No abstract

Inappropriate stimulation or defective negative regulation of the type I interferon response can lead to autoinflammation. In genetically uncharacterized cases of the type I interferonopathy Aicardi-Goutières syndrome, we identified biallelic mutations in LSM11 and RNU7-1, encoding components of the replication-dependent histone pre-mRNA processing complex. Mutations were associated with the misprocessing of canonical histone transcripts, and a disturbance of linker histone stoichiometry. Additionally, we observed an altered distribution of nuclear cyclic GMP-AMP synthase (cGAS), and enhanced interferon signaling mediated by the cGAS-stimulator of interferon genes (STING) pathway in patient fibroblasts. Finally, we established that chromatin without linker histone more efficiently stimulates cGAS production in vitro. We conclude that nuclear histones, as key constituents of chromatin, are essential in suppressing the immunogenicity of self-DNA.

BackgroundAmyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disease involving loss of motor neurons and having no known cure and uncertain etiology. Several studies have drawn connections between altered retrotransposon expression and ALS. Certain features of the LINE-1 (L1) retrotransposon-encoded ORF1 protein (ORF1p) are analogous to those of neurodegeneration-associated RNA-binding proteins, including formation of cytoplasmic aggregates. In this study we explore these features and consider possible links between L1 expression and ALS.ResultsWe first considered factors that modulate aggregation and subcellular distribution of LINE-1 ORF1p, including nuclear localization. Changes to some ORF1p amino acid residues alter both retrotransposition efficiency and protein aggregation dynamics, and we found that one such polymorphism is present in endogenous L1s abundant in the human genome. We failed, however, to identify CRM1-mediated nuclear export signals in ORF1p nor strict involvement of cell cycle in endogenous ORF1p nuclear localization in human 2102Ep germline teratocarcinoma cells. Some proteins linked with ALS bind and colocalize with L1 ORF1p ribonucleoprotein particles in cytoplasmic RNA granules. Increased expression of several ALS-associated proteins, including TAR DNA Binding Protein (TDP-43), strongly limits cell culture retrotransposition, while some disease-related mutations modify these effects. Using quantitative reverse transcription PCR (RT-qPCR) of ALS tissues and reanalysis of publicly available RNA-Seq datasets, we asked if changes in expression of retrotransposons are associated with ALS. We found minimal altered expression in sporadic ALS tissues but confirmed a previous report of differential expression of many repeat subfamilies in C9orf72 gene-mutated ALS patients.ConclusionsHere we extended understanding of the subcellular localization dynamics of the aggregation-prone LINE-1 ORF1p RNA-binding protein. However, we failed to find compelling evidence for misregulation of LINE-1 retrotransposons in sporadic ALS nor a clear effect of ALS-associated TDP-43 protein on L1 expression. In sum, our study reveals that the interplay of active retrotransposons and the molecular features of ALS are more complex than anticipated. Thus, the potential consequences of altered retrotransposon activity for ALS and other neurodegenerative disorders are worthy of continued investigation.Electronic supplementary materialThe online version of this article (10.1186/s13100-018-0138-z) contains supplementary material, which is available to authorized users.

Nearly half of the human genome is made of transposable elements (TEs) whose activity continues to impact its structure and function. Among them, Long INterspersed Element class 1 (LINE-1 or L1) elements are the only autonomously active TEs in humans. L1s are expressed and mobilized in different cancers, generating mutagenic insertions that could affect tumor malignancy. Tumor suppressor microRNAs are ∼22nt RNAs that post-transcriptionally regulate oncogene expression and are frequently downregulated in cancer. Here we explore whether they also influence L1 mobilization. We show that downregulation of let-7 correlates with accumulation of L1 insertions in human lung cancer. Furthermore, we demonstrate that let-7 binds to the L1 mRNA and impairs the translation of the second L1-encoded protein, ORF2p, reducing its mobilization. Overall, our data reveals that let-7, one of the most relevant microRNAs, maintains somatic genome integrity by restricting L1 retrotransposition.

Here, the role of non-invasive biomarkers in liquid biopsy was evaluated, mainly in exosomes and mitochondrial DNA (mtDNA) as promising, novel, and stable biomarkers for renal cell carcinoma (RCC). A total of 140 fractions (named from B to F) obtained by ultracentrifugations of whole blood samples from 28 individuals (13 patients and 15 controls) were included. Nanoparticle Tracking Analysis (NTA) was conducted to characterized exosomal fraction. Subsequently, an analysis of digital PCR (dPCR) using the QuantStudio™ 3D Digital PCR platform was performed and the quantification of mtDNA copy number by QuantStudioTM 12K Flex Real-Time PCR System (qPCR) was developed. Moreover, Next Generation Sequencing (NGS) analyses were included using MiSeq system (Illumina, San Diego, CA, USA). An F fraction, which contains all exosome data and all mitochondrial markers, was identified in dPCR and qPCR with statistically significant power (adjusted p values ≤ 0.03) when comparing cases and controls. Moreover, present analysis in mtDNA showed a relevant significance in RCC aggressiveness. To sum up, this is the first time a relation between exosomal mtDNA markers and clinical management of RCC is analyzed. We suggest a promising strategy for future liquid biopsy RCC analysis, although more analysis should be performed prior to application in routine clinical practice.

