Transposable element activation promotes neurodegeneration in a Drosophila model of Huntington's disease

Huntington's disease (HD) is an autosomal dominant disorder with progressive motor dysfunction and cognitive decline. The disease is caused by a CAG repeat expansion in the IT15 gene, which elongates a polyglutamine stretch of the HD protein, Huntingtin. No therapeutic treatments are available, and new pharmacological targets are needed. Retrotransposons are transposable elements (TEs) that represent 40% and 30% of the human and Drosophila genomes and replicate through an RNA intermediate. Mounting evidence suggests that mammalian TEs are active during neurogenesis and may be involved in diseases of the nervous system. Here we show that TE expression and mobilization are increased in a Drosophila melanogaster HD model. By inhibiting TE mobilization with Reverse Transcriptase inhibitors, polyQ-dependent eye neurodegeneration and genome instability in larval brains are rescued and fly lifespan is increased. These results suggest that TE activation may be involved in polyQ-induced neurotoxicity and a potential pharmacological target

A human minisatellite hosts an alternative transcription start site for NPRL3 driving its expression in a repeat number-dependent manner

Minisatellites, also called variable number of tandem repeats (VNTRs), are a class of repetitive elements that may affect gene expression at multiple levels and have been correlated to disease. Their identification and role as expression quantitative trait loci (eQTL) have been limited by their absence in comparative genomic hybridization and single nucleotide polymorphisms arrays. By taking advantage of cap analysis of gene expression (CAGE), we describe a new example of a minisatellite hosting a transcription start site (TSS) which expression is dependent on the repeat number. It is located in the third intron of the gene nitrogen permease regulator like protein 3 (NPRL3). NPRL3 is a component of the GAP activity toward rags 1 protein complex that inhibits mammalian target of rapamycin complex 1 (mTORC1) activity and it is found mutated in familial focal cortical dysplasia and familial focal epilepsy. CAGE tags represent an alternative TSS identifying TAGNPRL3 messenger RNAs (mRNAs). TAGNPRL3 is expressed in red blood cells both at mRNA and protein levels, it interacts with its protein partner NPRL2 and its overexpression inhibits cell proliferation. This study provides an example of a minisatellite that is both a TSS and an eQTL as well as identifies a new VNTR that may modify mTORC1 activity

The prolyl-isomerase PIN1 is essential for nuclear Lamin-B structure and function and protects heterochromatin under mechanical stress

Chromatin organization plays a crucial role in tissue homeostasis. Heterochromatin relaxation and consequent unscheduled mobilization of transposable elements (TEs) are emerging as key contributors of aging and aging-related pathologies, including Alzheimer's disease (AD) and cancer. However, the mechanisms governing heterochromatin maintenance or its relaxation in pathological conditions remain poorly understood. Here we show that PIN1, the only phosphorylation-specific cis/trans prolyl isomerase, whose loss is associated with premature aging and AD, is essential to preserve heterochromatin. We demonstrate that this PIN1 function is conserved from Drosophila to humans and prevents TE mobilization-dependent neurodegeneration and cognitive defects. Mechanistically, PIN1 maintains nuclear type-B Lamin structure and anchoring function for heterochromatin protein 1\u3b1 (HP1\u3b1). This mechanism prevents nuclear envelope alterations and heterochromatin relaxation under mechanical stress, which is a key contributor to aging-related pathologies

Blood transcriptomics of drug-na\uefve sporadic Parkinson's disease patients

BACKGROUND: Parkinson's disease (PD) is a chronic progressive neurodegenerative disorder that is clinically defined in terms of motor symptoms. These are preceded by prodromal non-motor manifestations that prove the systemic nature of the disease. Identifying genes and pathways altered in living patients provide new information on the diagnosis and pathogenesis of sporadic PD. METHODS: Changes in gene expression in the blood of 40 sporadic PD patients and 20 healthy controls ("Discovery set") were analyzed by taking advantage of the Affymetrix platform. Patients were at the onset of motor symptoms and before initiating any pharmacological treatment. Data analysis was performed by applying Ranking-Principal Component Analysis, PUMA and Significance Analysis of Microarrays. Functional annotations were assigned using GO, DAVID, GSEA to unveil significant enriched biological processes in the differentially expressed genes. The expressions of selected genes were validated using RT-qPCR and samples from an independent cohort of 12 patients and controls ("Validation set"). RESULTS: Gene expression profiling of blood samples discriminates PD patients from healthy controls and identifies differentially expressed genes in blood. The majority of these are also present in dopaminergic neurons of the Substantia Nigra, the key site of neurodegeneration. Together with neuronal apoptosis, lymphocyte activation and mitochondrial dysfunction, already found in previous analysis of PD blood and post-mortem brains, we unveiled transcriptome changes enriched in biological terms related to epigenetic modifications including chromatin remodeling and methylation. Candidate transcripts as CBX5, TCF3, MAN1C1 and DOCK10 were validated by RT-qPCR. CONCLUSIONS: Our data support the use of blood transcriptomics to study neurodegenerative diseases. It identifies changes in crucial components of chromatin remodeling and methylation machineries as early events in sporadic PD suggesting epigenetics as target for therapeutic intervention

Effect of dihydroartemisinin (DHA) on human erythroid cell differentiation : implications for malaria treatment in pregnancy

