Regulation of LINE-1 in mammals

Transposable elements (TEs) are mobile DNA elements that represent almost half of the human genome. Transposition of TEs has been implicated as a source of genome evolution and acquisition of new traits but also as an origin of diseases. The activity of these elements is therefore tightly regulated during the life cycle of each individual, and many recent discoveries involved the genetic and epigenetic mechanisms in their control. In this review, we present recent findings in this field of research, focusing on the case of one specific family of TEs: the long-interspersed nuclear elements-1 (LINE-1 or L1). LINE-1 elements are the most representative class of retrotransposons in mammalian genomes. We illustrate how these elements are conserved between mice and humans, and how they are regulated during the life cycle. Additionally, recent advances in genome-wide sequencing approaches allow us not only to better understand the regulation of LINE-1 but also highlight new issues specifically at the bioinformatics level. Therefore, we discuss the state of the art in analyzing such bioinformatics datasets to identify epigenetic regulators of repeated elements in the human genome

Regulation of LINE-1 Elements by miR-128 Is Not Conserved in Mouse Embryonic Stem Cells

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IP3 signalling regulates exogenous RNAi in Caenorhabditis elegans.

RNA interference (RNAi) is a widespread and widely exploited phenomenon. Here, we show that changing inositol 1,4,5-trisphosphate (IP3) signalling alters RNAi sensitivity in Caenorhabditis elegans. Reducing IP3 signalling enhances sensitivity to RNAi in a broad range of genes and tissues. Conversely up-regulating IP3 signalling decreases sensitivity. Tissue-specific rescue experiments suggest IP3 functions in the intestine. We also exploit IP3 signalling mutants to further enhance the sensitivity of RNAi hypersensitive strains. These results demonstrate that conserved cell signalling pathways can modify RNAi responses, implying that RNAi responses may be influenced by an animal's physiology or environment.We thank A. Fire, K. Ford, S. Mitani and H. Peterkin for the provision of plasmids and strains. Some strains were provided by the CGC, which is funded by NIH Office of Research Infrastructure Programs (P40 OD010440). Other strains were provided by the Mitani Lab through the National Bio‐Resource Project of the MEXT, Japan. We are grateful to J. Ahringer, B. Olofsson and members of the Baylis group for helpful discussions. AIN was funded by Trinity Hall College, Cambridge and the Cambridge European Trust. The work of MDS and RPV‐M was partially funded by a Miguel Servet Grant (CP11/00090) from the Health Research Institute Carlos III, which is partially supported by the European Regional Development Fund. RPV‐M is a Marie Curie fellow (CIG322034). RG was funded by the MRC (G0601106).This is the accepted manuscript. The final version is available from Embo Press at http://embor.embopress.org/content/early/2015/01/21/embr.201439585

Noncanonical function of DGCR8 controls mESC exit from pluripotency

Mouse embryonic stem cells (mESCs) deficient for DGCR8, a key component of the microprocessor complex, present strong differentiation defects. However, the exact reasons impairing their commitment remain elusive. The analysis of newly generated mutant mESCs revealed that DGCR8 is essential for the exit from the pluripotency state. To dissociate canonical versus noncanonical functions of DGCR8, we complemented the mutant mESCs with a phosphomutant DGCR8, which restored microRNA levels but did not rescue the exit from pluripotency defect. Integration of omics data and RNA immunoprecipitation experiments established DGCR8 as a direct interactor of Tcf7l1 mRNA, a core component of the pluripotency network. Finally, we found that DGCR8 facilitated the splicing of Tcf7l1, an event necessary for the differentiation of mESCs. Our data reveal a new noncanonical function of DGCR8 in the modulation of the alternative splicing of Tcf7l1 mRNA in addition to its established function in microRNA biogenesis

Regulation of LINE-1 in mammals

ISSN:1868-503XISSN:1868-502

The Role of RNA Interference in Stem Cell Biology: Beyond the Mutant Phenotypes

Complex gene regulation systems ensure the maintenance of cellular identity during early development in mammals. Eukaryotic small RNAs have emerged as critical players in RNA interference (RNAi) by mediating gene silencing during embryonic stem cell self-renewal. Most of the proteins involved in the biogenesis of small RNAs are essential for proliferation and differentiation into the three germ layers of mouse embryonic stem cells. In the last decade, new functions for some RNAi proteins, independent of their roles in RNAi pathways, have been demonstrated in different biological systems. In parallel, new concepts in stem cell biology have emerged. Here, we review and integrate the current understanding of how RNAi proteins regulate stem cell identity with the new advances in the stem cell field and the recent non-canonical functions of the RNAi proteins. Finally, we propose a reevaluation of all RNAi mutant phenotypes, as non-canonical (small non-coding RNA independent) functions may contribute to the molecular mechanisms governing mouse embryonic stem cells commitment.ISSN:0022-2836ISSN:1089-863

Argonaute 2 Is Required for Extra-embryonic Endoderm Differentiation of Mouse Embryonic Stem Cells

In mouse, although four Argonaute (AGO) proteins with partly overlapping functions in small-RNA pathways exist, only Ago2 deficiency causes embryonic lethality. To investigate the role of AGO2 during mouse early development, we generated Ago2-deficient mouse embryonic stem cells (mESCs) and performed a detailed characterization of their differentiation potential. Ago2 disruption caused a global reduction of microRNAs, which resulted in the misregulation of only a limited number of transcripts. We demonstrated, both in vivo and in vitro, that AGO2 is dispensable for the embryonic germ-layer formation. However, Ago2-deficient mESCs showed a specific defect during conversion into extra-embryonic endoderm cells. We proved that this defect is cell autonomous and can be rescued by both a catalytically active and an inactive Ago2, but not by Ago2 deprived of its RNA binding capacity or by Ago1 overexpression. Overall, our results suggest a role for AGO2 in stem cell differentiation.ISSN:2213-671

Integrative analysis allows a global and precise identification of functional miRNA target genes in mESCs

MicroRNA (miRNA) loaded Argonaute (AGO) complexes regulate gene expression via direct base pairing with their mRNA targets. Previous works suggest that up to 60% of mammalian transcripts might be subject to miRNA-mediated regulation, but it remains largely unknown which fraction of these interactions are functional in a specific cellular context. Here, we integrate transcriptome data from a set of miRNA-depleted mouse embryonic stem cell (mESC) lines with published miRNA interaction predictions and AGO-binding profiles. Using this integrative approach, combined with molecular validation data, we present evidence that < 10% of expressed genes are functionally and directly regulated by miRNAs in mESCs. In addition, analyses of the stem cell-specific miR-290-295 cluster target genes identify TFAP4 as an important transcription factor for early development. The extensive datasets developed in this study will support the development of improved predictive models for miRNA-mRNA functional interactions.ISSN:1469-221XISSN:1469-317