High throughput characterization of the m6A demethylase FTO by CLIP and RNAseq
Ontology highlight
ABSTRACT:
ORGANISM(S): Homo sapiens
PROVIDER: GSE79577 | GEO | 2017/08/25
SECONDARY ACCESSION(S): PRJNA316214
REPOSITORIES: GEO
Similar Datasets
Project description:To gain insight into FTO function, we knocked down and overexpressed FTO in HEK293 cells.Genetrail analyses of expression profiles pointed to the RNA splicing and processing machinery. Intriguingly, using immunocytochemistry and confocal laser scanning microscopy, we observed strong enrichment of FTO in nuclear speckles and - to a lesser extent - in nucleoli, but not in other known nuclear bodies. We also studied RNA samples of Fto knockout and wild type mice with regard to content of methylated and unmethylated nucleosidesand observed that ratios of modified and unmodified uracil and adenine were different depending on the presence of FTO. Taken together, our data suggest that FTO is involved in RNA processing and modification. We used microarrays to investigate global gene expression changes depending on the level of FTO We compared FTO overexpressing and FTO depleted cells to cells with endogeneus level of FTO to determine global gene expression changes.
2012-11-13 | E-GEOD-33870 | biostudies-arrayexpress
Project description:The fat mass and obesity-associated (FTO) protein is a well-characterized demethylase that removes N6-methyladenosine (m6A) from animal mRNAs. However, it is unclear yet how the demethylation operates in living cells. In this study, we applied genome-wide approaches to study how FTO finds its demethylation targets in human cells. We overexpressed FTO in human HeLa cells, demonstrating that FTO effectively removes m6A from the RRACH motif enriched in the 3’UTR regions and leaves m6A at other motifs unaffected. RRACH elements are clearly enriched at FTO binding sites; however, m6A has a lower tendency to be removed from the FTO-bound RRACH. Taken together with the experimental validaton results, we propose a model in which FTO effectivly recognizes m6A-containing RRACH motifs in RNAs, leading to a faster demethylation and dissociation kinetic than the association, and consequently undectable FTO-mRNA association. However, when FTO binds to the non-m6A RRACH motif, the binding has a lower dissociation kinetics, yielding the detectable FTO binding signals which would enable FTO to have roles in regulating other mRNA processing events.
2020-03-22 | GSE101953 | GEO
Project description:The RNA modification N6-methyladenosine (m6A) plays a key role in the life cycles of several RNA viruses. Whether this applies to SARS-CoV-2 and whether m6A affects the outcome of COVID-19 disease is still poorly explored. Here we report that the RNA demethylase FTO strongly affects both m6A marking of SARS-CoV-2 and COVID-19 severity. By m6A profiling of SARS-CoV-2, we confirmed in infected cultured cells and showed for the first time in vivo in hamsters that the regions encoding TRS_L and the nucleocapsid protein are multiply marked by m6A, preferentially within RRACH motifs that are specific to β-coronaviruses and well conserved across SARS-CoV-2 variants. In cells, downregulation of the m6A demethylase FTO, occurring upon SARS-CoV-2 infection, increased m6A marking of SARS-CoV-2 RNA and slightly promoted viral replication. In COVID-19 patients, a negative correlation was found between FTO expression and both SARS-CoV-2 expression and disease severity. FTO emerged as a classifier of disease severity and hence a potential stratifier of COVID-19 patients.
2023-04-26 | GSE201626 | GEO
Project description:The fat mass and obesity-associated (FTO) protein is a well-characterized demethylase that removes N6-methyladenosine (m6A) from animal mRNAs. However, it is unclear yet how the demethylation operates in living cells. In this study, we applied genome-wide approaches to study how FTO finds its demethylation targets in human cells. We overexpressed FTO in human HeLa cells, demonstrating that FTO effectively removes m6A from the RRACH motif enriched in the 3’UTR regions and leaves m6A at other motifs unaffected. RRACH elements are clearly enriched at FTO binding sites; however, m6A has a lower tendency to be removed from the FTO-bound RRACH. Taken together with the experimental validaton results, we propose a model in which FTO effectivly recognizes m6A-containing RRACH motifs in RNAs, leading to a faster demethylation and dissociation kinetic than the association, and consequently undectable FTO-mRNA association. However, when FTO binds to the non-m6A RRACH motif, the binding has a lower dissociation kinetics, yielding the detectable FTO binding signals which would enable FTO to have roles in regulating other mRNA processing events.
