Transcriptome-wide mapping of N6-methyladenosine (m6A) at base resolution remains an issue, impeding our understanding of m6A roles at the nucleotide level. Here, we report a metabolic labeling ...method to detect mRNA m6A transcriptome-wide at base resolution, called ‘m6A-label-seq’. Human and mouse cells could be fed with a methionine analog, Se-allyl-l-selenohomocysteine, which substitutes the methyl group on the enzyme cofactor SAM with the allyl. Cellular RNAs could therefore be metabolically modified with N6-allyladenosine (a6A) at supposed m6A-generating adenosine sites. We pinpointed the mRNA a6A locations based on iodination-induced misincorporation at the opposite site in complementary DNA during reverse transcription. We identified a few thousand mRNA m6A sites in human HeLa, HEK293T and mouse H2.35 cells, carried out a parallel comparison of m6A-label-seq with available m6A sequencing methods, and validated selected sites by an orthogonal method. This method offers advantages in detecting clustered m6A sites and holds promise to locate nuclear nascent RNA m6A modifications.The authors developed a metabolic labeling method via incorporation of allyl-SAM analogs to profile transcriptome-wide m6A at base resolution, which enables identification of m6A motifs and clustered m6A sites.
Abstract
Faithful genome integrity maintenance plays an essential role in cell survival. Here, we identify the RNA demethylase ALKBH5 as a key regulator that protects cells from DNA damage and ...apoptosis during reactive oxygen species (ROS)-induced stress. We find that ROS significantly induces global mRNA N6-methyladenosine (m6A) levels by modulating ALKBH5 post-translational modifications (PTMs), leading to the rapid and efficient induction of thousands of genes involved in a variety of biological processes including DNA damage repair. Mechanistically, ROS promotes ALKBH5 SUMOylation through activating ERK/JNK signaling, leading to inhibition of ALKBH5 m6A demethylase activity by blocking substrate accessibility. Moreover, ERK/JNK/ALKBH5-PTMs/m6A axis is activated by ROS in hematopoietic stem/progenitor cells (HSPCs) in vivo in mice, suggesting a physiological role of this molecular pathway in the maintenance of genome stability in HSPCs. Together, our study uncovers a molecular mechanism involving ALKBH5 PTMs and increased mRNA m6A levels that protect genomic integrity of cells in response to ROS.
Abstract
Here we show that FTO as an
N
6
-methyladenosine (m
6
A) RNA demethylase is degraded by selective autophagy, which is impaired by low-level arsenic exposure to promote tumorigenesis. We ...found that in arsenic-associated human skin lesions, FTO is upregulated, while m
6
A RNA methylation is downregulated. In keratinocytes, chronic relevant low-level arsenic exposure upregulated FTO, downregulated m
6
A RNA methylation, and induced malignant transformation and tumorigenesis.
FTO
deletion inhibited arsenic-induced tumorigenesis. Moreover, in mice, epidermis-specific
FTO
deletion prevented skin tumorigenesis induced by arsenic and UVB irradiation. Targeting FTO genetically or pharmacologically inhibits the tumorigenicity of arsenic-transformed tumor cells. We identified
NEDD4L
as the m
6
A-modified gene target of FTO. Finally, arsenic stabilizes FTO protein through inhibiting p62-mediated selective autophagy. FTO upregulation can in turn inhibit autophagy, leading to a positive feedback loop to maintain FTO accumulation. Our study reveals FTO-mediated dysregulation of mRNA m
6
A methylation as an epitranscriptomic mechanism to promote arsenic tumorigenicity.
TET enzymes convert 5-methylcytosine to 5-hydroxymethylcytosine and higher oxidized derivatives. TETs stably associate with and are post-translationally modified by the nutrient-sensing enzyme OGT, ...suggesting a connection between metabolism and the epigenome. Here, we show for the first time that modification by OGT enhances TET1 activity in vitro. We identify a TET1 domain that is necessary and sufficient for binding to OGT and report a point mutation that disrupts the TET1-OGT interaction. We show that this interaction is necessary for TET1 to rescue hematopoetic stem cell production in tet mutant zebrafish embryos, suggesting that OGT promotes TET1's function during development. Finally, we show that disrupting the TET1-OGT interaction in mouse embryonic stem cells changes the abundance of TET2 and 5-methylcytosine, which is accompanied by alterations in gene expression. These results link metabolism and epigenetic control, which may be relevant to the developmental and disease processes regulated by these two enzymes.
