Swine influenza: Epidemiological situation in France

Serological and virus identification studies have been carried out in France to assess the epidemiological situation of the swine population. At the end of the 70’s, the serological profile in pig farms was dominated by the presence of A/H3N2 antibodies, associated with epidemics of human influenza. Since then, epizootic outbreaks have succeeded one another in the pig. The disease is now both enzootic and epizootic. Since the early 2000s, swine influenza in France occurs mainly in Brittany, where pig density is the highest. Its economic impact is considerable in pig farms of that area. The disease is caused by the influenza A/H1 virus of avian origin (A/H1N1) or by reassortants (A/H1N2). As influenza viruses are unstable, detection tools need permanent updating to guarantee an effective epidemiological surveillance.La situation épidémiologique du cheptel porcin français est appréhendée au travers d'études sérologiques ainsi que de recherches virales. À la fin des années 1970,le profil sérologique des élevages est dominé par la présence d'anticorps A/H3N2 correspondant à des épidémies de grippe humaine. Par la suite, des vagues épizootiques ont déferlé. La maladie se présente désormais sous une forme enzootique et épizootique. Depuis le début des années 2000, la grippe du porc en France concerne avant tout les élevages de Bretagne, où la densité porcine est la plus élevée. Elle a un impact économique considérable dans les élevages de cette région. L'activité grippale est le fait de virus A/H1 d'origine aviaire (A/H1N1) ou de réassortants (A/H1N2). L'instabilité des virus grippaux suppose d'adapter régulièrement les outils de détection afin de permettre une épidémiosurveillance efficace

Interaction of HP1 and Brg1/Brm with the Globular Domain of Histone H3 Is Required for HP1-Mediated Repression

The heterochromatin-enriched HP1 proteins play a critical role in regulation of transcription. These proteins contain two related domains known as the chromo- and the chromoshadow-domain. The chromo-domain binds histone H3 tails methylated on lysine 9. However, in vivo and in vitro experiments have shown that the affinity of HP1 proteins to native methylated chromatin is relatively poor and that the opening of chromatin occurring during DNA replication facilitates their binding to nucleosomes. These observations prompted us to investigate whether HP1 proteins have additional histone binding activities, envisioning also affinity for regions potentially occluded by the nucleosome structure. We find that the chromoshadow-domain interacts with histone H3 in a region located partially inside the nucleosomal barrel at the entry/exit point of the nucleosome. Interestingly, this region is also contacted by the catalytic subunits of the human SWI/SNF complex. In vitro, efficient SWI/SNF remodeling requires this contact and is inhibited in the presence of HP1 proteins. The antagonism between SWI/SNF and HP1 proteins is also observed in vivo on a series of interferon-regulated genes. Finally, we show that SWI/SNF activity favors loading of HP1 proteins to chromatin both in vivo and in vitro. Altogether, our data suggest that HP1 chromoshadow-domains can benefit from the opening of nucleosomal structures to bind chromatin and that HP1 proteins use this property to detect and arrest unwanted chromatin remodeling

Epigenetic component in the onset of inflammation in the context of multiple sclerosis

