Molecular Basis for Interaction of let-7 MicroRNAs with Lin28

Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, Boston, MA 02115, USA.
Cell (Impact Factor: 32.24). 11/2011; 147(5):1080-91. DOI: 10.1016/j.cell.2011.10.020
Source: PubMed


MicroRNAs (miRNAs) are small noncoding RNA molecules that regulate gene expression. Among these, members of the let-7 miRNA family control many cell-fate determination genes to influence pluripotency, differentiation, and transformation. Lin28 is a specific, posttranscriptional inhibitor of let-7 biogenesis. We report crystal structures of mouse Lin28 in complex with sequences from let-7d, let-7-f1, and let-7 g precursors. The two folded domains of Lin28 recognize two distinct regions of the RNA and are sufficient for inhibition of let-7 in vivo. We also show by NMR spectroscopy that the linker connecting the two folded domains is flexible, accommodating Lin28 binding to diverse let-7 family members. Protein-RNA complex formation imposes specific conformations on both components that could affect downstream recognition by other processing factors. Our data provide a molecular explanation for Lin28 specificity and a model for how it regulates let-7.

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    • "One study in C. elegans also suggested binding of Lin28A to the let-7 genomic locus (Van Wynsberghe et al., 2011). Structurally, Lin28A is composed of two classical nucleic acid binding domains: a cold-shock domain (CSD) and a pair of CCHC-type zinc finger motifs (Moss et al., 1997), both of which contribute to the affinity for pre-let7 and repression of let-7 maturation (Loughlin et al., 2012; Nam et al., 2011). Notably, CSD domains in other molecules have been shown to bind DNA (Hudson and Ortlund, 2014), raising the possibility that Lin28A may also exhibit DNA binding activity. "
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    ABSTRACT: Lin28, a well-known RNA-binding protein, regulates diverse cellular properties. All physiological functions of Lin28A characterized so far have been attributed to its repression of let-7 miRNA biogenesis or modulation of mRNA translational efficiency. Here we show that Lin28A directly binds to a consensus DNA sequence in vitro and in mouse embryonic stem cells in vivo. ChIP-seq and RNA-seq reveal enrichment of Lin28A binding around transcription start sites and a positive correlation between its genomic occupancy and expression of many associated genes. Mechanistically, Lin28A recruits 5-methylcytosine-dioxygenase Tet1 to genomic binding sites to orchestrate 5-methylcytosine and 5-hydroxymethylcytosine dynamics. Either Lin28A or Tet1 knockdown leads to dysregulated DNA methylation and expression of common target genes. These results reveal a surprising role for Lin28A in transcriptional regulation via epigenetic DNA modifications and have implications for understanding mechanisms underlying versatile functions of Lin28A in mammalian systems.
    Full-text · Article · Dec 2015 · Molecular cell
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    • "There are two paralogs of LIN28, LIN28A, and LIN28B, both containing a cold shock domain (CSD) and CCHC-zinc finger RNA-binding domain. They regulate let-7 miRNA levels by CSD binding to the NGN- GAYNNN (N = any base and Y = pyrimidine) sequence on the terminal loop of let-7 and CCHC-zinc finger binding to the GGAG sequence on the same terminal loop [26]. The linker between the CSD and the CCHC-zinc finger allows for binding of all twelve let-7 miRNA family members. "
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    ABSTRACT: Epithelial ovarian cancer is the most aggressive and deadly form of ovarian cancer and is the most lethal gynecological malignancy worldwide; therefore, efforts to elucidate the molecular factors that lead to epithelial ovarian cancer are essential to better understand this disease. Recent studies reveal that tumor cells release cell-secreted vesicles called exosomes and these exosomes can transfer RNAs and miRNAs to distant sites, leading to cell transformation and tumor development. The RNA-binding protein LIN28 is a known marker of stem cells and when expressed in cancer, it is associated with poor tumor outcome. We hypothesized that high LIN28 expressing ovarian cancer cells secrete exosomes that can be taken up by nontumor cells and cause changes in gene expression and cell behavior associated with tumor development. IGROV1 cells were found to contain high LIN28A and secrete exosomes that were taken up by HEK293 cells. Moreover, exposure to these IGROV1 secreted exosomes led to significant increases in genes involved in Epithelial-to-Mesenchymal Transition (EMT), induced HEK293 cell invasion and migration. These changes were not observed with exosomes secreted by OV420 cells, which contain no detectable amounts of LIN28A or LIN28B. No evidence was found of LIN28A transfer from IGROV1 exosomes to HEK293 cells.
    Full-text · Article · Nov 2015
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    • "These functions of let-7 miRNAs primarily are accomplished in differentiated cells where they are expressed abundantly. Let-7 pri-and pre-miRNAs harbor a typical hairpin structure with a stem containing the let-7-5p miRNA sequence base paired extensively with the partially complementary let- 7-3p miRNA sequence, connected by a so-called terminal loop region of variable lengths and structures among different let-7 family members, a region referred to as pre-element (preE) (Nam et al., 2011). Let-7 preE serves as a platform to recruit RNA-binding proteins, such as LIN28, KHSRP (also known as KSRP), hnRNPA1, and TRIM25, in order to posttranscriptionally regulate let-7 biogenesis (Heo et al., 2008; Michlewski and Cá ceres, 2010; Newman et al., 2008; Rybak et al., 2008; Trabucchi et al., 2009; Viswanathan et al., 2008; Zhang et al., 2015). "
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    ABSTRACT: Let-7 microRNAs (miRNAs) are critical regulators of animal development, stem cell differentiation, glucose metabolism, and tumorigenesis. Mammalian genomes contain 12 let-7 isoforms that suppress expression of a common set of target mRNAs. LIN28 proteins selectively block let-7 biogenesis in undifferentiated cells and in cancer. The current model for coordinate let-7 repression involves the LIN28 cold-shock domain (CSD) binding the terminal loop and the two CCHC-type zinc fingers recognizing a GGAG sequence motif in precursor let-7 (pre-let-7) RNAs. Here, we perform a systematic analysis of all let-7 miRNAs and find that a single let-7 family member, human let-7a-3 (and its murine ortholog let-7c-2), escapes LIN28-mediated regulation. Mechanistically, we find that the pre-let-7c-2 loop precludes LIN28A binding and regulation. These findings refine the current model of let-7 regulation by LIN28 proteins and have important implications for understanding the LIN28/let-7 axis in development and disease.
    Full-text · Article · Oct 2015 · Cell Reports
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