In this study, changes in the microbial communities and the metabolic pathway involved in anaerobic digestion were examined as the temperature was changed from 9 to 55 °C. The results from six batch ...biogas reactors, which were designed to ferment at 9, 15, 21, 35, 45 and 55 °C, showed that: 1) The components of the microbial communities and metabolic pathways were found to be very similar but the metabolic activities increased significantly when the temperature grew from 15 to 35 °C. 2) Hydrogenotrophic metabolism was almost exclusively observed when the temperature was lowered to 9 °C, whereas greater bacterial diversity involving hydrolisis, acidogenesis, and acetogenesis was observed when the temperature was raised to 45 °C. Finally, these dominant bacteria involved in hydrolisis, acidogenesis, and acetogenesis decreased in number when the temperature was raised to 55 °C and acetotrophic-type metabolism became the major metabolic pathway. That means the turning point for microbial community composition and metabolic pathway from mesophilic to thermophilic was at 45 °C and the metabolic pathway at 9 °C was somewhat different to those observed at 15–35 °C.
•Microbial community structures were connected to their metabolic pathways.•Microbial communities were similar but metabolic activity increased with temperature grew.•Turning point for metabolic pathway from mesophilic to thermophilic was at 45 °C.•Key microorganisms from hydrolysis to methanogenesis comprised not more than 20 species.
The genus Ophiocordyceps, which includes Ophiocordyceps sinensis, has been demonstrated to be one of the most valuable medicinal taxa. The low rate of larval infection and slow development that ...characterize the cultivation of this genus should be urgently addressed. To identify potential bioinoculants that stimulate the growth of Ophiocordyceps, O. highlandensis was selected as a model system, and a total of 72 samples were collected to systematically compare the microbial communities present during fruiting body development. By applying high-throughput 16S and ITS2 amplicon sequencing technology, the bacterial and fungal communities were identified in O. highlandensis and its surrounding soil, and the functional dynamics of the bacteria were explored. The results indicate that the most abundant bacteria across all the samples from O. highlandensis were Proteobacteria, Firmicutes and Bacteroidetes, while members of Ascomycota were detected among the fungi. The pathways enriched in the developmental stages were associated with carbohydrate degradation, nucleotides and pyridoxal biosynthesis, and the TCA cycle. Compared with that in the fungal community, an unexpectedly high taxonomic and functional fluctuation was discovered in the bacterial community during the maturation of O. highlandensis. Furthermore, bipartite network analysis identified four potential supercore OTUs associated with O. highlandensis growth. All the findings of this study suggest unexpectedly high taxonomic and functional fluctuations in the bacterial community of O. highlandensis during its maturation. O. highlandensis may recruit different endogenous bacteria across its life cycle to enhance growth and support rapid infection. These results may facilitate Ophiocordyceps cultivation and improve the development of strategies for the identification of potential bioinoculant resources.
Bacteriophages function as a regulator of host communities and metabolism. Many phages have been isolated and sequenced in environments such as the ocean, but very little is known about hypersaline ...environments. Phages infecting members of the genus
remain poorly understood, and no
phage genome has been reported. In this study, a halovirus infecting
sp. F3, YPCBV-1, was isolated from Yipinglang salt mine. YPCBV-1 could only infect host strain F3 with burst size of 6.3 PFU/cell. It could produce progeny in 5%-20% (w/v) NaCl with an optimal concentration of 10% (w/v), but the optimal adsorption NaCl concentration was 5%-8% (w/v). YPCBV-1 is sensitive to pure water and depends on NaCl or KCl solutions to survive. YPCBV-1 stability increased with increasing salinity but decreased in NaCl saturated solutions, and it has a broader salinity adaptation than the host. YPCBV-1 has a double-stranded DNA of 36,002 bp with a G + C content of 67.09% and contains a total of 55 predicted ORFs and no tRNA genes. Phylogenetic analysis and genomic network analysis suggested that YPCBV-1 is a novel Mu-like phage under the class Caudoviricetes. Auxiliary metabolic gene, SUMF1/EgtB/PvdO family non-heme iron enzyme, with possible roles in antioxidant was found in YPCBV-1. Moreover, DGR-associated genes were predicted in YPCBV-1 genome, which potentially produce hypervariable phage tail fiber. These findings shed light on the halovirus-host interaction in hypersaline environments.
