Non-microRNA Binding Competitively Inhibits LIN28 Regulation
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ABSTRACT:
PROVIDER: PRJNA738154 | ENA |
REPOSITORIES: ENA
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Project description:RNA binding protein (RBP) expression is finite. For RBPs that are vastly outnumbered by their potential target sites, a simple competition for binding can set the magnitude of post-transcriptional control. Here we show that LIN28, best known for its direct regulation of let-7 miRNA biogenesis, is also indirectly regulated by its widespread binding of non-miRNA transcripts. Approximately 99% of LIN28 binding sites are found on non-miRNA transcripts, like protein coding and ribosomal RNAs. These sites are bound specifically and strongly, but they do not appear to mediate direct post-transcriptional regulation. Instead, non-miRNA sites act to sequester LIN28 protein and effectively change its functional availability, thus impeding the regulation of let-7 in cells. Together, these data show that the binding properties of the transcriptome broadly influence the ability of an RBP to mediate changes in RNA metabolism and gene expression.
2022-01-07 | GSE178259 | GEO
Project description: Not available
| S-EPMC8670721 | biostudies-literature
Project description:LIN28 (also known as LIN28A), a highly conserved RNA-binding protein, has emerged as a central post-transcriptional regulator of cell fate through blockade of let-7 microRNA (miRNA) biogenesis and direct modulation of mRNA translation. To gain insight into how LIN28 is integrated with the pluripotency signaling network, we investigated the role of LIN28 phosphorylation. We employed a targeted phosphoproteomics strategy in mouse ESCs (mESCs) and were able to map four phosphosites, two of which, S184 and S200, were confidently assigned to specific serine residues.
2016-10-30 | MSV000080302 | MassIVE
Project description:LIN28 is a conserved RNA binding protein implicated in pluripotency, reprogramming and oncogenesis. Previously shown to act primarily by blocking let-7 microRNA (miRNA) biogenesis, here we elucidate distinct roles of LIN28 regulation via its direct messenger RNA (mRNA) targets. Through cross-linking and immunoprecipitation coupled with high-throughput sequencing (CLIP-seq) in human embryonic stem cells and somatic cells expressing exogenous LIN28, we have defined discrete LIN28 binding sites in a quarter of human transcripts. These sites revealed that LIN28 binds to GGAGA sequences enriched within loop structures in mRNAs, reminiscent of its interaction with let-7 miRNA precursors. Among LIN28 mRNA targets, we found evidence for LIN28 autoregulation and also direct but differing effects on the protein abundance of splicing regulators in somatic and pluripotent stem cells. Splicing-sensitive microarrays demonstrated that exogenous LIN28 expression causes widespread downstream alternative splicing changes. These findings identify important regulatory functions of LIN28 via direct mRNA interactions.
2012-09-10 | GSE39872 | GEO
Project description:The conserved human LIN28 RNA-binding proteins function in development, maintenance of pluripotency and oncogenesis. We used PAR-CLIP and a newly developed variant of this method, iDo-PAR-CLIP, to identify LIN28B targets as well as sites bound by the individual RNA binding domains of LIN28B in the human transcriptome at nucleotide resolution. The position of target binding sites reflected the known structural relative orientation of individual LIN28B binding domains, validating iDo-PAR-CLIP. Our data suggest that LIN28B directly interacts with most expressed mRNAs and members of the let-7 microRNA family. The Lin28 binding motif detected in pre-let-7 was enriched in mRNA sequences bound by LIN28B. Upon LIN28B knock down, cell proliferation and the cell cycle were strongly impaired. Quantitative shotgun proteomics of LIN28B depleted cells revealed significant reduction of protein synthesis from its RNA targets that function in translation, mRNA splicing and cell cycle control. Computational analyses provided evidence that the strength of protein synthesis reduction correlated with the location of LIN28B binding sites within target transcripts.
2013-06-25 | GSE46908 | GEO
Project description:LIN28 is a conserved RNA binding protein implicated in pluripotency, reprogramming and oncogenesis. Previously shown to act primarily by blocking let-7 microRNA (miRNA) biogenesis, here we elucidate distinct roles of LIN28 regulation via its direct messenger RNA (mRNA) targets. Through cross-linking and immunoprecipitation coupled with high-throughput sequencing (CLIP-seq) in human embryonic stem cells and somatic cells expressing exogenous LIN28, we have defined discrete LIN28 binding sites in a quarter of human transcripts. These sites revealed that LIN28 binds to GGAGA sequences enriched within loop structures in mRNAs, reminiscent of its interaction with let-7 miRNA precursors. Among LIN28 mRNA targets, we found evidence for LIN28 autoregulation and also direct but differing effects on the protein abundance of splicing regulators in somatic and pluripotent stem cells. Splicing-sensitive microarrays demonstrated that exogenous LIN28 expression causes widespread downstream alternative splicing changes. These findings identify important regulatory functions of LIN28 via direct mRNA interactions.
