Solusyklin G2/M-vaiheen säätelyn vaikutus koiran parvoviruksen tumasta vapautumiseen

Koiran parvovirus (engl. canine parvovirus, CPV) on pieni (halkaisijaltaan ~25 nm), vaipaton protoparvovirus. Sillä on ikosahedraalinen kapsidi joka ympäröi noin 5 kb pitkää, yksijuosteista DNA -genomia. CPV kulkeutuu solun sisään klatriinivälitteellä endosytoosilla. Vapauduttuaan endosomeista se ku...

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Main Author: Järvensivu, Jani
Other Authors: Matemaattis-luonnontieteellinen tiedekunta, Faculty of Sciences, Bio- ja ympäristötieteiden laitos, Department of Biological and Environmental Science, University of Jyväskylä, Jyväskylän yliopisto
Format: Master's thesis
Language:fin
Published: 2017
Subjects:
Online Access: https://jyx.jyu.fi/handle/123456789/54879
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author Järvensivu, Jani
author2 Matemaattis-luonnontieteellinen tiedekunta Faculty of Sciences Bio- ja ympäristötieteiden laitos Department of Biological and Environmental Science University of Jyväskylä Jyväskylän yliopisto
author_facet Järvensivu, Jani Matemaattis-luonnontieteellinen tiedekunta Faculty of Sciences Bio- ja ympäristötieteiden laitos Department of Biological and Environmental Science University of Jyväskylä Jyväskylän yliopisto Järvensivu, Jani Matemaattis-luonnontieteellinen tiedekunta Faculty of Sciences Bio- ja ympäristötieteiden laitos Department of Biological and Environmental Science University of Jyväskylä Jyväskylän yliopisto
author_sort Järvensivu, Jani
datasource_str_mv jyx
description Koiran parvovirus (engl. canine parvovirus, CPV) on pieni (halkaisijaltaan ~25 nm), vaipaton protoparvovirus. Sillä on ikosahedraalinen kapsidi joka ympäröi noin 5 kb pitkää, yksijuosteista DNA -genomia. CPV kulkeutuu solun sisään klatriinivälitteellä endosytoosilla. Vapauduttuaan endosomeista se kuljetetaan tumaan, jossa genomi replikoidaan ja pakataan uusiin kapsideihin. Lopulta valmiit virukset maturoituvat ja poistuvat tumasta infektoidakseen uusia soluja. Parvovirusten on havaittu tarvitsevan isäntäsolunsa solusyklin säätelyä edistääkseen genominsa replikaatiota tai uusien partikkeleiden kasaamista. Parvovirusinfektion seurauksena solusykli pysähtyy S/G2-vaiheeseen. On toistaiseksi epäselvää tarvitsevatko parvovirukset solusyklin säätelyä myös vapautuakseen isäntäsolunsa tumasta. Tämän tutkimuksen tarkoituksena oli selvittää CPV:n tumasta vapautumiseen johtavia muutoksia solusyklissä ja tumadynamiikassa. Parvoviruksen tumasta vapautumisen tiedettiin ajoittuvan S/G2-vaiheen loppuun, joten solusyklin etenemistä kohti M-vaihetta tarkasteltiin kyseiselle siirtymälle keskeisen säätelyproteiinin, sykliini B1:n avulla. Aikaisemmissa tutkimuksissa CPV:n on havaittu aiheuttavan katkoksia isäntäsolujensa tumakuljetukseen myöhäisessä vaiheessa infektiota. Tässä tutkimuksessa pyrittiin selvittämään liittyvätkö nämä ilmiöt viruksen tumasta poistumiseen ja G2/M -siirtymään. Tutkimus osoitti ensimmäistä kertaa, että CPV:n tumasta poistuminen ajoittuu G2/M –vaiheeseen. Tutkimuksessa havaittiin, että myöhäisen vaiheen CPV-infektio aiheuttaa erityisesti importiini β:n sekä sykliini B1:n, mutta myös sykliini B1:n fosforyloitujen muotojen akkumuloitumista tumaan. Virusten marginalisoituessa tumakotelon sisäpinnalle ennen tumasta poistumistaan, sytoplasmista sykliini B1:tä fosforyloitiin seriineistä 126 ja 147, jonka seurauksena molemmat fosforylaatiot akkumuloituivat samanaikaisesti tumaan. Nopea tuma-akkumulaatio osoittaa solusyklin edenneen G2/M-vaiheeseen. Lisäksi