La sclérose en plaques est une maladie auto-immune dirigée contre les protéines de la myéline du cerveau. Plusieurs mécanismes physiopathologiques sont impliqués dans la SEP tels que l'inflammation, la démyélinisation et l’atteinte axonale. La SEP est associée à une expression accrue de cytokines et à une activation des rétrovirus endogènes humains (HERVs). Dans les conditions physiologiques, ces unités transcriptionnelles sont maintenues dans un état réprimé par un même mécanisme répresseur dépendant de la chromatine : la tri-méthylation de la lysine 9 de l’histone H3 (H3K9me3), qui crée un site de liaison aux protéines de la famille HP1. Nous avons trouvé qu’à la fois, les gènes de l’immunité et les HERVs nécessitent les protéines hétérochromatiniennes HP1α pour leur répression transcriptionnelle. Nous avons montré que la peptidylarginine déiminase 4, une enzyme qui joue un rôle dans la SEP, affaiblit la liaison de HP1α à la lysine 9 tri-méthylée de l'histone H3 en citrullinant l’arginine 8. Nous avons apporté la preuve que de multiples événements de la réactivation de la transcription chez les patients atteints de la SEP peuvent être expliqués par un défaut du mécanisme unique de répression génique. Nous avons également montré qu'il est possible de renforcer la répression de HP1 à l'aide de petites molécules. Par exemple, l’EGCG, un composé de thé vert, est en mesure de réduire à la fois l’expression des HERVs et des cytokines en augmentant l'activité de l’histone méthyltransférase SUV39H1. Cela conduit à une accumulation de la marque répressive H3K9me3, qui va favoriser la liaison de HP1.Ensemble, ces résultats suggèrent que HP1 est une composante importante de la SEP au niveau de la régulation des cytokines et des HERVs.Multiple Sclerosis is an autoimmune disease resulting in damage to myelin structures of the brain. Several physiopathological mechanisms are involved in MS including inflammation, demyelination, and axonal damage. MS is associated with increased cytokine expression and activation of human endogenous retroviruses (HERVs). These two types of transcriptional units are kept in check by chromatin-dependent silencing associated with lysine 9 trimethylation of histone H3, and subsequent of HP1 proteins. We find that both the cytokine genes and the HERVs require the heterochromatin protein HP1 for their transcriptional repression. Furthermore, we have shown that the peptidylarginine deiminase 4, an enzyme with a suspected role in MS, weakens the binding of HP1 to tri-methylated histone H3 lysine 9 by citrullinating histone H3 arginine 8. We thereby evidence that multiple events of transcriptional reactivation in MS patients can be explained by deficiency of a single mechanism of gene silencing. We have also shown that it is possible to reinforce HP1 repression by using small molecules. For example, EGCG, a green tea compound, is able to reduce both HERVs and cytokines expression by increasing histone methyltransferase activity SUV39H1. This leads to increased accumulation of H3K9me3 repressive marks and favors binding of HP1. All together, these results suggest that HP1 is an important component of the regulation of cytokine genes and HERVs in MS patients

Expression of endogenous retroviruses reflects increased usage of atypical enhancers in T cells

International audienceSeveral autoimmune diseases including multiple sclerosis (MS) cause increased transcription of endogenous retroviruses (HERVs) normally repressed by heterochromatin. In parallel, HERV-derived sequences were reported to drive gene expression. Here, we have examined a possible link between promoter and enhancer divergent transcription and the production of HERV transcripts. We find that HERV-derived sequences are in general counter-selected at regulatory regions, a counter-selection that is strongest in brain tissues while very moderate in stem cells. By exposing T cells to the pesticide dieldrin, we further found that a series of HERV-driven enhancers otherwise active only at stem cell stages can be reactivated by stress. This in part relies on peptidylarginine deiminase activity, possibly participating in the reawakening of silenced enhancers. Likewise, usage of HERV-driven enhancers was increased in myelin-reactive T cells from patients with MS, correlating with activation of nearby genes at several sites. Altogether, we propose that HERV-driven enhancers constitute a reservoir of auxiliary enhancers transiently induced by stress while chronically active in diseases like MS

Citrullination of Histone H3 Interferes with HP1-Mediated Transcriptional Repression

International audienceMultiple Sclerosis (MS) is an autoimmune disease associated with abnormal expression of a subset of cytokines, resulting in inappropriate T-lymphocyte activation and uncontrolled immune response. A key issue in the field is the need to understand why these cytokines are transcriptionally activated in the patients. Here, we have examined several transcription units subject to pathological reactivation in MS, including the TNFa and IL8 cytokine genes and also several Human Endogenous RetroViruses (HERVs). We find that both the immune genes and the HERVs require the heterochromatin protein HP1a for their transcriptional repression. We further show that the Peptidylarginine Deiminase 4 (PADI4), an enzyme with a suspected role in MS, weakens the binding of HP1a to tri-methylated histone H3 lysine 9 by citrullinating histone H3 arginine 8. The resulting de-repression of both cytokines and HERVs can be reversed with the PADI-inhibitor Cl-amidine. Finally, we show that in peripheral blood mononuclear cells (PBMCs) from MS patients, the promoters of TNFa, and several HERVs share a deficit in HP1a recruitment and an augmented accumulation of histone H3 with a double citrulline 8 trimethyl lysine 9 modifications. Thus, our study provides compelling evidence that HP1a and PADI4 are regulators of both immune genes and HERVs, and that multiple events of transcriptional reactivation in MS patients can be explained by the deficiency of a single mechanism of gene silencing

