L1 retrotransposition in the soma: a field jumping ahead

Retrotransposons are transposable elements (TEs) capable of "jumping" in germ, embryonic and tumor cells and, as is now clearly established, in the neuronal lineage. Mosaic TE insertions form part of a broader landscape of somatic genome variation and hold significant potential to generate phenotypic diversity, in the brain and elsewhere. At present, the LINE-1 (L1) retrotransposon family appears to be the most active autonomous TE in most mammals, based on experimental data obtained from disease-causing L1 mutations, engineered L1 reporter systems tested in cultured cells and transgenic rodents, and single-cell genomic analyses. However, the biological consequences of almost all somatic L1 insertions identified thus far remain unknown. In this review, we briefly summarize the current state-of-the-art in the field, including estimates of L1 retrotransposition rate in neurons. We bring forward the hypothesis that an extensive subset of retrotransposition-competent L1s may be de-repressed and mobile in the soma but largely inactive in the germline. We discuss recent reports of non-canonical L1-associated sequence variants in the brain and propose that the elevated L1 DNA content reported in several neurological disorders may predominantly comprise accumulated, unintegrated L1 nucleic acids, rather than somatic L1 insertions. Finally, we consider the main objectives and obstacles going forward in elucidating the biological impact of somatic retrotransposition

Is somatic retrotransposition a parasitic or symbiotic phenomenon?

The extraordinary evolutionary success of transposable elements (TEs) invites us to question the nature of the co-evolutionary dynamics between TE and host. Although sometimes assumed to be wholly parasitic, TEs have penetrated and spread throughout eukaryotic genomes at a rate unparalleled by other parasites. This near-ubiquity, occurring despite the potentially deleterious effects of insertional mutagenesis, raises the possibility that a counterbalancing benefit exists for the host. Such a benefit may act at the population level to generate genomic diversity within a species and hence greater adaptability under new selective pressures, or at the level of primary gain for the individual. Recent studies have highlighted the occurrence of retrotransposition events in the germline and discovered a surprisingly high rate of mobilization in somatic cells. Here we examine the available evidence for somatic retrotransposition and discuss how this phenomenon may confer a selective advantage upon an individual or species

Retrotransposon-induced mosaicism in the neural genome

Over the past decade, major discoveries in retrotransposon biology have depicted the neural genome as a dynamic structure during life. In particular, the retrotransposon LINE-1 (L1) has been shown to be transcribed and mobilized in the brain. Retrotransposition in the developing brain, as well as during adult neurogenesis, provides a milieu in which neural diversity can arise. Dysregulation of retrotransposon activity may also contribute to neurological disease. Here, we review recent reports of retrotransposon activity in the brain, and discuss the temporal nature of retrotransposition and its regulation in neural cells in response to stimuli. We also put forward hypotheses regarding the significance of retrotransposons for brain development and neurological function, and consider the potential implications of this phenomenon for neuropsychiatric and neurodegenerative conditions

RNA-MATE: a recursive mapping strategy for high-throughput RNA-sequencing data

Summary: Mapping of next-generation sequencing data derived from RNA samples (RNAseq) presents different genome mapping challenges than data derived from DNA. For example, tags that cross exon-junction boundaries will often not map to a reference genome, and the strand specificity of the data needs to be retained. Here we present RNA-MATE, a computational pipeline based on a recursive mapping strategy for placing strand specific RNAseq data onto a reference genome. Maximizing the mappable tags can provide significant savings in the cost of sequencing experiments. This pipeline provides an automatic and integrated way to align color-space sequencing data, collate this information and generate files for examining gene-expression data in a genomic context

Genetic control of the innate immune response

BACKGROUND: Susceptibility to infectious diseases is directed, in part, by the interaction between the invading pathogen and host macrophages. This study examines the influence of genetic background on host-pathogen interactions, by assessing the transcriptional responses of macrophages from five inbred mouse strains to lipopolysaccharide (LPS), a major determinant of responses to gram-negative microorganisms. RESULTS: The mouse strains examined varied greatly in the number, amplitude and rate of induction of genes expressed in response to LPS. The response was attenuated in the C3H/HeJlps(d )strain, which has a mutation in the LPS receptor Toll-like receptor 4 (TLR4). Variation between mouse strains allowed clustering into early (C57Bl/6J and DBA/2J) and delayed (BALB/c and C3H/ARC) transcriptional phenotypes. There was no clear correlation between gene induction patterns and variation at the Bcg locus (Slc11A1) or propensity to bias Th1 versus Th2 T cell activation responses. CONCLUSION: Macrophages from each strain responded to LPS with unique gene expression profiles. The variation apparent between genetic backgrounds provides insights into the breadth of possible inflammatory responses, and paradoxically, this divergence was used to identify a common transcriptional program that responds to TLR4 signalling, irrespective of genetic background. Our data indicates that many additional genetic loci control the nature and the extent of transcriptional responses promoted by a single pathogen-associated molecular pattern (PAMP), such as LPS

Nanopore Sequencing Enables Comprehensive Transposable Element Epigenomic Profiling

Transposable elements (TEs) drive genome evolution and are a notable source of pathogenesis, including cancer. While CpG methylation regulates TE activity, the locus-specific methylation landscape of mobile human TEs has to date proven largely inaccessible. Here, we apply new computational tools and long-read nanopore sequencing to directly infer CpG methylation of novel and extant TE insertions in hippocampus, heart, and liver, as well as paired tumor and non-tumor liver. As opposed to an indiscriminate stochastic process, we find pronounced demethylation of young long interspersed element 1 (LINE-1) retrotransposons in cancer, often distinct to the adjacent genome and other TEs. SINE-VNTR-Alu\ua0(SVA) retrotransposons, including their internal tandem repeat-associated CpG island, are near-universally methylated. We encounter allele-specific TE methylation and demethylation of aberrantly expressed young LINE-1s in normal tissues. Finally, we recover the complete sequences of tumor-specific LINE-1 insertions and their retrotransposition hallmarks, demonstrating how long-read sequencing can simultaneously survey the epigenome and detect somatic TE mobilization

Visualization and analysis of RNA-Seq assembly graphs.

