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N6-methyladenosine (m6A) RIP-seq in SK-Hep-1 cells

ABSTRACT: To explore the m6a modification in SK-Hep-1 cells, we performed RNA immunoprecipitation with an antibody against m6a (SYSY), and sequencing was performed using Illumina NovaSeq 6000

ORGANISM(S): Homo sapiens

PROVIDER: GSE243538 | GEO | 2023/09/20

REPOSITORIES: GEO

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N6-methyladenosine (m6A) RIP-seq in SK-Hep-1 cells

Project description:N6-methyladenosine (m6A) RIP-seq in SK-Hep-1 cells

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Expression analysis of sk-hep-1(sk) cell and sk-hep-1-sh EZH2(sks) cell

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Genome-wide maps of H3K27me3 modification sites in sk-hep-1,sk-hep-1-shEZH2 and LO2 cells.

Project description:We report the application of chromatin immunoprecipitation coupled with high-throughput sequencing (ChIP-seq) for high-throughput profiling of H3K27me3 modifications in sk-hep-1(sk), sk-hep-1-shEZH2 (sks) and LO2 cells. Following standard ChIP procedure, 10 ng of each DNA sample was used for Illumina sequencing. Statistically significant ChIP-enriched regions (peaks) were identified by sk vs sks (p-value threshold of 10-2) or sk vs LO2 (p-value threshold of 10-3). Enriched peaks were annotated by the nearest gene using the newest UCSC RefSeq database. We identify genes modified by EZH2-mediated H3K27me3. This study provides insights into these down stream genes modified by EZH2 regulated H3K27me3 in HCC cell lines.

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H3K27ac ChIP-seq of SK-Hep-1, Huh7, SNU387, and SNU182 cells

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Transcriptome-wide N6-methyladenosine modification profiling in patients with recurrent implantation failure (RIF)

Project description:Purpose:using m6A modified RNA immunoprecipitation sequence (m6A-seq), to establish the m6A methylation transcription profiles in recurrent implantation failure (RIF). Methods: GenSeq® Low Input Whole RNA Library Prep Kit (GenSeq, Inc.) was used to construct RNA libraries for IP and input samples by following the manufacturer's instructions. The library quality was evaluated using Agilent 2100 bioanalyzer and then sequenced in a NovaSeq platform (Illumina 6000). Result:using m6A modified RNA immunoprecipitation sequence (m6A-seq), methylated sites in mRNA,lncRNA and circRNA are identified. Conclusions: Our study revealed the m6A methylation landscape by MERIP sequencing.

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Transcriptome-wide N6-Methyladenosine (m6A) modifications in Anopheles sinensis

Project description:We profile transcriptome-wide m6A in female and male Anopheles sinensis and reveal that m6A is also a highly conserved modification of mRNA in mosquitoes were generated by deep sequencing, in triplicate, using illumina Novaseq™ 6000. Distinct from mammals and yeast, but similar to Arabidopsis thaliana, m6A in An. sinensis is enriched not only around the stop codon and within 3’-untranslated regions, but also around the start codon and 5’-UTR. Gene ontology analysis indicates that the unique distribution pattern of m6A in An. sinensis is associated with mosquito sex-specific pathways. In addition, the positive correlation between m6A deposition and mRNA abundance indicates that m6A can play roles in regulating gene expression in mosquitoes. Our study proposed a transcriptional regulatory network of m6A in An. sinensis, which may provide a new clue for the control of this disease-transmitting vector.

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ATAC-seq for SK-HEP-1 cell line with or without sirolimus treatment

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N6-methyladenosine Modulates Messenger RNA Translation Efficiency

Project description:N6-methyladenosine (m6A) is the most abundant internal messenger (mRNA) modification in mammalian mRNA. This modification is reversible and non-stoichiometric, which potentially adds an additional layer of variety and dynamic control of mRNA metabolism. The m6A-modified mRNA can be selectively recognized by the YTH family “reader” proteins. The preferential binding of m6A-containing mRNA by YTHDF2 is known to reduce the stability of the target transcripts; however, the exact effects of m6A on translation has yet to be elucidated. Here we show that another m6A reader protein, YTHDF1, promotes ribosome loading of its target transcripts. YTHDF1 forms a complex with translation initiation factors to elevate the translation efficiency of its bound mRNA. In a unified mechanism of translation control through m6A, the YTHDF2-mediated decay controls the lifetime of target transcripts; whereas, the YTHDF1-based translation promotion increases the translation efficiency to ensure effective protein production from relatively short-lived transcripts that are marked by m6A. PAR-CLIP and RIP was used to identify YTHDF1 binding sites followed by ribosome profling and RNA seq to assess the consequences of YTHDF1 siRNA knock-down

2015-06-09 | E-GEOD-63591 | biostudies-arrayexpress

Dynamic N6-methyladenosine Epitranscriptomic Landscape in Lung Adenocarcinoma

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Genome-wide identification of N6-methyladenosine by microarray

Project description:Although internal PolyA RNA modification N6-methyladenosine (m6A) plays essential roles in diverse biological processes, technology to detect precise m6A sites at transcriptome-wide scale is lacking. Here, we discovered that m6A interferes A (Adenine) – U (Uracil) or A-T (Thymidine) pairing. Based on differential hybridization between methylated vs. unmethylated RNAs to a DNA probe, we developed tiling microarray to pinpoint m6A sites in mouse transcriptome. We validated some of the identified sites and provided evidence to suggest that one functional mechanism of m6A is to block small RNA targeting to methylated mRNA. We designed a custom tiling array with to examine the precise location of m6A within meRIP-seq peaks from mouse embryonic stem cells determined in our previous publication (Wang et al., 2014). Each custom two-channel Agilent tiling array harbors 947,952 probes. Each probe is 25 nucleotides (nt), and any two adjacent probes in the genomic coordinate overlap each other by 19 nt. The Cy5 or red channel corresponds to Mettl14 knockout (M14) or DZA mutant mESC cell line, and Cy3 or green channel is associated with wild type cell line treated with scramble hairpin (SCR). Thus, in principle a higher Cy5/Cy3 signal for each probe reflects an increased hybridization to the oligonucleotide due to de-methylation of a particular RNA molecule in M14 or DZA condition relative to the SCR control. Moreover, we employed additional arrays with both channels dedicated for M14 as an external control for technical difference between the Cy5 and Cy3 dye (details below). For each comparison, we have three biological replicates, and therefore there are 9 tiling arrays in total (i.e., 3 arrays for M14 vs SCR, 3 arrays for DZA vs SCR, and 3 arrays for M14 vs M14).

2015-09-08 | E-GEOD-62530 | biostudies-arrayexpress

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