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

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

Extensive somatic L1 retrotransposition in colorectal tumors

L1 retrotransposons comprise 17% of the human genome and are its only autonomous mobile elements. Although L1-induced insertional mutagenesis causes Mendelian disease, their mutagenic load in cancer has been elusive. Using L1-targeted resequencing of 16 colorectal tumor and matched normal DNAs, we found that certain cancers were excessively mutagenized by human-specific L1s, while no verifiable insertions were present in normal tissues. We confirmed de novo L1 insertions in malignancy by both validating and sequencing 69/107 tumor-specific insertions and retrieving both 5′ and 3′ junctions for 35. In contrast to germline polymorphic L1s, all insertions were severely 5′ truncated. Validated insertion numbers varied from up to 17 in some tumors to none in three others, and correlated with the age of the patients. Numerous genes with a role in tumorigenesis were targeted, including ODZ3, ROBO2, PTPRM, PCM1, and CDH11. Thus, somatic retrotransposition may play an etiologic role in colorectal cancer

Targeting the macrophage in equine post-operative ileus

Post-operative ileus (POI) is the functional inhibition of propulsive intestinal motility which is a frequent occurrence following abdominal surgery in the horse and in humans. Rodent and human-derived data have shown that manipulation-induced activation of the resident muscularis externa (ME) macrophages in the intestine contributes to the pathophysiology of the disease. Most studies of the disease, specifically in the horse, have focussed on identification of risk factors, descriptive studies of the disease or the assessment of the efficacy of various therapeutic and prophylactic interventions. As a result, the proposed pathogenesis of equine POI is largely reliant on the translation of data from rodent models. The aims of this thesis were to identify macrophage populations in the normal equine gastrointestinal tract (GIT) and to study equine macrophage activation by stimulating equine bone marrow-derived macrophages (eqBMDMs) with lipopolysaccharide (LPS) as a model for intestinal macrophage activation. Firstly, the normal population of macrophages in the equine GIT was determined. Using CD163 as an immunohistochemical marker for macrophages. CD163+ve cells were present in all tissue layers of the equine intestine: mucosa, submucosa, ME and serosa. CD163+ve cells were regularly distributed within the ME, with accumulations adjacent to the myenteric plexus, and therefore to intestinal motility effector cells such as neurons and the Interstitial Cells of Cajal. The differentiation and survival of intestinal macrophages depends upon signals from the macrophage colony-stimulating factor (CSF-1) receptor. LPS translocation from the gut lumen is thought to be a key activator of ME macrophages. To provide a model for gut macrophages, a protocol was optimised to produce pure populations of equine bone marrow-derived macrophages (eqBMDMs) by cultivation of equine bone marrow in CSF-1. Macrophage functionality was assessed using microscopy, flow cytometry and phagocytosis assays. EqBMDMs responded to LPS stimulation with increases in expression of positive control genes, tumour necrosis factor alpha (TNF-α) and Indoleamine 2,3-dioxygenase (IDO1). The same mRNA was subjected to transcriptomic (RNA-Seq) analysis. Differential gene expression and network cluster analysis demonstrated an inflammatory response characterised by the production of pro-inflammatory cytokines such as interleukin 1 beta (IL-1β) and interleukin 6 (IL-6). However, in contrast to rodent macrophages, eqBMDMs failed to produce nitric oxide in response to LPS, showing species-specific variation in innate immune biology. Using these data, we compared gene expression in normal equine intestine and in intestine from horses undergoing abdominal surgery for colic (abdominal pain). Horses undergoing abdominal surgery showed evidence of increased expression of IL-1β, IL-6 and TNF-α in the mucosa and ME when compared to control tissue. Horses with post-operative reflux (POR), a clinical sign of POI, had increased gene expression of IL-1β, IL-6 and TNF-α compared to horses that did not develop POR following abdominal surgery. These preliminary data suggest that there is macrophage activation within the ME of the intestine during abdominal surgery in the horse, and that a greater activation state is present in horses that subsequently develop POR. The final part of this study was to investigate the effect of a long-acting form of CSF- 1, an Fc fusion protein (CSF1-Fc), as a potential treatment for POI using a mouse model. This work, performed in collaboration with another research group, found that mice lacking the C-C chemokine receptor type 2 (CCR2) gene, which is required for monocyte recruitment into tissues, had a longer recovery period following intestinal manipulation (IM) than wild type (WT) mice. With the administration of CSF1-Fc, infiltration of neutrophils to the ME was reduced and the number of macrophages in the ME was increased in both WT and CCR2-/- mice following IM. Administration of CSF1-Fc in CCR2-/- mice improved recovery of gastrointestinal transit three days following IM, to the same extent as WT mice. Network cluster analysis and RT-qPCR of the ME revealed clusters of genes induced and downregulated by CSF1-Fc, with increased expression of anti-inflammatory and pro-resolving genes after IM in WT and CCR2-/- mice following treatment with CSF1-Fc