Fragaria nilgerrensis is a wild strawberry species widely distributed in southwest China and has strong ecological adaptability. Akihime ( F. × ananassa Duch. cv. Akihime) is one of the main ...cultivated strawberry varieties in China and is prone to infection with a variety of diseases. In this study, high-throughput sequencing was used to analyze and compare the soil and root microbiomes of F. nilgerrensis and Akihime. Results indicate that the wild species F. nilgerrensis showed higher microbial diversity in nonrhizosphere soil and rhizosphere soil and possessed a more complex microbial network structure compared with the cultivated variety Akihime. Genera such as Bradyrhizobium and Anaeromyxobacter , which are associated with nitrogen fixation and ammonification, and Conexibacter , which is associated with ecological toxicity resistance, exhibited higher relative abundances in the rhizosphere and nonrhizosphere soil samples of F. nilgerrensis compared with those of Akihime. Meanwhile, the ammonia-oxidizing archaea Candidatus Nitrososphaera and Candidatus Nitrocosmicus showed the opposite tendencies. We also found that the relative abundances of potential pathogenic genera and biocontrol bacteria in the Akihime samples were higher than those in the F. nilgerrensis samples. The relative abundances of Blastococcus, Nocardioides, Solirubrobacter, and Gemmatimonas , which are related to pesticide degradation, and genus Var iovorax , which is associated with root growth regulation, were also significantly higher in the Akihime samples than in the F. nilgerrensis samples. Moreover, the root endophytic microbiomes of both strawberry species, especially the wild F. nilgerrensis , were mainly composed of potential biocontrol and beneficial bacteria, making them important sources for the isolation of these bacteria. This study is the first to compare the differences in nonrhizosphere and rhizosphere soils and root endogenous microorganisms between wild and cultivated strawberries. The findings have great value for the research of microbiomes, disease control, and germplasm innovation of strawberry.
The aim of this study was to compare the microbial community structure and diversity in powdery mildew-infected and noninfected strawberry plant rhizosphere soils in the greenhouse based on ...variations in the 16S rRNA gene V3–V4 and fungal ITS2 regions by Illumina amplicon sequencing. Powdery mildew infection reduced the number of operational taxonomic units (OTUs) and prokaryotic and fungal community richness/diversity indexes in the rhizosphere soils compared with those in healthy plant soils. Furthermore, 3543 prokaryotic and 581 fungal OTUs were obtained at the 97% similarity level. Proteobacteria, Actinobacteria, Bacteroidetes, Acidobacteria, and Chloroflexi were the dominant bacterial phyla; Woesearchaeota_DHVEG-6, Bathyarchaeota, and Thaumarchaeota were the dominant archaea; and Ascomycota, Basidiomycota, unclassified_fungi, and Zygomycota were the dominant fungal phyla. Their proportions differed significantly among samples.
Wolbachia
,
Devosia
,
Pseudolabrys
,
Streptomyces
, and
Rhizomicrobium
were the most abundant bacterial genera; their proportions differed significantly among samples. Most
Pseudomonas
,
Streptomyces
, and ‘norank’ group members might be potential antagonistic microorganisms of powdery mildew pathogens, and
Wolbachia
and
Rickettsia
might be pathogen-transmitting vectors.
Microascus
,
Clitopilus
, and
Ciliophora
were the dominant fungi, and their community structures and abundances significantly differed among samples.
Microascus
,
Talaromyces
,
Zopfiella
, and
Cryptococcus
were relatively more abundant in the powdery mildew-infected strawberry plant rhizosphere soils.
Fusarium
,
Trichoderma
,
Clitopilus
, and ‘unclassified’ group members may be potential antagonistic populations. The results suggested that powdery mildew-infected strawberry fruits and plants cannot be consumed. This report is the first study to illustrate differences in the rhizosphere soil prokaryotic and fungal communities between powdery mildew-infected and noninfected strawberry plants in a greenhouse.
With continuous progress in science and technology, a large amount of data are produced in all fields of the world at anytime and anywhere. These data are unmarked and lack marking information, while ...manual marking is time-consuming and laborious. Herein, this paper introduces a distributed semi-supervised labeling framework. This framework addresses the issue of missing data by proposing an attribute-filling method based on subspace learning. Furthermore, this paper presents a distributed semi-supervised learning strategy that trains sub-models (private models) within each sub-system. Finally, this paper develops a distributed graph convolutional neural network fusion technique with enhanced interpretability grounded on the attention mechanism. This paper assigns weights of importance to the edges of each layer in the graph neural network based on sub-models and public data, thereby enabling distributed and interpretable graph convolutional attention. Extensive experimentation using public datasets demonstrates the superiority of the proposed scheme over other state-of-the-art baselines, achieving a reduction in loss of 50% compared to the original approach.
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