S126 ja S147 fosforyoitujen sykliinien määrän todettiin kasvavan infektion edetessä. Nämä tulokset kertoivat solun vähitellen siirtyvän G2/M-tarkastuspisteeseen infektion aikana. Tutkimuksessa havaittiin lisäksi, että viruksen poistuessa tumasta sytoplasmaan, S147 fosforyloitu sykliini B1 akkumuloitui tuman sijasta sytoplasmaan, mikä kertoo sykliini välitteisestä G2/M –säätelystä viruksen tumasta poistumisen aikana. Tutkimuksen perusteella on kiistatonta, että myös myöhäinen infektio vaikuttaa solusyklin etenemiseen ja G2/M-vaihe on keskeistä viruksen tumasta poistumiselle. Lisätutkimuksia kuitenkin tarvitaan näihin G2/M -vaihetta edistävien signalointireittien selvittämiseksi. Erityisesti infektion seurauksena aktivoituvan DNA-vauriovasteen merkitys havaitulle G2/M-siirtymälle on tärkeä tulevaisuuden tutkimuskohde. Canine parvovirus (CPV) is a small (~25 nm in diameter) nonenveloped protoparvovirus. An icosahedral capsid encloses a single stranded DNA genome of ~5 kb in length. CPV internalisation into a host cell occurs via endocytosis followed by transport into the nucleus. Viral genome is replicated and packaged into newly assembled capsids in the nucleoplasm. Following maturation progeny viruses egress from the nucleus and are ready to infect other cells. Parvoviruses are dependent on the host cell cycle regulation in order to promote their replication or the capsid assembly resulting in S/G2 cell cycle arrest. However, it has remained unknown whether they require further cell cycle regulation during their egress from the nucleus. The aim of this study was to analyse the role of cell cycle regulation and nuclear dynamics in parvoviral nuclear egress. The nuclear egress of parvoviruses is known to occur after S/G2. To study if the cell cycle proceeds towards G2/M during the egress, the intracellular distribution and phosphorylation levels of cyclin B1, a key regulator of the G2/M transition, was investigated. Recent studies with CPV have indicated that the late-infection causes temporal interruptions in the host cell nuclear import. In this study, the relation of these phenomena on the virus nuclear egress or G2/M-phase transition was evaluated. This study shows for the first time that the nuclear egress of CPV occurs in the G2/M –phase of the cell cycle. The results indicate that late CPV-infection causes nuclear accumulation of not only importin β and cyclin B1 but also of the serine 126 and 147 phosphorylated forms of cyclin B1. Virus marginalization to the nucleus periphery near to the inner nuclear membrane prior to nuclear escape was accompanied with cyclin B1 phosphorylation at S126 and S147 and nuclear accumulation. Moreover, the intranuclear levels of these phosphorylated cyclins were increased during the progression of the infection showing gradual cell cycle progression into G2/M. Virus escape from the nucleus into the cytoplasm was followed by synchronous cytoplasmic retention of S147P cyclin B1 indicating cyclin-mediated cell cycle regulation during the nuclear egress of parvovirus capsids. Together, these results indicate that the nuclear egress of CPV is accompanied with G2/M transition or checkpoint activation. This study shows undisputably that parvoviral infection causes cell cycle progression from late S/G2 to G2/M. However, further studies are needed to clarify the signalling pathways behind the G2/M phase transition. Especially the effects of parvovirus-induced DNA damage response on the detected G2/M arrest are a significant aspect in future experiments.