Somatic retrotransposition alters the genetic landscape of the human brain

Retrotransposons are mobile genetic elements that use a germline 'copy-and-paste' mechanism to spread throughout metazoan genomes. At least 50 per cent of the human genome is derived from retrotransposons, with three active families (L1, Alu and SVA) associated with insertional mutagenesis and disease. Epigenetic and post-transcriptional suppression block retrotransposition in somatic cells, excluding early embryo development and some malignancies. Recent reports of L1 expression and copy number variation in the human brain suggest that L1 mobilization may also occur during later development. However, the corresponding integration sites have not been mapped. Here we apply a high-throughput method to identify numerous L1, Alu and SVA germline mutations, as well as 7,743 putative somatic L1 insertions, in the hippocampus and caudate nucleus of three individuals. Surprisingly, we also found 13,692 somatic Alu insertions and 1,350 SVA insertions. Our results demonstrate that retrotransposons mobilize to protein-coding genes differentially expressed and active in the brain. Thus, somatic genome mosaicism driven by retrotransposition may reshape the genetic circuitry that underpins normal and abnormal neurobiological processes

Impact of COVID-19 on cardiovascular testing in the United States versus the rest of the world

Objectives: This study sought to quantify and compare the decline in volumes of cardiovascular procedures between the United States and non-US institutions during the early phase of the coronavirus disease-2019 (COVID-19) pandemic. Background: The COVID-19 pandemic has disrupted the care of many non-COVID-19 illnesses. Reductions in diagnostic cardiovascular testing around the world have led to concerns over the implications of reduced testing for cardiovascular disease (CVD) morbidity and mortality. Methods: Data were submitted to the INCAPS-COVID (International Atomic Energy Agency Non-Invasive Cardiology Protocols Study of COVID-19), a multinational registry comprising 909 institutions in 108 countries (including 155 facilities in 40 U.S. states), assessing the impact of the COVID-19 pandemic on volumes of diagnostic cardiovascular procedures. Data were obtained for April 2020 and compared with volumes of baseline procedures from March 2019. We compared laboratory characteristics, practices, and procedure volumes between U.S. and non-U.S. facilities and between U.S. geographic regions and identified factors associated with volume reduction in the United States. Results: Reductions in the volumes of procedures in the United States were similar to those in non-U.S. facilities (68% vs. 63%, respectively; p = 0.237), although U.S. facilities reported greater reductions in invasive coronary angiography (69% vs. 53%, respectively; p < 0.001). Significantly more U.S. facilities reported increased use of telehealth and patient screening measures than non-U.S. facilities, such as temperature checks, symptom screenings, and COVID-19 testing. Reductions in volumes of procedures differed between U.S. regions, with larger declines observed in the Northeast (76%) and Midwest (74%) than in the South (62%) and West (44%). Prevalence of COVID-19, staff redeployments, outpatient centers, and urban centers were associated with greater reductions in volume in U.S. facilities in a multivariable analysis. Conclusions: We observed marked reductions in U.S. cardiovascular testing in the early phase of the pandemic and significant variability between U.S. regions. The association between reductions of volumes and COVID-19 prevalence in the United States highlighted the need for proactive efforts to maintain access to cardiovascular testing in areas most affected by outbreaks of COVID-19 infection

The evolving SARS-CoV-2 epidemic in Africa: Insights from rapidly expanding genomic surveillance