Broad spectrum compounds targeting early stages of rabies virus (RABV) infection

International audienceABMA and its analogue DABMA are two molecules of the adamantane family known to perturbate the endosomal pathway and to inhibit cell infection of several RNA and DNA viruses. Their activity against Rabies Virus (RABV) infection has already been demonstrated in vitro. (Wu et al., 2017, 2019). Here, we describe in more details their mechanism of action by comparison to Arbidol (umifenovir) and Ribavirin, two broad spectrum antivirals against emerging viruses such as Lassa, Ebola, influenza and Hantaan viruses. ABMA and DABMA, delivered 2 h pre-infection, inhibit RABV infection in vitro with an EC50 of 7.8 μM and 14 μM, respectively. They act at post-entry, by causing RABV accumulation within the endosomal compartment and DABMA specifically diminishes the expression of the GTPase Rab7a controlling the fusion of early endosomes to late endosomes or lysosomes. This may suggest that ABMA and DABMA act at different stages of the late endosomal pathway as supported by their different profile of synergy/antagonism with the fusion inhibitor Arbidol. This difference is further confirmed by the RABV mutants induced by successive passages under increasing selective pressure showing a particular involvement of the viral G protein in the DABMA inhibition while ABMA inhibition induces less mutations dispersed in the M, G and L viral proteins. These results suggest new therapeutic perspectives against rabies

Model.

<p>HP1α silences HERVs and represses several cytokines until they are stimulated. It binds the tri-methylated lysine 9 of histone H3 on these promoters.This binding is in part regulated by PADI4 conversion of histone H3 arginine 8 into a citrulline, thereby reducing the affinity of HP1α for the neighboring methylated lysine 9. In MS patients, PADI4 induces an enrichment of H3cit8K9me3 double histone modification resulting in reduced accumulation of HP1α on HERVs, on the promoter of TNFα and possibly on other cytokines genes. This in turn may be responsible for excessive expression of HERVs and cytokines that inappropriately activate T lymphocytes and ultimately damages the central nervous system. It may be possible to interrupt this sequence of event with the PADI4 specific inhibitor Cl-amidine.</p

PADI4 activity controls HP1α occupancy and histone H3 citrullination at TNFα and HERV promoters.

<p>(A) Total protein extracts from MCF7 cells treated either estradiol (E2) or ionophore (A23187) and/or Cl-amidine were immunoblotted with the indicated antibodies. (B) Total RNA from MCF7 cells either untreated or treated with estradiol followed by ionophore (E2+A23187) and/or Cl-amidine (E2+A23187+Cl-amidine) treatment was quantified by RT-qPCR. Data are shown relative to the un-induced condition (set to 1). Values are mean ± SEM from four experimental replicates. (C–E) ChIP with the listed antibodies was carried out with chromatin from MCF7 cells either untreated or treated with estradiol and ionophore (E2+A23187) in the absence or presence of Cl-amidine. The relative enrichments of the indicated antibodies on the shown LTRs or promoters were measured by qPCR. Data are presented as a percentage of histone H3 or relative to non-immune IgG as indicated. Enrichments are presented relative to indicated controls (set to 1). Values are means ± SEM from four PCR measures of representative ChIP experiments.</p

The H3cit8K9me3 double histone modification reduces the affinity of HP1α for the H3K9me3 single modification.

<p>(A) Dot blot showing that HP1α does not bind to histone H3 peptides carrying a H3Cit8K9me3 double modification. Indicated histone H3 peptides of 1, 0.2, and 0.04 µg were spotted on the membranes. Then binding of recombinant GST-HP1α was tested by labeling of the retained protein with anti-GST antibodies. Blots are representative of the experimental replicates. (B) Purified recombinant GST-HP1α (red) was bound to fixed MCF7 cells in “overlay” assays, and then challenged by competition with 1 µg of the indicated peptides. DNA is labeled with DAPI (blue), scale bar: 5 µm. (C) Real-time association and dissociation surface plasmon resonance (SPR) profiles corresponding to the injection of the indicated H3 peptides at 12.5 µM over immobilized GST-HP1α. (D) Percentage of bound GST-HP1α sites as a function of the peptide concentration. (E–F) Total RNA from MCF7 cells transfected with the PADI4 small interfering RNAs (siRNAs) was quantified with RT-qPCR. Changes in mRNA levels are shown relative to the siGAPDH transfection (set to 1), which was not affecting the mRNA levels of the genes of interest (<a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1002934#pgen.1002934.s001" target="_blank">Figure S1C</a>). The data are presented as the means ± SEM of triplicate experiments.</p