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Rôle des corps nucléaires PML et des chaperons de l’histone H3.3 dans la chromatinisation du génome du virus Herpès Simplex 1 pendant la latence / Role of PML Nuclear Bodies and H3.3 chaperones in Herpes Simplex Virus 1 genomes chromatinization during latencyCohen, Camille 20 October 2017 (has links)
L'établissement de latence du virus de l'Herpès simplex 1 (HSV1) est contrôlé par les corps nucléaires PML (PML-NBs) mais leur implication exacte reste encore confuse. Une des caractéristiques majeures de la latence du virus est l'interaction entre le génome viral et les PML-NBs formant des structures nommées viral DNA-containing PML-NBs (vDCP-NBs). L'utilisation d'un modèle d'infection de fibroblastes primaires humains, qui reproduit la formation des vDCP-NBs, combinée à une approche par immuno-FISH, a permis de montrer que les vDCP-NBs contiennent l'histone H3.3 et ses chaperons, les complexes DAXX-ATRX et HIRA. La protéine HIRA a été également observé au sein des vDCP-NBs dans les neurones des ganglions trijumeaux de souris infectées par HSV1. Des expériences de ChIP-qPCR dans des cellules exprimant H3.3 ou H3.1 tagguées, nous a permis de déterminer que le génome viral est spécifiquement chromatinisé avec l'histone H3.3. La déplétion d'une seule protéine des complexes chaperons de H3.3 affecte légèrement l'incorporation de H3.3 dans les génomes viraux latents. Au contraire, l'absence de PML diminue significativement la chromatinisation H3.3 du génome HSV-1 latent sans remplacement par H3.1. Cette étude démontre une régulation épigénétique du génomes HSV1 latent par une chromatinisation dépendante de H3.3 impliquant les complexes chaperons DAXX-ATRX et HIRA. De plus, cette étude révèle un rôle majeur des PML-NBs dans la chromatinisation H3.3 des génomes HSV1 latent / Herpes simplex virus 1 (HSV-1) latency establishment is tightly controlled by PML nuclear bodies (PML-NBs) although their exact implication is still elusive. A hallmark of HSV-1 latency is the interaction between latent viral genomes and PML-NBs leading to the formation of viral DNA-containing PML-NBs (vDCP-NBs). Using a replication defective HSV-1 infected human primary fibroblast model reproducing the formation of vDCP-NBs, combined with an IF-FISH approach developed to detect latent HSV-1, we show that vDCP-NBs contain both histone H3.3 and its chaperone complexes, i.e. the DAXX/ATRX and the HIRA complex. HIRA was also detected co-localizing with vDCP-NBs present in trigeminal ganglia neurons from HSV-1 infected WT mice. ChIP-qPCR performed on fibroblasts stably expressing tagged H3.3 or H3.1 show that latent HSV1 genomes are chromatinized almost exclusively with H3.3. Depletion of single proteins from the H3.3 chaperone complexes only mildly affects H3.3 deposition on the latent HSV1 genome. In contrast, absence of PML significantly impacts on the chromatinization of the latent genomes with H3.3 without replacement with H3.1. Consequently, the study demonstrates a specific epigenetic regulation of latent HSV-1 through an H3.3-dependent HSV-1 chromatinization involving both H3.3 chaperones DAXX/ATRX and HIRA complexes. Additionally, the study reveals that PML-NBs are major actors of the latent HSV-1 H3.3 chromatinization through a PML-NBs/histone H3.3/H3.3 chaperones axis
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Role metody PCR v diagnostice neuroinfekcí vyvolaných herpetickými viry / Diagnostics of neuroinfection caused by human herpesviruses using nucleic acid amplification methodsLabská, Klára January 2021 (has links)
of thesis Diagnostics of neuroinfection caused by human herpesviruses using nucleic acid amplification methods author: MUDr. Klára Labská supervisor: doc. MUDr. Vilma Marešová, CSc. In recent years, the diagnosis of neuroinfections has undergone a shift towards molecular biology methods. Our research focused on the predictive value of the capture of herpesvirus (HV) DNA in cerebrospinal fluid. In the first study, we examined the presence of DNA neurotropic herpes viruses (HSV1, HSV2, VZV and HHV6) in cerebrospinal fluid in immunocompetent patients with laboratory-confirmed tick-borne meningoencephalitis and enterovirus meningitis and meningoencephalitis. The control group consisted of patients with proven absence of an inflammation in the cerebrospinal fluid. Patients were followed for 6 months. The course of the disease and its consequences, including laboratory tests, were compared between groups of patients with and without the presence of HV DNA. In the second study, we tried to demonstrate the presence of HSV1 DNA in cerebrospinal fluid during its symptomatic reactivation in patients with purulent meningitis. In our group of immunocompetent patients with non-purulent inflammation in the cerebrospinal fluid, the proportion of HV DNA positive patients reached 7.5% (13 out of 173), we also...
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