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Title:Vpliv proteinov APOBEC na infekcijo z virusi HPV
Authors:Lojpur, Tjaša (Author)
Bergant Marušič, Martina (Mentor) More about this mentor... New window
Files:.pdf Tjasa_Lojpur.pdf (1,64 MB)
 
Language:Slovenian
Work type:Master's thesis/paper (mb22)
Tipology:2.09 - Master's Thesis
Organization:FZO - Faculty of Environmental Sciences
Abstract:Humani virusi papiloma (HPV) so povzročitelji najpogostejše spolno prenosljive okužbe, povezani pa so tudi z razvojem raka materničnega vratu. Proteini APOBEC so DNA/RNA editirajoči encimi, ki igrajo ključno vlogo pri prirojenih protivirusnih imunskih odzivih in so vpleteni tako v infekcijo s HPV kot tudi v razvoj rakavih obolenj pri dolgotrajni okužbi s HPV. Glavni cilj magistrskega dela je bil proučiti protivirusno delovanje proteinov APOBEC in določiti tudi ožji nabor proteinov APOBEC, vključenih v protivirusni odziv pri infekciji s HPV. Vpliv proteinov APOBEC na primarno infekcijo smo ugotavljali z uporabo psevdovirusnih delcev (PsV) HPV, v plašču katerih je bil vgrajen reporterski gen za luciferazo. Stopnjo infekcije virusov HPV v gostiteljskih celičnih linijah, transficiranih s proteini hAPOBEC, smo določali posredno preko encimske aktivnosti luciferaze. Ugotovili smo, da izražanje večine proteinov hAPOBEC zniža infektivnost PsV HPV. Izmed proučevanih proteinov hAPOBEC so imeli najmočnejši protivirusni odziv hAID, hA1, hA2, hA3A, hA3B, hA3C in hA3DE, saj so zmanjšali infektivnost PsV HPV-16 za več kot 50 %. Čeprav smo protivirusni učinek proteinov hAPOBEC ugotovili tudi pri dveh kožnih tipih, HPV-2 in HPV-5, je bil le ta značilno višji pri visokorizičnem sluzničnem tipu HPV-16. Mehanizme delovanja proteinov hAPOBEC smo ugotavljali s katalitskimi in nekatalitskimi mutantami proteinov hA3A (E72A) in hA3B (E255Q in E68Q), ki so za razliko od divjih tipov, infektivnost HPV-16 povečale. Mikroskopska analiza znotrajcelične lokalizacije izbranih proteinov hAPOBEC med infekcijo s HPV-16 pa je pokazala, da se 24 ur po infekciji zgodi premik deleža proteina hA3A iz citoplazme v jedro, kar bi lahko vplivalo na potek infekcije. V magistrskem delu smo potrdili prvotno hipotezo, da proteini APOBEC sodelujejo pri protivirusnem odzivu virusov HPV, njihov protivirusni učinek pa je odvisen tako od vrste proteina APOBEC kot tudi od tipa virusa HPV.
Keywords:HPV, proteini APOBEC, protivirusni odziv, infektivnost, znotrajcelična lokalizacija
Year of publishing:2018
Source:Nova Gorica
COBISS_ID:5263611 Link is opened in a new window
URN:URN:SI:UNG:REP:X0EL7E7Z
Views:687
Downloads:49
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Secondary language

Language:English
Title:Effect of APOBEC proteins on the HPV infection
Abstract:Human papillomaviruses (HPVs) cause the most common sexually transmitted infection, but they are also involved in the development of cervical cancer. APOBEC proteins are DNA/RNA editing enzymes, which play a key role in innate antiviral immune responses. They are involved in both, HPV infection and the development of cancerous diseases in case of persistent HPV infection. The main aim of this master’s thesis was to study the antiviral function of APOBEC proteins and to determine shorter set of APOBEC proteins, involved in the antiviral response in HPV infection. The effect of APOBEC proteins on primary infection had been studied using HPV pseudovirions (PsVs), carrying a luciferase reporter gene. The infectivity rate of HPVs in host cell lines transfected with hAPOBEC proteins was determined indirectly, by the luciferase enzyme activity. We found out that expression of most hAPOBEC proteins reduces infectivity of HPV PsVs. Among the studied hAPOBEC proteins, hAID, hA1, hA2, hA3A, hA3B, hA3C and hA3DE were those with the strongest antiviral response, as they reduced infectivity of HPV-16 by more than 50 %. Although the antiviral effect of hAPOBEC proteins was also found in two cutaneous HPV types, namely HPV-2 and HPV-5, such effect was significantly higher in the high-risk mucosal type – HPV-16. Mechanisms of hAPOBEC proteins' function were established using catalytic and non-catalytic mutants of hA3A (E72A) and hA3B (E255Q and E68Q) proteins which, unlike wild types, increased the infectivity of HPV-16. Microscopic analysis of the intracellular localization of selected hAPOBEC proteins during HPV-16 infection showed that 24 hours post infection the displacement of the hA3A from the cytoplasm to the nucleus occurs which could affect the process of infection. In this master's thesis we confirmed that APOBEC proteins participate in the antiviral response of HPV viruses and that their antiviral effect depends on both, type of APOBEC protein and the type of HPV virus, which was our initial hypothesis.
Keywords:HPV, APOBEC proteins, antiviral response, infectivity, intracellular localization


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