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Identify QKI-bound transcripts via RIP-seq

ABSTRACT: N7-methylguanosine (m7G) modification, routinely occurring at the 5’ cap of mRNA or within tRNA and rRNA, also exists internally in mRNA. Although essential for mRNA translation as well as stress response, the “reader” protein for mRNA internal m7G modification is still unrevealed. Here, we reported that Quaking protein (QKI), especially QKI7, can selectively recognize the internal mRNA m7G decoration in the cytosol of various cell types. We identified over 1000 confident m7G-modified and QKI binding RNA targets with a conserved motif, “GANGAN (N=A/U/G)”. More strikingly, internal m7G reader QKI7 directly interacts with the SG core protein G3BP1 and can shuttle a subset of m7G-modified transcripts into SG mRNA pool under oxidative stress condition. Additionally, by sequestering mRNA within SGs, QKI7 modulates the translation efficiency of selected transcripts. Moreover, in line with the observation that doxorubicin triggers the assembly of SGs, QKI7 mediates the sensitivity of cancer cells to chemotherapy drug treatment.

ORGANISM(S): Homo sapiens

PROVIDER: GSE193036 | GEO | 2023/05/17

REPOSITORIES: GEO

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Effect of QKI7 on global RNA stability

Project description:N7-methylguanosine (m7G) modification, routinely occurring at the 5’ cap of mRNA or within tRNA and rRNA, also exists internally in mRNA. Although essential for mRNA translation as well as stress response, the “reader” protein for mRNA internal m7G modification is still unrevealed. Here, we reported that Quaking protein (QKI), especially QKI7, can selectively recognize the internal mRNA m7G decoration in the cytosol of various cell types. We identified over 1000 confident m7G-modified and QKI binding RNA targets with a conserved motif, “GANGAN (N=A/U/G)”. More strikingly, internal m7G reader QKI7 directly interacts with the SG core protein G3BP1 and can shuttle a subset of m7G-modified transcripts into SG mRNA pool under oxidative stress condition. Additionally, by sequestering mRNA within SGs, QKI7 modulates the translation efficiency of selected transcripts. Moreover, in line with the observation that doxorubicin triggers the assembly of SGs, QKI7 mediates the sensitivity of cancer cells to chemotherapy drug treatment.

2023-05-18 | GSE218222 | GEO

Identify QKI7-bound transcripts via RIP-seq

2023-05-17 | GSE192847 | GEO

Determine the effects of QKI7 on translation efficiency via ribosome profiling (Ribo-seq)

2023-05-17 | GSE192846 | GEO

Effect of METTL1 KO on the binding ability of QKI7

2023-05-17 | GSE218221 | GEO

Recognition of internal mRNA N7-methylguanosine by QKI shuttles transcripts into stress granules and regulates translation

Project description:Recent studies reported that N7-methylguanosine (m7G) modification exists in internal mRNAs; however, the “reader” protein for mRNA internal m7G modification is still unrevealed. Here, by performing m7G MeRIP-seq and RIP-seq, we identified quaking protein (QKI) as a novel internal m7G modification reader. Internal mRNA m7G modification acts as a key player in mRNA translation under stress condition. To explore the mechanism underlying the function of QKI in translation regulation, we employed RNA-seq and Ribo-seq, and demonstrated that QKI7 regulates the translation efficiency of a subset of internal m7G-modified transcripts under stress condition.

2023-05-17 | GSE193039 | GEO

Transcriptome Mapping of Internal N7-methylguanosine Methylome in Mammalian mRNA

Project description:N7-methylguanosine (m7G) is a positively charged, essential modification at the 5′ cap of eukaryotic mRNA, regulating mRNA export, translation, and splicing. m7G also occurs internally within tRNA and rRNA, but its existence and distribution within eukaryotic mRNA remain to be investigated. Here, we show the presence of internal m7G sites within mammalian mRNA. We then performed transcriptome-wide profiling of internal m7G methylome using m7G-MeRIP sequencing (MeRIP-seq). To map this modification at base resolution, we developed a chemical-assisted sequencing approach that selectively converts internal m7G sites into abasic sites, inducing misincorporation at these sites during reverse transcription. This base-resolution m7G-seq enabled transcriptome-wide mapping of m7G in human tRNA and mRNA, revealing distribution features of the internal m7G methylome in human cells. We also identified METTL1 as a methyltransferase that installs a subset of m7G within mRNA and showed that internal m7G methylation could affect mRNA translation.

2019-04-26 | GSE112276 | GEO

Detection of internal N7-methylguanosine (m7G) RNA modifications by mutational profiling sequencing

Project description:Methylation of guanosine on position N7 (m7G) on internal RNA positions have been found in all domains of life and have been implicated in human disease, but m7G modifications has so far only been mapped in a limited number of RNA molecules. Here, we present m7G Mutational Profiling sequencing (m7G-MaP-seq), which allows high throughput detection of m7G modifications. In our method, m7G modified positions are converted to abasic sites by mild reduction, recorded as cDNA mutations through reverse transcription, sequenced and subsequently detected by identification of positions with increased mutation rates in the reduced sample compared to the control. We show that m7G-MaP-seq efficiently detects m7G modifications in rRNA, including a previously uncharacterised rRNA modification in Arabidopsis thaliana. Furthermore, we identify m7G tRNA modifications in budding yeast, human and arabidopsis tRNA and show that m7G modification occurs before tRNA splicing. We do not find any evidence for internal m7G modifications being present in other small RNA, such as miRNA, snoRNA and sRNA. Likewise, high coverage m7G-MaP-seq analysis of mRNA from E. coli or yeast cells failed to identify any internal m7G modifications.

2018-10-30 | GSE121927 | GEO

Transcriptome Mapping of Internal N7-methylguanosine modification via m7G MeRIP-seq [m7g.2]

Project description:Previous study found the presence of internal N7-methylguanosine (m7G) within mammalian mRNA. We performed antibody-based m7G MeRIP-seq for transcriptome-wide mapping the internal m7G sites in U2OS cells.

2023-05-17 | GSE192845 | GEO

Transcriptome Mapping of Internal N7-methylguanosine modification via m7G MeRIP-seq [m7g.1]

2023-05-17 | GSE192844 | GEO

RNA partitioning into stress granules is independent of G3BP

Project description:Stress granules (SGs) are stress induced RNA-protein assemblies formed from untranslating mRNPs. Mammalian SGs are non-uniform and contain “cores”, which are stable in lysates and heterogenous in protein composition. The interactions defining RNA recruitment into SGs, and whether the RNA composition varies between different cores remains unclear. Herein, we determine the SG transcriptome through purification of both PABPC1 and G3BP SG cores during both arsenite and sorbitol stress. We observe that similar RNAs are recruited to SGs independent of the protein used for core purification, suggesting individual RNA-binding proteins (RBPs) may play a limited role in defining the SG transcriptome. Analysis of the sorbitol-induced SG transcriptome reveals G3BP1/2 has little effect on RNAs recruited to SGs suggesting RNAs localize to SGs independent of individual RBPs. Taken together, we suggest that partitioning of RNAs to SGs is independent of at least some proteins, consistent with SGs forming through intermolecular RNA-RNA interactions.

2019-01-30 | GSE119977 | GEO

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