BACKGROUND: Severe malaria in pregnancy causes maternal anemia, low birth weight increased mortality of both mother and infants. WHO recommends few antimalarials due to safety problems. Artemisinin combination therapy is the first line treatment, however artemisinin derivatives showed animal embryotoxicity with a reduction of embryonic erythrocytes when treatment is performed on certain days of gestation. AIMS: To investigate the effect of Dihydroartemisinin (DHA), the metabolite of artemisinins, on an in vitro model reproducing human erythropoiesis and to characterize the erythroid target stage, in order to predict the window of susceptibility to DHA in human pregnancy. METHODS: The mononuclear cells derived from pheripheral blood of healthy volunteers were enriched for CD34+ cells by positive selection using anti-CD34-tagged magnetic beads. CD34+ cells were cultured for 14 days with a specific medium containing erythropoietin to induce erythroid differentiation. DHA at 0,5 or 2 \uc2\ub5M, according to the dosages of previous animal experiments, was added for the first time at day 0 (on isolated stem cell), at day 2 (on early erythroid progenitors), at day 4 (in presence of both early progenitors and pro-erythroblasts), at day 7 (on basophilic erythroblasts) or at day 11 (polychromatic erythroblasts) then continuously every 3 days up to 14 days, because of its short half life. Cells growth and viability were evaluated by trypan blue exclusion; erythroid differentiation was investigated by cytofluorimetric analysis of Glycophorin A (GPA) expression, by morphological analysis on benzidine-May-Grunwald-Giemsa stained smears and by erythroid specific gene expression analysis with real-time PCR. RESULTS: DHA was added on stem cells or early erythroid progenitors caused a transient inhibition of both cell growth and differentiation up to day 7, but then the treated cells started growing and completed their erythroid differentiation at day 14 of culture. When DHA was added on basophilic erythroblasts, a significant and long lasting effect decrease in proliferation as well as a delay in erythroid differentiation was observed. Up to day 14. DHA added on mature stages i.e. polychromatic erythroblasts, only a small reduction of cell growth has been observed without any consequence for the erythroid cell differentiation. CONCLUSIONS: These data suggest that DHA\ue2 s specific target is the basophilic erythroblast, since DHA added at this stage causes a significant inhibition of erythroid differentiation. Based on these in vitro results, we hypothesize that DHA could affect human primitive erythropoiesis, which occurs during the late phase of human secondary yolk sac erythropoiesis (weeks 4-8 of gestation), when foetal blood is formed of only primitive erythroblasts. This means that if the treatment with DHA or artemisinin derivatives is performed during the first trimester of human pregnancy, toxic effects on embryo could be expected

EFFECT OF DIHYDROARTEMISININ (DHA) ON HUMAN ERYTHROID CELL DIFFERENTIATION : IMPLICATIONS FOR MALARIA TREATMENT IN PREGNANCY

Objectives: WHO does not recommend the use of Artemisinin Combination Therapy (ACT) to treat malaria during pregnancy, because animal studies showed a depletion of embryonic erythrocytes. We investigated the effect of Dihydroartemisinin (DHA), the metabolite of artemisinins, on an in vitro model reproducing human erythropoiesis. Methods: CD34+ cells differentiate towards erythroblasts under erythropoietin stimulus in 14 days. DHA, 0,5 or 2 \uf06dM, was added on different erythroid stages. At different time cell growth, morphology, Glycophorin A expression as well as globin genes have been evaluated. Results: DHA added on stem cells or on early progenitors caused a transient inhibitory effect, which was then fully restored. On the contrary, DHA added on more differentiated erythroblasts significantly blocked the erythroid differentiation. This indicates that DHA specifically affects the primitive erythropoiesis, occurring in the yolk sac. Therefore, during the first trimester of pregnancy, ACT must be avoided. EU Antimal Project 18834 is acknowledge

Anti-inflammatory and vasoactive properties of quinoline and artemisinins antimalarials : Modulation of endothelin-1 and cytokines production by microvascular endothelial cells

Severe malaria pathogenesis involves both overproduction of inflammatory cytokines and cytoadherence of parasitized erythrocytes to the microvasculature, leading to local obstruction and hypoxia. Hypoxia and cytokines stimulate endothelial cells to produce endothelin-1 (ET-1), a vasoconstrictor peptide that, in turn, regulates the inflammatory response and, in pregnancy, is involved in pre-eclampsia. Active ET-1 derives from BigEndothelin-1 after processing by endothelin converting enzymes (ECE). To investigate whether, beyond parasite killing, antimalarial drugs could also play an anti-intlammatory role, the effects of quinolines and artemisinins antimalarials on the in vitro production of ET1 and inflammatory cytokines was tested. Human microvascular endothelial cell line (HMEC) treated with quinolines or arremisinin derivatives under normoxia or hypoxia were analyzed for the production of ET-1 and cytokines (IL-6, lL-8) by RT PCR and ELISA. The inhibition of constitutive or hypoxia-induced ET1 secretion by HMEC in the presence of quinolines or artemisinins was dose and time-dependent, and not due to direct toxicity. However, the mechanism of inhibition was different: quinolines, including chloroquine, inhibited the processing by ECE, but not the production, of the precursor Big-ET1 to active ET1. Activity was dependent upon the weak base properties of the drugs. On the contrary, artemisinins inhibited the synthesis of Big-ET1 indicating an effect on transcription. Modulation of cytokine secretion was also observed. These results may explain some of the pharmacological effects of antimalarial drugs such as hypotension after CQ treatment and suggest that their us in pregnancy could help preventing preeclarnpsia