Single-nucleus multiomic mapping of m6A methylome and transcriptome in native populations of cells with sn-m6A-CT
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PROVIDER: PRJNA898293 | ENA |
REPOSITORIES: ENA
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Project description:N6-methyladenosine (m6A) RNA modification plays important roles in the governance of gene expression and is temporally regulated in different cell states. In contrast to global m6A profiling in bulk sequencing, technologies for analyzing m6A heterogeneity across single-cells are not extensively established. Here, we report single-nucleus m6A CUT-Tag (sn-m6A-CT) for simultaneous profiling of m6A methylome and transcriptome within a single nucleus. m6A-CT is capable of enriching m6A-marked RNA molecules in situ, without isolating RNAs from cells, in a highly efficient manner. We adapted m6A-CT to the droplet-based single-cell omics platform and demonstrated high throughput performance in analyzing nuclei isolated from thousands of cells from various cell types. We show that sn-m6A-CT profiling is sufficient to determine cell identity and allows the generation of cell-type-specific m6A methylome landscapes from heterogeneous populations. Further, we exemplified the robust deconvolution capabilities in m6A heterogeneities with stem cells in multiple distinct pluripotency states. These indicate that sn-m6A-CT provides additional dimensions to multimodal datasets and new insights into epitranscriptomic landscape in defining cell fate identity and states.
2023-09-05 | GSE217254 | GEO
Project description:N6-methyladenosine (m6A) RNA modification plays important roles in the governance of gene expression and is temporally regulated in different cell states. In contrast to global m6A profiling in bulk sequencing, technologies for analyzing m6A heterogeneity across single-cells are not extensively established. Here, we report single-nucleus m6A CUT-Tag (sn-m6A-CT) for simultaneous profiling of m6A methylome and transcriptome within a single nucleus. m6A-CT is capable of enriching m6A-marked RNA molecules in situ, without isolating RNAs from cells, in a highly efficient manner. We adapted m6A-CT to the droplet-based single-cell omics platform and demonstrated high throughput performance in analyzing nuclei isolated from thousands of cells from various cell types. We show that sn-m6A-CT profiling is sufficient to determine cell identity and allows the generation of cell-type-specific m6A methylome landscapes from heterogeneous populations. Further, we exemplified the robust deconvolution capabilities in m6A heterogeneities with stem cells in multiple distinct pluripotency states. These indicate that sn-m6A-CT provides additional dimensions to multimodal datasets and new insights into epitranscriptomic landscape in defining cell fate identity and states.
2023-09-05 | GSE217255 | GEO
Project description:Single-nucleus multiomic mapping of m6A methylome and transcriptome in native populations of cells with sn-m6A-CT [Bulk CUT&Tag]
| PRJNA898301 | ENA
Project description:Single-nucleus multiomic mapping of m6A methylome and transcriptome in native populations of cells with sn-m6A-CT [scRNA-seq, scCUT&Tag]
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2023-09-05 | GSE217256 | GEO
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Project description:Despite effective antiretroviral therapy at reducing HIV-1 viral loads to undetectable levels, the presence of latently infected CD4+ T cells poses a major barrier to HIV-1 cure. N6-methyladenosine (m6A) modification of viral and cellular RNA has a functional role in regulating HIV-1 infection. m6A modification of HIV-1 RNA can affect its stability, translation, and splicing in cells and suppresses type-I interferon induction in macrophages. However, the function of m6A modification in regulating HIV-1 latency reactivation remains unknown. We used the Jurkat T cell line-derived HIV-1 latency model (J-Lat cells) to investigate changes in m6A levels of cellular RNA in response to latency reversal. We observed a significant increase in m6A levels of total cellular RNA upon reactivation of latent HIV-1 in J-Lat cells. This increase in m6A levels was transient and returned to steady-state levels despite continued high levels of viral gene expression in reactivated cells compared to control cells. Upregulation of m6A levels occurred without significant changes in the protein expression of m6A writers or erasers that add or remove m6A, respectively. Knockdown of m6A writers in J-Lat cells significantly reduced HIV-1 reactivation. Treatment of a m6A writer inhibitor reduced cellular RNA m6A levels along with reduction in HIV-1 reactivation. Furthermore, using m6A-specific sequencing, we identified cellular RNAs that are differentially m6A-modified during HIV-1 reactivation in J-Lat cells. Knockdown of identified m6A-modified RNA validate these results with established primary CD4+ T-cell models of HIV-1 latency. These results showed the importance of m6A RNA modification in HIV-1 latency reversal.
2024-10-01 | GSE273614 | GEO
Project description:N6-methyladenosine (m6A) is a widespread reversible chemical modification of RNAs, implicated in many aspects of RNA metabolism. Little quantitative information exists as to either how many transcript copies of particular genes are m6A modified (âm6A levelsâ), or the relationship of m6A modification(s) to alternative RNA isoforms. To deconvolute the m6A epitranscriptome, we developed m6A level and isoform-characterization sequencing (m6A-LAIC-seq). We found that cells exhibit a broad range of non-stoichiometric m6A levels with cell type specificity. At the level of isoform characterization, we discovered widespread differences in use of tandem alternative polyadenylation (APA) sites by methylated and nonmethylated transcript isoforms of individual genes. Strikingly, there is a strong bias for methylated transcripts to be coupled with proximal APA sites, resulting in shortened 3â untranslated regions (3â-UTRs), while nonmethylated transcript isoforms tend to use distal APA sites. m6A-LAIC-seq yields a new perspective on transcriptome complexity and links APA usage to m6A modifications. m6A-LAIC-seq of H1-ESC and GM12878 cell lines, each cell line has two replicates
2016-06-24 | E-GEOD-66086 | biostudies-arrayexpress
Project description:Here we report a metabolic labeling method to map mRNA N6-methyladenosine (m6A) modification transcriptome-wide at base resolution, termed m6A-label-seq. The cells were fed with Se-allyl-L-selenohomocysteine, an analog of methoine, which serves as the precursor of methylation enzyme cofactor, so that cellular RNAs were continuously deposited with N6-allyladenosine (a6A) at supposed m6A sites. We enriched a6A-containing mRNAs and sequenced their a6A sites which are identical to m6A sites, based on iodination-induced misincorporation during reverse transcription.
2020-04-21 | GSE131316 | GEO
Project description:To find the m6A methylation targets of ABRO1, we performed m6A methylated RNA immunoprecipitation sequencing (MeRIP-seq) in ABRO1 Knockout or Wild type (WT) mice heart. The analysis of the distribution of m6A peak density in mRNA transcripts showed that m6A peaks are mainly found in coding sequences (CDS) and a considerable amount of m6A peaks enriched around start codon and stop codon regions in mRNA transcripts from ABRO1 KO and WT hearts. Among the total RNA transcripts with m6A sites, more than half (59.4%) of mRNA transcripts contain ≤ 2 m6A peaks, 27.4% mRNA transcripts comprise 3 to 5 m6A peaks and more than 5 m6A peaks exist in 13.2% of mRNA transcripts. MeRIP-seq analysis results from ABRO1 deleted hearts showed that a total of 3444 m6A peaks were upregulated and 4631 m6A were downregulated compared to WT hearts.
2023-12-01 | GSE188518 | GEO