N6-methyladenosine (m6A) profiling in 60 YRI LCLs
Ontology highlight
ABSTRACT:
ORGANISM(S): Homo sapiens
PROVIDER: GSE125377 | GEO | 2020/05/04
REPOSITORIES: GEO
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Project description:N6-methyladenosine (m6A) profiling in 60 YRI LCLs
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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
Project description:m6A-seq of undifferentiated and differentiated mouse embryonic stem cell m6A-mRNA library for undifferentiated and differentiated mouse embryonic stem cell each having one biological replicate were generated using HiSeq2000 v3 flowcell (Illumina) and sequenced for 100 bases with separate 7 base indexing read in a single lane.
2015-12-31 | E-GEOD-44086 | biostudies-arrayexpress
Project description:N6-methyladenosine (m6A) is the most abundant internal modification in the messenger RNA (mRNA) of all higher eukaryotes. This modification has been shown to be reversible in mammals; it is installed by a methyltransferase heterodimer complex of METTL3 and METTL14 bound with WTAP, and reversed by iron(II)- and α-ketoglutarate-dependent demethylases FTO and ALKBH5. This modification exhibits significant functional roles in various biological processes. The m6A modification as a RNA mark is recognized by reader proteins, such as YTH domain family proteins and HNRNPA2B1; m6A can also act as a structure switch to affect RNA-protein interactions for biological regulation. In Arabidopsis thaliana, the methyltransferase subunit MTA (the plant orthologue of human METTL3, encoded by At4g10760) was well characterized and FIP37 (the plant orthologue of human WTAP) was first identified as the interacting partner of MTA. Here we report the discovery and characterization of reversible m6A methylation mediated by AtALKBH10B (encoded by At4g02940) in A. thaliana, and noticeable roles of this RNA demethylase in affecting plant development and floral transition. Our findings reveal potential broad functions of reversible mRNA methylation in plants. m6A peaks were identified from wild type Columbia-0 and atalkbh10b-1 mutant in two biological replicates
2016-03-24 | E-GEOD-79523 | biostudies-arrayexpress
Project description:We show that N6-methyladenosine (m6A), the most abundant internal modification in mRNA/lncRNA with still poorly characterized function, alters RNA structure to facilitate the access of RBM for heterogeneous nuclear ribonucleoprotein C (hnRNP C). We term this mechanism m6A-switch. Through combining PAR-CLIP with Me-RIP, we identify 39,060 m6A-switches among hnRNP C binding sites transcriptome-wide. We show that m6A-methyltransferases METTL3 or METTL14 knockdown decreases hnRNP C binding at 16,582 m6A-switches. Taken together, 2,798 m6A-switches of high confidence are identified to mediate RNA-hnRNP C interactions and affect diverse biological processes including cell cycle regulation. These findings reveal the biological importance of m6A and provide insights into the sophisticated regulation of RNA-RBP interactions through m6A-induced RNA structural remodeling. Measure the m6A methylated hnRNP C binding sites transcriptome-wide by PARCLIP-MeRIP; measure the differential hnRNP C occupancies upon METTL3/METTL14 knockdown by PAR-CLIP; measure RNA abundance and splicing level changes upon HNRNPC, METTL3 and METTL14 knockdown
2015-02-27 | E-GEOD-56010 | biostudies-arrayexpress
Project description:Phenotypic differences within populations and between closely related species are often driven by variation and evolution of gene expression. However, most analyses have focused on the effects of genomic variation at cis-regulatory elements such as promoters and enhancers that control transcriptional activity, and little is understood about the influence of post-transcriptional processes on transcript evolution. Post-transcriptional modification of RNA by N6-methyladenosine (m6A) has been shown to be widespread throughout the transcriptome, and this reversible mark can affect transcript stability and translation dynamics. Here we analyze m6A mRNA modifications in lymphoblastoid cell lines (LCLs) from human, chimpanzee and rhesus, and we identify patterns of m6A evolution among species. We find that m6A evolution occurs in parallel with evolution of consensus RNA sequence motifs known to be associated with the enzymatic complexes that regulate m6A dynamics, and expression evolution of m6A-modified genes occurs in a parallel evolutionary pattern with m6A evolution. Further, genes modified by evolved m6A in humans are significantly enriched in transcriptional regulatory processes and disease related pathways, suggesting an important role in human biology for the evolution of m6A modification.
2016-10-14 | GSE70299 | GEO
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
Project description:Noncoding variants play a central role in the genetics of complex traits, but we still lack a full description of the main molecular pathways through which they act. Here we used molecular data to quantify the contribution of cis-acting genetic effects at each major stage of gene regulation from chromatin to proteins, within a population sample of Yoruba lymphoblastoid cell lines (LCLs). We performed 4sU metabolic labeled transcripts in 65 YRI LCLs to identify genetic variants that affect transcription rates. As expected, we found an important contribution of genetic variation via chromatin, contributing â¼65% of eQTLs (expression Quantitative Trait Loci). The remaining eQTLs, which are not asso- ciated with chromatin-level variation, are highly enriched in transcribed regions, and hence may affect expression through co- or post-transcriptional processes. International HapMap lymphoblastoid cell lines (LCLs) derived from YRI (Yoruba in Ibadan, Nigeria); We adapted the 4sU labelling method from (PMID 21516085). Briefly, cell cultures were grown to log phase in volumes sufficient to yield about 300 ng of 4sU-labeled RNA. Cells were incubated with 4sU for the required length of time (0, 30, or 60 minutes), then washed, pelleted, and frozen. Total RNA was extracted, and 4sU-labeled RNA was separated from total RNA using a bead-based biotin-streptavidin purification protocol. We sequenced metabolic labeled transcripts in 65 YRI LCLs 30 minutes and 60 minutes after incubation.
2016-03-29 | E-GEOD-75220 | biostudies-arrayexpress
Project description:Noncoding variants play a central role in the genetics of complex traits, but we still lack a full description of the main molecular pathways through which they act. Here we used molecular data to quantify the contribution of cis-acting genetic effects at each major stage of gene regulation from chromatin to proteins, within a population sample of Yoruba lymphoblastoid cell lines (LCLs). We performed 4sU metabolic labeled transcripts in 65 YRI LCLs to identify genetic variants that affect transcription rates. As expected, we found an important contribution of genetic variation via chromatin, contributing ∼65% of eQTLs (expression Quantitative Trait Loci). The remaining eQTLs, which are not asso- ciated with chromatin-level variation, are highly enriched in transcribed regions, and hence may affect expression through co- or post-transcriptional processes.
2016-03-29 | GSE75220 | GEO
Project description:Systematically evaluate the global effect of dysregulated m6A regulators on the m6A epitranscriptome in patient tumors, and identify prognostic m6A markers by linking with patients' clinicopathological and outcome information.
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