Hakai is required for stabilizing core components of the m6A mRNA methylation machinery
Ontology highlight
ABSTRACT:
PROVIDER: PRJNA666156 | ENA |
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| SRR12730390.fastq.gz | Fastqsanger.gz | |||
| SRR12730391.fastq.gz | Fastqsanger.gz | |||
| SRR12730392.fastq.gz | Fastqsanger.gz | |||
| SRR12730393.fastq.gz | Fastqsanger.gz | |||
| SRR12730394.fastq.gz | Fastqsanger.gz |
Items per page: 5 1 - 5 of 6 |
Similar Datasets
Project description:N6-methyladenosine RNA (m6A) is the most abundant internal modification on mRNA which influences most steps of mRNA metabolism and is involved in several biological functions, including circadian clock, metabolism and embryonic stem cell differentiation. The E3 ubiquitin ligase Hakai was previously found in complex with components of the m6A methylation machinery in plants and mammalian cells but its precise function remained to be investigated. Here we show that Hakai is a conserved component of the methyltransferase complex in Drosophila. Its depletion results in reduced m6A levels and altered m6A-dependent functions including sex determination. We show that its ubiquitination domain is required for dimerisation and interaction with other members of the m6A machinery, while its catalytic activity seems dispensable. Finally, we demonstrate that the loss of Hakai destabilizes the level of several subunits of the methyltransferase complex, resulting in impaired m6A deposition. Thus, our work adds new functional and molecular insights into the mechanism of the m6A mRNA writer complex.
2021-04-26 | GSE158663 | GEO
Project description:N6-methyladenosine (m6A) is the most abundant internal modification on mRNA which influences most steps of mRNA metabolism and is involved in several biological functions. The E3 ubiquitin ligase Hakai was previously found in complex with components of the m6A methylation machinery in plants and mammalian cells but its precise function remained to be investigated. Here we show that Hakai is a conserved component of the methyltransferase complex in Drosophila and human cells. In Drosophila, its depletion results in reduced m6A levels and altered m6A-dependent functions including sex determination. We show that its ubiquitination domain is required for dimerisation and interaction with other members of the m6A machinery, while its catalytic activity is dispensable. Finally, we demonstrate that the loss of Hakai destabilizes several subunits of the methyltransferase complex, resulting in impaired m6A deposition. Our work adds new functional and molecular insights into the mechanism of the m6A mRNA writer complex.
2021-04-29 | PXD022294 | Pride
Project description:Using in vivo interaction proteomics, two HAKAI-interacting zinc finger proteins, HIZ1 and HIZ2, were discovered as novel components of the Arabidopsis m6A writer complex. HAKAI is required for the interaction between HIZ1 and MTA. Whilst HIZ1 knockout plants have normal levels of m6A, plants in which it is overexpressed show reduced methylation. In addition, HIZ1 was found to be involved in root hair development upon auxin transport inhibition in a HAKAI-dependent manner. Mutant plants lacking HIZ2 are viable but have an 85% reduction in m6A abundance and show severe developmental defects. Our findings suggest that HIZ1 appears to be a HAKAI-dependent negative regulator of m6A deposition and HIZ2 is a novel and essential member of the Arabidopsis m6A writer complex.
2022-01-21 | GSE188610 | GEO
Project description:Using in vivo interaction proteomics, two HAKAI-interacting zinc finger proteins, HIZ1 and HIZ2, were discovered as novel components of the Arabidopsis m6A writer complex. HAKAI is required for the interaction between HIZ1 and MTA. Whilst HIZ1 knockout plants have normal levels of m6A, plants in which it is overexpressed show reduced methylation. In addition, HIZ1 was found to be involved in root hair development upon auxin transport inhibition in a HAKAI-dependent manner. Mutant plants lacking HIZ2 are viable but have an 85% reduction in m6A abundance and show severe developmental defects. Our findings suggest that HIZ1 appears to be a HAKAI-dependent negative regulator of m6A deposition and HIZ2 is a novel and essential member of the Arabidopsis m6A writer complex.
2022-01-20 | GSE174573 | GEO
Project description:The methyltransferase complex (m6A writer), which catalyzes the deposition of N6-methyladenosine (m6A) in mRNAs, is highly conserved across most eukaryotic organisms, but its components and interactions between them are still far from fully understood. Using in vivo interaction proteomics, two HAKAI-interacting zinc finger proteins, HIZ1 and HIZ2, were discovered as novel components of the Arabidopsis m6A writer complex. HAKAI is required for the interaction between HIZ1 and MTA. Whilst HIZ1 knockout plants have normal levels of m6A, plants in which it is overexpressed show reduced methylation. In addition, HIZ1 was found to be involved in root hair development upon auxin transport inhibition in a HAKAI-dependent manner. Mutant plants lacking HIZ2 are viable but have an 85% reduction in m6A abundance and show severe developmental defects. Our findings suggest that HIZ1 appears to be a HAKAI-dependent negative regulator of m6A deposition and HIZ2 is a novel and essential member of the Arabidopsis m6A writer complex.
2022-02-09 | PXD026287 | Pride
Project description:In order to obtain a stringent m6A methylome in Drosophila adults, we performed m6A-RIP-seq in yw control (male and female), Mettl3 (male), Mettl14 (male) and Hakai (male) mutant flies. We find that the effective m6A modification, which depends on the writer complex, is mostly distributed in 5’ UTR and near start codon in Drosophila, in contrast to the mammalian system. We define a set of high-confident m6A methylation sites shared by Mettl3, Mettl14 and Hakai, indicating that Hakai is a core component of the m6A writer complex. We also find differential methylation pattern in certain loci between male and female flies.