19Nearly half of the human genome is made of transposable elements (TEs) 20 whose activity continues to impact its structure and function. Among them, 21 Long INterspersed Element class 1 (LINE-1 or L1) elements are the only 22 autonomously active TEs in humans. L1s are expressed and mobilized in 23 different cancers, generating mutagenic insertions that could affect 24 malignancy. Tumor suppressor microRNAs are 22nt RNAs that post-25 transcriptionally regulate oncogene expression and are frequently 26 downregulated in cancer. Here we explore whether they also influence L1 27 mobilization. We found that downregulation of let-7 correlates with 28 accumulation of L1 insertions in human lung cancer. Furthermore, we 29 demonstrate that let-7 binds to the L1 mRNA and impairs the translation of the 30 second L1-encoded protein, ORF2p, reducing its mobilization. Overall, our 31 data uncover a new role for let-7, one of the most relevant microRNAs, which 32 is to maintain somatic genome integrity by restricting L1 retrotransposition. 33 34 35 Transposable elements (TEs) account for nearly half of the human genome 1 . 36 However, the only TE that remains autonomously active nowadays is a non-37 Long Terminal Repeat (LTR) retrotransposon known as Long INterspersed 38 Element class 1 (LINE-1 or L1), whose mobilization continues to impact our 39 genome 2 . LINE-1s comprise >20% of our DNA 3 but only about 80-100 of the 40 ~500,000 L1 copies present in the average human genome are full-length 41 elements that retain the ability to mobilize and are thus called 42 Retrotransposition-Competent L1s (RC-L1s) 4 . RC-L1s belong to the human-43 2 specific L1Hs subfamily, are 6kb long and encode two proteins (L1-ORF1p and 44 L1-ORF2p) that are indispensable for retrotransposition 5 . However, ORF2p is 45 expressed at a significantly lower level than ORF1p 6,7 , and these differences are 46 thought to be controlled at the level of translation 8 . L1-ORF1p is a 40kDa RNA 47 binding protein with nucleic acid chaperone activity 9,10 , whereas L1-ORF2p is a 48 150 kDa protein with Endonuclease (EN) and Reverse Transcriptase (RT) 49 activities 11,12 . RC--and-ism, involving 50 reverse transcription of an RNA intermediate and insertion of its cDNA copy at 51 a new site in the genome (reviewed in 2 ). Briefly, retrotransposition starts with 52 the transcription of a full-length RC-L1 bicistronic mRNA, which is exported to 53 the cytoplasm and translated, giving rise to L1-ORF1p and L1-ORF2p that bind 54 preferentially to the same L1 mRNA to form a ribonucleoparticle (RNP) 13 . The 55 RNP gains access to the nucleus where retrotransposition occurs by a 56 mechanism known as Target Primed Reverse Transcription (TPRT) 14,15 . During 57 TPRT, the endonuclease activity of L1-ORF2p nicks the genomic DNA, and its 58 reverse transcriptase activity uses the L1 mRNA as a template to generate a new 59 copy of the element in a different genomic location. L1 can target all regions of 60 the genome, but integration is locally dictated by the presence of...

BackgroundTransposable elements are biologically important components of eukaryote genomes. In particular, non-LTR retrotransposons (N-LTRrs) played a key role in shaping the human genome throughout evolution. In this study, we compared retrotransposon insertions differentially present in the genomes of Anatomically Modern Humans, Neanderthals, Denisovans and Chimpanzees, in order to assess the possible impact of retrotransposition in the differentiation of the human lineage.ResultsWe first identified species-specific N-LTRrs and established their distribution in present day human populations. These analyses shortlisted a group of N-LTRr insertions that were found exclusively in Anatomically Modern Humans. These insertions are associated with an increase in the number of transcriptional/splicing variants of those genes they inserted in. The analysis of the functionality of genes containing human-specific N-LTRr insertions reflects changes that occurred during human evolution. In particular, the expression of genes containing the most recent N-LTRr insertions is enriched in the brain, especially in undifferentiated neurons, and these genes associate in networks related to neuron maturation and migration. Additionally, we identified candidate N-LTRr insertions that have likely produced new functional variants exclusive to modern humans, whose genomic loci show traces of positive selection.ConclusionsOur results strongly suggest that N-LTRr impacted our differentiation as a species, most likely inducing an increase in neural complexity, and have been a constant source of genomic variability all throughout the evolution of the human lineage.Electronic supplementary materialThe online version of this article (10.1186/s13100-018-0133-4) contains supplementary material, which is available to authorized users.

Long Interspersed Element 1 (LINE-1/L1) is an abundant retrotransposon that has greatly impacted human genome evolution. LINE-1s are responsible for the generation of millions of insertions in the current human population. The characterization of sporadic cases of mosaic individuals carrying pathogenic L1-insertions, suggest that heritable insertions occurs during early embryogenesis.However, the timing and potential genomic impact of LINE-1 mobilization during early embryogenesis is unknown. Here, we demonstrate that inner cell mass of human pre-implantation embryos support the expression and retrotransposition of LINE -1s. Additionally, we show that LINE-1s are expressed in trophectoderm cells of embryos , and identify placenta-restricted endogenous LINE-1 insertions in newborns. Using human embryonic stem cells as a model of postimplantation epiblast cells, we demonstrate ongoing LINE-1 retrotransposition, which can impact expression of targeted genes. Our data demonstrate that LINE-1 retrotransposition starts very shortly after fertilization and may represent a previously underappreciated factor in human biology and disease.