequence types (STs), including 22 novel ones, were assigned, indicating a relatively high genetic diversity of the Cronobacter in this food type. Pathogenic ST148, ST7, and ST1 were the main STs in this study. ST4, epidemiologically related to neonatal meningitis, was also identified. All strains were sensitive to cefepime, tobramycin, ciprofloxacin, and imipenem, in which the resistance to cephalothin was the highest (64.94%).Two isolates exhibited multidrug resistance to five and seven antimicrobial agents, respectively. In conclusion, these findings suggest that the comparatively high contamination level of Cronobacter spp. in quick-frozen foods is a potential risk warranting public attention.

Evolutionary Divergence of the Novel Staphylococcal Species Staphylococcus argenteus.

Currently, invasive infections caused by Staphylococcus argenteus, which is a recently named staphylococcal species, are increasingly reported worldwide. However, only a few genomic studies of S. argenteus have offered comprehensive information regarding its genetic diversity, epidemiological characteristics, antimicrobial resistance genes (ARGs), virulence genes and other profiles. Here, we describe a comparative genomic analysis by population structure, pangenome, panmobilome, region-specific accessory genes confer an adaptive advantage in 153 S. argenteus strains which comprised 24 strains sequenced in this study and 129 strains whose genome sequences were available from GenBank. As a result, the population of S. argenteus comprised seven genetically distinct clades, including two major clades (C1 and C2), with distinct isolation source patterns. Pangenome analysis revealed that S. argenteus has an open pangenome composed of 7,319 genes and a core genome composed of 1,508 genes. We further determined the distributions of 75 virulence factors (VFs) and 30 known ARGs and identified at least four types of plasmids and 93 complete or partial putative prophages. It indicate that S. argenteus may show a similar level of pathogenicity to that of S. aureus. This study also provides insights into the evolutionary divergence of this pathogen, indicating that the geographical distribution was a potential driving force behind the evolutionary divergence of S. argenteus. The preferential horizontal acquisition of particular elements, such as staphylococcal cassette chromosome mec elements and plasmids, was observed in specific regions, revealing potential gene exchange between S. argenteus strains and local S. aureus strains. Moreover, multiple specific genes related to environmental adaptation were identified in strains isolated from East Asia. However, these findings may help promote our understanding of the evolutionary divergence of this bacterium at a high genetic resolution by providing insights into the epidemiology of S. argenteus and may help combat its spread.

[Evaluation on effects of chromogenic medium in rapid detection of Coliform and Escherichia coli].

OBJECTIVE: To evaluate the detective efficacy of Chromogenic Coliform and Escherichia Coli Agar (CCEA). METHODS: A new chromogenic medium CCEA prepared by Huankai laboratory was used to compare with a classical medium of violet red bile agar (VRBA), and other two Chromogenic media Agar I and Agar II by detecting separately 11 reference strains, thirteen sterile samples with Coliform or E.coli and other four samples, and the accordant rates of detection were observed. RESULTS: CCEA had the good selectivity. To seven kinds of quality strains in the resultant analysis, CCEA with VRBA and Agar I had not shown salience difference (P > 0.05), and CCEA with Agar II had significant difference (P < 0.05). CCEA showed more advantages than the Agar II. To thirteen sterile samples with Coliform or E.coli in resultant analysis, CCEA with Agar I and Agar II had shown no significant difference (P > 0.05), while CCEA with VRBA had significant difference (P < 0.05). CCEA might be more advantageous than the VRBA. In analysis of the four actual samples of Coliform, CCEA with VRBA, Agar I and Agar II showed no significant difference (P > 0.05). The accordant rates were 90%, 71.88%, 86.25% and 81.25% respectively, showing CCEA > Agar I > Agar II > VRBA. To two actual samples of E.coli in the resultant analysis, the CCEA with Agar I and Agar II had not shown significant difference (P > 0.05). The accordant rates were 100% respectively. CONCLUSIONS: The CCEA might be more advantageous than the VRBA, having the same efficacy as with Agar I and Agar II.

Synthesis of precipitating chromogenic/fluorogenic ß-glucosidase/ß-galactosidase substrates by a new method and their application in the visual detection of foodborne pathogenic bacteria.

We developed a new efficient method for the synthesis of important indoxyl glycoside substrates for ß-glucosidase and ß-galactosidase by using 1-acetylindol-3-ones as intermediates. This method was used to synthesise novel precipitating fluorogenic substrates for ß-glucosidase based on 2-(benzothiazol-2'-yl)-phenols. We also assessed the application of these substrates in the detection of foodborne pathogenic bacteria.