2012-09-10 | GSE39855 | GEO
Project description:The conserved human LIN28 RNA-binding proteins function in development, maintenance of pluripotency and oncogenesis. We used PAR-CLIP and a newly developed variant of this method, iDo-PAR-CLIP, to identify LIN28B targets as well as sites bound by the individual RNA binding domains of LIN28B in the human transcriptome at nucleotide resolution. The position of target binding sites reflected the known structural relative orientation of individual LIN28B binding domains, validating iDo-PAR-CLIP. Our data suggest that LIN28B directly interacts with most expressed mRNAs and members of the let-7 microRNA family. The Lin28 binding motif detected in pre-let-7 was enriched in mRNA sequences bound by LIN28B. Upon LIN28B knock down, cell proliferation and the cell cycle were strongly impaired. Quantitative shotgun proteomics of LIN28B depleted cells revealed significant reduction of protein synthesis from its RNA targets that function in translation, mRNA splicing and cell cycle control. Computational analyses provided evidence that the strength of protein synthesis reduction correlated with the location of LIN28B binding sites within target transcripts. We used PAR-CLIP and a newly developed variant of this method, iDo-PAR-CLIP, to identify LIN28B targets as well as sites bound by the individual RNA binding domains of LIN28B in the human transcriptome at nucleotide resolution.
2013-06-25 | E-GEOD-46908 | biostudies-arrayexpress
Project description:LIN28 is a conserved RNA binding protein implicated in pluripotency, reprogramming and oncogenesis. Previously shown to act primarily by blocking let-7 microRNA (miRNA) biogenesis, here we elucidate distinct roles of LIN28 regulation via its direct messenger RNA (mRNA) targets. Through cross-linking and immunoprecipitation coupled with high-throughput sequencing (CLIP-seq) in human embryonic stem cells and somatic cells expressing exogenous LIN28, we have defined discrete LIN28 binding sites in a quarter of human transcripts. These sites revealed that LIN28 binds to GGAGA sequences enriched within loop structures in mRNAs, reminiscent of its interaction with let-7 miRNA precursors. Among LIN28 mRNA targets, we found evidence for LIN28 autoregulation and also direct but differing effects on the protein abundance of splicing regulators in somatic and pluripotent stem cells. Splicing-sensitive microarrays demonstrated that exogenous LIN28 expression causes widespread downstream alternative splicing changes. These findings identify important regulatory functions of LIN28 via direct mRNA interactions. CLIP-seq for LIN28-V5 in stable human Flp-In-293 cells, and LIN28 in hES cells; strand-specific mRNA-seq for uninfected, control KD, and LIN28 KD human H9 ES cells; and strand-specific smallRNA-seq for uninfected, control KD, and LIN28 KD human H9 ES cells.
2012-09-10 | E-GEOD-39872 | biostudies-arrayexpress
Project description:LIN28 is an RNA binding protein with important roles in early embryo development by interaction with let-7 microRNA precursors or mRNAs in cytoplasm. Here, we showed novel roles of LIN28 within the nucleolus in cultured pluripotent stem cells (PSCs). We found Lin28 deficiency led to activation of the 2-cell (2C)-like transcriptional program and repression of the ES cell program. Mechanistically, LIN28 maintains a peri-nucleolar heterochromatin-residing complex to repress 2C activator Dux via association with NCL and TRIM28. Meanwhile, it promotes liquid-liquid phase separation (LLPS) of nucleolar proteins FBL and NCL to support nucleolar function through its RNA binding domains and intrinsic disordered regions. These findings elucidate novel roles for nucleolar LIN28 in coordinately regulating the ES/2C state homeostasis, and shed light on RNA binding protein-mediated nucleolar phase separation mechanisms in PSCs and early embryo development.
2021-09-15 | GSE164414 | GEO
Project description:LIN28 is an RNA binding protein with important roles in early embryo development by interaction with let-7 microRNA precursors or mRNAs in cytoplasm. Here, we showed novel roles of LIN28 within the nucleolus in cultured pluripotent stem cells (PSCs). We found Lin28 deficiency led to activation of the 2-cell (2C)-like transcriptional program and repression of the ES cell program. Mechanistically, LIN28 maintains a peri-nucleolar heterochromatin-residing complex to repress 2C activator Dux via association with NCL and TRIM28. Meanwhile, it promotes liquid-liquid phase separation (LLPS) of nucleolar proteins FBL and NCL to support nucleolar function through its RNA binding domains and intrinsic disordered regions. These findings elucidate novel roles for nucleolar LIN28 in coordinately regulating the ES/2C state homeostasis, and shed light on RNA binding protein-mediated nucleolar phase separation mechanisms in PSCs and early embryo development.
2021-09-15 | GSE164296 | GEO