RNA-Seq is a powerful transcriptome profiling technology enabling transcript discovery and quantification. Whilst most commonly used for gene-level quantification, the data can be used for the analysis of transcript isoforms. However, when the underlying transcript assemblies are complex, current visualization approaches can be limiting, with splicing events a challenge to interpret. Here, we report on the development of a graph-based visualization method as a complementary approach to understanding transcript diversity from short-read RNA-Seq data. Following the mapping of reads to a reference genome, a read-to-read comparison is performed on all reads mapping to a given gene, producing a weighted similarity matrix between reads. This is used to produce an RNA assembly graph, where nodes represent reads and edges similarity scores between them. The resulting graphs are visualized in 3D space to better appreciate their sometimes large and complex topology, with other information being overlaid on to nodes, e.g. transcript models. Here we demonstrate the utility of this approach, including the unusual structure of these graphs and how they can be used to identify issues in assembly, repetitive sequences within transcripts and splice variants. We believe this approach has the potential to significantly improve our understanding of transcript complexity

Evidence for L1-associated DNA rearrangements and negligible L1 retrotransposition in glioblastoma multiforme

Background: LINE-1 (L1) retrotransposons are a notable endogenous source of mutagenesis in mammals. Notably, cancer cells can support unusual L1 retrotransposition and L1-associated sequence rearrangement mechanisms following DNA damage. Recent reports suggest that L1 is mobile in epithelial tumours and neural cells but, paradoxically, not in brain cancers. Results: Here, using retrotransposon capture sequencing (RC-seq), we surveyed L1 mutations in 14 tumours classified as glioblastoma multiforme (GBM) or as a lower grade glioma. In four GBM tumours, we characterised one probable endonuclease-independent L1 insertion, two L1-associated rearrangements and one likely Alu-Alu recombination event adjacent to an L1. These mutations included PCR validated intronic events in MeCP2 and EGFR. Despite sequencing L1 integration sites at up to 250× depth by RC-seq, we found no tumour-specific, endonuclease-dependent L1 insertions. Whole genome sequencing analysis of the tumours carrying the MeCP2 and EGFR L1 mutations also revealed no endonuclease-dependent L1 insertions. In a complementary in vitro assay, wild-type and endonuclease mutant L1 reporter constructs each mobilised very inefficiently in four cultured GBM cell lines. Conclusions: These experiments altogether highlight the consistent absence of canonical L1 retrotransposition in GBM tumours and cultured cell lines, as well as atypical L1-associated sequence rearrangements following DNA damage in vivo

Genome-wide methylated CpG island profiles of melanoma cells reveal a melanoma coregulation network

Metastatic melanoma is a malignant cancer with generally poor prognosis, with no targeted chemotherapy. To identify epigenetic changes related to melanoma, we have determined genome-wide methylated CpG island distributions by next-generation sequencing. Melanoma chromosomes tend to be differentially methylated over short CpG island tracts. CpG islands in the upstream regulatory regions of many coding and noncoding RNA genes, including, for example, TERC, which encodes the telomerase RNA, exhibit extensive hypermethylation, whereas several repeated elements, such as LINE 2, and several LTR elements, are hypomethylated in advanced stage melanoma cell lines. By using CpG island demethylation profiles, and by integrating these data with RNA-seq data obtained from melanoma cells, we have identified a co-expression network of differentially methylated genes with significance for cancer related functions. Focused assays of melanoma patient tissue samples for CpG island methylation near the noncoding RNA gene SNORD-10 demonstrated high specificity

HCV activates somatic L1 retrotransposition–A potential hepatocarcinogenesis pathway

Hepatitis C virus (HCV) is a common cause of hepatocellular carcinoma (HCC). The activation and mutagenic consequences of L1 retrotransposons in virus-associated-HCC have been documented. However, the direct influence of HCV upon L1 elements is unclear, and is the focus of the present study. L1 transcript expression was evaluated in a publicly available liver tissue RNA-seq dataset from patients with chronic HCV hepatitis (CHC), as well as healthy controls. L1 transcript expression was significantly higher in CHC than in controls. L1orf1p (a L1 encoded protein) expression was observed in six out of 11 CHC livers by immunohistochemistry. To evaluate the influence of HCV on retrotransposition efficiency, in vitro engineered-L1 retrotransposition assays were employed in Huh7 cells in the presence and absence of an HCV replicon. An increased retrotransposition rate was observed in the presence of replicating HCV RNA, and persisted in cells after viral clearance due to sofosbuvir (PSI7977) treatment. Increased retrotransposition could be due to dysregulation of the DNA-damage repair response, including homologous recombination, due to HCV infection. Altogether these data suggest that L1 expression can be activated before oncogenic transformation in CHC patients, with HCV-upregulated retrotransposition potentially contributing to HCC genomic instability and a risk of transformation that persists post-viral clearance