2020-03-22 | GSE101954 | GEO
Project description: Not available
| S-EPMC6789218 | biostudies-literature
Project description:To gain insight into FTO function, we knocked down and overexpressed FTO in HEK293 cells.Genetrail analyses of expression profiles pointed to the RNA splicing and processing machinery. Intriguingly, using immunocytochemistry and confocal laser scanning microscopy, we observed strong enrichment of FTO in nuclear speckles and - to a lesser extent - in nucleoli, but not in other known nuclear bodies. We also studied RNA samples of Fto knockout and wild type mice with regard to content of methylated and unmethylated nucleosidesand observed that ratios of modified and unmodified uracil and adenine were different depending on the presence of FTO. Taken together, our data suggest that FTO is involved in RNA processing and modification. We used microarrays to investigate global gene expression changes depending on the level of FTO
2012-11-13 | GSE33870 | GEO
Project description:Our understanding of posttranscriptional modifications that decorate RNA, a regulatory layer positioned between DNA and proteins, is in its infancy. N6-methyladenosine (m6A) is the most prevalent internal modification in messenger RNAs that is installed and erased by m6A methyltransferases and demethylases. The importance of these enzymes in cancer is rapidly emerging, yet information of their specific mode of actions during disease progression remain largely unknown. In the present study, we report that the m6A RNA demethylase FTO controls EMT and invasion in cancer through regulation of the Wnt pathway. We find that loss of FTO, in contrast to acute myeloid leukemia, is frequent in many cancer types, including breast and prostate cancers. Knockdown of FTO promotes tumor progression – specifically migration and invasion – in breast and prostate cancer cells. Furthermore, implantation of these cells accelerates tumor progression in recipient mice in vivo. In these tumors, FTO depletion leads to m6A-dependent activation of Wnt signaling, which drives an enhanced EMT program and invasion, thus leading to poor clinical outcome. However, loss of FTO also sensitizes cancers cells to Wnt inhibition, offering a rationale for the therapeutic targeting of Wnt for cancer patients with low FTO levels. Together, our work reveals FTO as a novel regulator of EMT and an unexpected mechanism by which Wnt signals are dysregulated in tumors, providing a rationale to stratify cancer patients treated with Wnt inhibitor. These data uncover a previously unrecognized relationship between RNA modification and EMT in cancer.
2021-01-11 | GSE128581 | GEO
Project description:Our understanding of posttranscriptional modifications that decorate RNA, a regulatory layer positioned between DNA and proteins, is in its infancy. N6-methyladenosine (m6A) is the most prevalent internal modification in messenger RNAs that is installed and erased by m6A methyltransferases and demethylases. The importance of these enzymes in cancer is rapidly emerging, yet information of their specific mode of actions during disease progression remain largely unknown. In the present study, we report that the m6A RNA demethylase FTO controls EMT and invasion in cancer through regulation of the Wnt pathway. We find that loss of FTO, in contrast to acute myeloid leukemia, is frequent in many cancer types, including breast and prostate cancers. Knockdown of FTO promotes tumor progression – specifically migration and invasion – in breast and prostate cancer cells. Furthermore, implantation of these cells accelerates tumor progression in recipient mice in vivo. In these tumors, FTO depletion leads to m6A-dependent activation of Wnt signaling, which drives an enhanced EMT program and invasion, thus leading to poor clinical outcome. However, loss of FTO also sensitizes cancers cells to Wnt inhibition, offering a rationale for the therapeutic targeting of Wnt for cancer patients with low FTO levels. Together, our work reveals FTO as a novel regulator of EMT and an unexpected mechanism by which Wnt signals are dysregulated in tumors, providing a rationale to stratify cancer patients treated with Wnt inhibitor. These data uncover a previously unrecognized relationship between RNA modification and EMT in cancer.
2021-01-11 | GSE128580 | GEO
Project description:Background: Despite its functional importance in various fundamental bioprocesses, the studies of N6-methyladenosine (m6A) in the heart are lacking. Methods: We performed methylated (m6A) RNA immunoprecipitation sequencing (MeRIP-seq) to map transcriptome-wide m6A in healthy and failing hearts. Results: Improving expression of FTO in failing mouse hearts attenuated the ischemia-induced increase in m6A and decrease in cardiac contractile function. This is carried out by the demethylation activity of FTO, which selectively demethylates cardiac contractile transcripts Conclusion: Collectively, our study demonstrates the functional importance of FTO-dependent cardiac m6A methylome in cardiac contraction during heart failure and provides a novel mechanistic insight into the therapeutic mechanisms of FTO.
2018-08-13 | GSE112789 | GEO
Project description:Single nucleotide polymorphisms in intron 1 of the fat mass and obesity-associated (FTO) gene were found to be associated with an increased risk of adult obesity. Enhanced FTO expression in mice leads to hyperphagia, increased fat mass, and higher body weight. Neuronal-specific FTOâ??deleted mice have an identical lean body weight phenotype to global FTO-deleted mice. The physiological role of adipose FTO in the homeostasis of energy regulation remains to be elucidated. We used microarrays to elucidate the metabolic pathways that are regulated by FTO in the white fat. FTO flox/flox and Adiponectin-cre FTO flox/flox (AFO) mice were fed with chow diet. White fat tissues from epididymal adipose pad were harvested under ad lib condition for RNA isolation. Three independent pools of FTO flox/flox and AFO mouse white fat RNA were included in the study.
2016-07-03 | E-GEOD-65098 | biostudies-arrayexpress