INTRODUCTION Investment in Africa over the past year with regard to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) sequencing has led to a massive increase in the number of sequences, which, to date, exceeds 100,000 sequences generated to track the pandemic on the continent. These sequences have profoundly affected how public health officials in Africa have navigated the COVID-19 pandemic. RATIONALE We demonstrate how the first 100,000 SARS-CoV-2 sequences from Africa have helped monitor the epidemic on the continent, how genomic surveillance expanded over the course of the pandemic, and how we adapted our sequencing methods to deal with an evolving virus. Finally, we also examine how viral lineages have spread across the continent in a phylogeographic framework to gain insights into the underlying temporal and spatial transmission dynamics for several variants of concern (VOCs). RESULTS Our results indicate that the number of countries in Africa that can sequence the virus within their own borders is growing and that this is coupled with a shorter turnaround time from the time of sampling to sequence submission. Ongoing evolution necessitated the continual updating of primer sets, and, as a result, eight primer sets were designed in tandem with viral evolution and used to ensure effective sequencing of the virus. The pandemic unfolded through multiple waves of infection that were each driven by distinct genetic lineages, with B.1-like ancestral strains associated with the first pandemic wave of infections in 2020. Successive waves on the continent were fueled by different VOCs, with Alpha and Beta cocirculating in distinct spatial patterns during the second wave and Delta and Omicron affecting the whole continent during the third and fourth waves, respectively. Phylogeographic reconstruction points toward distinct differences in viral importation and exportation patterns associated with the Alpha, Beta, Delta, and Omicron variants and subvariants, when considering both Africa versus the rest of the world and viral dissemination within the continent. Our epidemiological and phylogenetic inferences therefore underscore the heterogeneous nature of the pandemic on the continent and highlight key insights and challenges, for instance, recognizing the limitations of low testing proportions. We also highlight the early warning capacity that genomic surveillance in Africa has had for the rest of the world with the detection of new lineages and variants, the most recent being the characterization of various Omicron subvariants. CONCLUSION Sustained investment for diagnostics and genomic surveillance in Africa is needed as the virus continues to evolve. This is important not only to help combat SARS-CoV-2 on the continent but also because it can be used as a platform to help address the many emerging and reemerging infectious disease threats in Africa. In particular, capacity building for local sequencing within countries or within the continent should be prioritized because this is generally associated with shorter turnaround times, providing the most benefit to local public health authorities tasked with pandemic response and mitigation and allowing for the fastest reaction to localized outbreaks. These investments are crucial for pandemic preparedness and response and will serve the health of the continent well into the 21st century

Blood from 'junk': the LTR chimeric transcript Pu.2 promotes erythropoiesis

Transposable elements (TEs) are a prominent feature of most eukaryotic genomes. Despite rapidly accumulating evidence for the role of TE-driven insertional mutagenesis and structural variation in genome evolution, few clear examples of individual TEs impacting biology via perturbed gene regulation are available. A recent report describes the discovery of an alternative promoter for the murine erythroid transcription factor Pu.1. This promoter is located in an ORR1A0 long terminal repeat (LTR) retrotransposon intronic to Pu.1 and is regulated by the Krüppel-like factors KLF1 and KLF3. Expression of the resultant chimeric transcript, called Pu.2, spontaneously induces erythroid differentiation in vitro. These experiments illustrate how transcription factor binding sites spread by retrotransposition have the potential to impact networks encoding key biological processes in the host genome

Widespread exaptation of l1 transposons for transcription factor binding in breast cancer

L1 transposons occupy 17% of the human genome and are widely exapted for the regulation of human genes, particularly in breast cancer, where we have previously shown abundant cancer-specific transcription factor (TF) binding sites within the L1PA2 subfamily. In the current study, we performed a comprehensive analysis of TF binding activities in primate-specific L1 subfamilies and identified pervasive exaptation events amongst these evolutionarily related L1 transposons. By motif scanning, we predicted diverse and abundant TF binding potentials within the L1 transposons. We confirmed substantial TF binding activities in the L1 subfamilies using TF binding sites consolidated from an extensive collection of publicly available ChIP-seq datasets. Young L1 subfamilies (L1HS, L1PA2 and L1PA3) contributed abundant TF binding sites in MCF7 cells, primarily via their 5′ UTR. This is expected as the L1 5′ UTR hosts cis-regulatory elements that are crucial for L1 replication and mobilisation. Interestingly, the ancient L1 subfamilies, where 5′ truncation was common, displayed comparable TF binding capacity through their 3′ ends, suggesting an alternative exaptation mechanism in L1 transposons that was previously unnoticed. Overall, primate-specific L1 transposons were extensively exapted for TF binding in MCF7 breast cancer cells and are likely prominent genetic players modulating breast cancer transcriptional regulation.</p