2021-03-01 | GSE155662 | GEO
Project description:N6-methyladenosine (m6A) is the most abundant internal modification on mRNA which influences most steps of mRNA metabolism and is involved in several biological functions. The E3 ubiquitin ligase Hakai was previously found in complex with components of the m6A methylation machinery in plants and mammalian cells but its precise function remained to be investigated. Here we show that Hakai is a conserved component of the methyltransferase complex in Drosophila and human cells. In Drosophila, its depletion results in reduced m6A levels and altered m6A-dependent functions including sex determination. We show that its ubiquitination domain is required for dimerization and interaction with other members of the m6A machinery, while its catalytic activity is dispensable. Finally, we demonstrate that the loss of Hakai destabilizes several subunits of the methyltransferase complex, resulting in impaired m6A deposition. Our work adds functional and molecular insights into the mechanism of the m6A mRNA writer complex.
| S-EPMC8213727 | biostudies-literature
Project description:E3 ubiquitin-ligases are important for the cellular protein homeostasis and their deregulation is implicated in cancer. The E3 ubiquitin-ligase Hakai is involved in tumour progression and metastasis, through the regulation of the tumour suppressor E-cadherin. Hakai is overexpressed in colon cancer, however, the implication in colitis-associated cancer is unknown. Here, we investigated the potential role of Hakai in intestinal inflammation and cancer bowel disease.Several mouse models of colitis and associated cancer were used including AOM-DSS, acute colitis, and genetically modified mice deficient for the IL-10 gene, to analyse Hakai expression by immunohistochemistry. Interactome analysis of Hakai was performed and effect on selected protein was determined by plasmid and siRNA transfection, western-blotting, immunoprecipitation, immunofluorescence and ubiquitination assays. Lipid accumulation was assayed by oil red staining. Immunohistochemistry was also performed in inflamed colon biopsies from ulcerative colitis, Crohn's disease and colorectal cancer patients. Our results show that Hakai was downregulated in inflammatory tissues in different mouse models. Fatty Acid Synthase (FASN) protein was identified as a novel Hakai-interacting protein. Hakai induces FASN ubiquitination and degradation via lysosome, thus regulating FASN-mediated lipid accumulation. An inverse expression of FASN with Hakai expression was detected in inflammatory AOM/DSS mouse model. In conclusion, Hakai regulates FASN ubiquitination and degradation, resulting in the regulation of FASN-mediated lipid accumulation, which is associated to the development of inflammatory bowel disease. The interaction between Hakai and FASN may be an important mechanism for the homeostasis of intestinal barrier function and in the pathogenesis of this disease.
2022-10-25 | PXD029136 | Pride
Project description:N6-methyladenosine (m6A) is an abundant modification in eukaryotic mRNA, regulating mRNA dynamics by influencing mRNA stability, splicing, export and translation. Recent studies discovered m6A methyltransferases (?writer?), demethylases (?eraser?) and binding proteins (?reader?), which modulate m6A methylation. However, the precise m6A regulating machinery still remains incompletely understood. Here we demonstrate that ZC3H13, a zinc finger protein, plays an essential role in modulating m6A methylation on polyadenylated RNA in the nucleus. ZC3H13 exists in an evolutionary-conserved macromolecular complex containing WTAP, Virilizer and Hakai. We confirm the interaction among those proteins and demonstrate that knockdown of Zc3h13 in mouse embryonic stem cell (mESC) significantly decreases global m6A level on mRNA, mainly at 3? untranslated regions (3? UTR). Interestingly, fractionation assays show that upon Zc3h13 knockdown a great majority of WTAP, Virilizer and Hakai translocate to the cytoplasm and the nuclear presence of the methyltransferase Mettl3 and Mettl14 also decrease significantly. In contrast, knockdown of WTAP, Virilizer or Hakai does not change the nuclear localization of Zc3h13. This suggests that Zc3h13 is required for nuclear localization of the Zc3h13-WTAP-Virilizer-Hakai complex, which is important for RNA m6A methylation. Finally, Zc3h13 depletion, as does WTAP, Virilizer or Hakai, impairs self-renewal and triggers mESC differentiation. Taken together, our findings demonstrate that Zc3h13 plays an essential role in anchoring WTAP, Virilizer and Hakai in the nucleus to facilitate m6A methylation and to regulate mESC self-renewal.
2018-03-14 | GSE94148 | GEO
Project description:N6-methyladenosine (m6A) is an abundant RNA modification in eukaryotes, playing crucial roles in multiple biological processes. m6A is catalyzed by the activity of Mettl3, which depends on additional proteins whose precise functions remain poorly understood. Here we identified Flacc/Zc3h13 as a novel interactor of m6A methyltransferase complex components in Drosophila and mouse. Like other components, Flacc controls m6A levels and is involved in sex determination in Drosophila. We demonstrate that Flacc promotes the recruitment of the methyltransferase to mRNA by bridging Fl(2)d to the mRNA binding factor Spenito. Altogether, our work advances our molecular understanding of conservation and regulation of the m6A machinery.
2018-12-20 | PXD008111 | Pride