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.
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 FTO–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.
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.
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.
Project description:RNA methylation plays an important role fine tuning translation and subsequently regulating cellular responses and cell fate. The fat mass- and obesity-associated protein (FTO) was recognized as an m6A demethylase and described as an oncogenic factor in leukemia and brain tumors. FTO expression levels are suppressed in ovarian tumors and ovarian cancer stem cells (CSCs). FTO induce cyclic AMP activity through targeting PDE4B and PDE1C by down-regulation of m6A levels in the mRNA transcript. In all or findings point to a tumor suppressor function of FTO in high grade serous OC
Project description:We identified the target genes of FTO ("fat mass and obesity associated") in primary cultures of human skeletal muscle cells using adenoviral vectors expressing FTO or GFP and oligonucleotide microarrays.
Project description:We identified the target genes of FTO ("fat mass and obesity associated") in primary cultures of human skeletal muscle cells using adenoviral vectors expressing FTO or GFP and oligonucleotide microarrays. Human myotubes were prepared from 4 different skeletal muscle biopsies. After differentiation, myotubes were infected for 48 hours with recombinant adenovirus expressing either GFP or FTO. Each FTO-infected myotubes culture was compared to GFP-infected myotubes culture. GFP-infected myotubes were regarded as the control. Four biological replicates were processed.
Project description:RNA methylation plays an important role fine tuning translation and subsequently regulating cellular responses and cell fate. The fat mass- and obesity-associated protein (FTO) was recognized as an m6A demethylase and described as an oncogenic factor in leukemia and brain tumors. FTO expression levels are suppressed in ovarian tumors and ovarian cancer stem cells (CSCs). FTO induce cyclic AMP activity through targeting PDE4B and PDE1C by down-regulation of m6A levels in the mRNA transcript. In all or findings point to a tumor suppressor function of FTO in high grade serous OC. FTO induce cyclic AMP activity through targeting PDE4B and PDE1C by down-regulation of m6A levels in the mRNA transcript. In all or findings point to a tumor suppressor function of FTO in high grade serous OC
Project description:Genome-wide association studies in diverse populations have reproducibly associated variants within introns of FTO with increased risk for obesity and type-2 diabetes.While the molecular mechanisms linking these noncoding variants with obesity are not immediately obvious, subsequent studies in mice demonstrated that FTO expression levels influence body mass and composition phenotypes. Yet, no direct connection between the obesity-associated intronic variants and FTO expression or function has been made. We show that the obesity-associated noncoding sequences within FTO are functionally connected, at megabase distances, primarily with the homeobox gene IRX3, rather than with FTO. 4C-seq samples for Fto/fto and Irx3/irx3a genes promoters in different samples: adult mouse brain, E9.5 mouse embryos and 24hpf zebrafish embryos.
Project description:N6-methyladenosine (m6A) is the most abundant internal modification on mammalian messenger RNA (mRNA). It is installed by a writer complex and can be reversed by erasers such as the fat mass and obesity-associated protein (FTO). Despite extensive research, the primary physiological substrates of FTO in mammalian tissues and development remain elusive. Here, we show that FTO mediates m6A demethylation of long-interspersed element-1 (LINE1) RNA in mouse embryonic stem cells (mESCs), regulating LINE1 RNA abundance and the local chromatin state, which in turn modulates transcription of LINE1-containing genes. FTO-mediated LINE1 RNA m6A demethylation also plays regulatory roles in shaping chromatin state and gene expression during mouse oocyte and embryonic development. Our results suggest broad effects of LINE1 RNA m6A demethylation by FTO in mammals.