first_indexed 2023-03-22T09:59:00Z
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Sill\u00e4 on ikosahedraalinen kapsidi joka ymp\u00e4r\u00f6i noin 5 kb pitk\u00e4\u00e4, yksijuosteista DNA -genomia. CPV kulkeutuu solun sis\u00e4\u00e4n klatriiniv\u00e4litteell\u00e4 endosytoosilla. Vapauduttuaan endosomeista se kuljetetaan tumaan, jossa genomi replikoidaan ja pakataan uusiin kapsideihin. Lopulta valmiit virukset maturoituvat ja poistuvat tumasta infektoidakseen uusia soluja. Parvovirusten on havaittu tarvitsevan is\u00e4nt\u00e4solunsa solusyklin s\u00e4\u00e4tely\u00e4 edist\u00e4\u00e4kseen genominsa replikaatiota tai uusien partikkeleiden kasaamista. Parvovirusinfektion seurauksena solusykli pys\u00e4htyy S/G2-vaiheeseen. On toistaiseksi ep\u00e4selv\u00e4\u00e4 tarvitsevatko parvovirukset solusyklin s\u00e4\u00e4tely\u00e4 my\u00f6s vapautuakseen is\u00e4nt\u00e4solunsa tumasta. T\u00e4m\u00e4n tutkimuksen tarkoituksena oli selvitt\u00e4\u00e4 CPV:n tumasta vapautumiseen johtavia muutoksia solusykliss\u00e4 ja tumadynamiikassa. Parvoviruksen tumasta vapautumisen tiedettiin ajoittuvan S/G2-vaiheen loppuun, joten solusyklin etenemist\u00e4 kohti M-vaihetta tarkasteltiin kyseiselle siirtym\u00e4lle keskeisen s\u00e4\u00e4telyproteiinin, sykliini B1:n avulla. Aikaisemmissa tutkimuksissa CPV:n on havaittu aiheuttavan katkoksia is\u00e4nt\u00e4solujensa tumakuljetukseen my\u00f6h\u00e4isess\u00e4 vaiheessa infektiota. T\u00e4ss\u00e4 tutkimuksessa pyrittiin selvitt\u00e4m\u00e4\u00e4n liittyv\u00e4tk\u00f6 n\u00e4m\u00e4 ilmi\u00f6t viruksen tumasta poistumiseen ja G2/M -siirtym\u00e4\u00e4n. Tutkimus osoitti ensimm\u00e4ist\u00e4 kertaa, ett\u00e4 CPV:n tumasta poistuminen ajoittuu G2/M \u2013vaiheeseen. Tutkimuksessa havaittiin, ett\u00e4 my\u00f6h\u00e4isen vaiheen CPV-infektio aiheuttaa erityisesti importiini \u03b2:n sek\u00e4 sykliini B1:n, mutta my\u00f6s sykliini B1:n fosforyloitujen muotojen akkumuloitumista tumaan. Virusten marginalisoituessa tumakotelon sis\u00e4pinnalle ennen tumasta poistumistaan, sytoplasmista sykliini B1:t\u00e4 fosforyloitiin seriineist\u00e4 126 ja 147, jonka seurauksena molemmat fosforylaatiot akkumuloituivat samanaikaisesti tumaan. Nopea tuma-akkumulaatio osoittaa solusyklin edenneen G2/M-vaiheeseen. Lis\u00e4ksi S126 ja S147 fosforyoitujen sykliinien m\u00e4\u00e4r\u00e4n todettiin kasvavan infektion edetess\u00e4. N\u00e4m\u00e4 tulokset kertoivat solun v\u00e4hitellen siirtyv\u00e4n G2/M-tarkastuspisteeseen infektion aikana. Tutkimuksessa havaittiin lis\u00e4ksi, ett\u00e4 viruksen poistuessa tumasta sytoplasmaan, S147 fosforyloitu sykliini B1 akkumuloitui tuman sijasta sytoplasmaan, mik\u00e4 kertoo sykliini v\u00e4litteisest\u00e4 G2/M \u2013s\u00e4\u00e4telyst\u00e4 viruksen tumasta poistumisen aikana. Tutkimuksen perusteella on kiistatonta, ett\u00e4 my\u00f6s my\u00f6h\u00e4inen infektio vaikuttaa solusyklin etenemiseen ja G2/M-vaihe on keskeist\u00e4 viruksen tumasta poistumiselle. Lis\u00e4tutkimuksia kuitenkin tarvitaan n\u00e4ihin G2/M -vaihetta edist\u00e4vien signalointireittien selvitt\u00e4miseksi. Erityisesti infektion seurauksena aktivoituvan DNA-vauriovasteen merkitys havaitulle G2/M-siirtym\u00e4lle on t\u00e4rke\u00e4 tulevaisuuden tutkimuskohde.", "language": "fi", "element": "description", "qualifier": "abstract", "schema": "dc"}, {"key": "dc.description.abstract", "value": "Canine parvovirus (CPV) is a small (~25 nm in diameter) nonenveloped protoparvovirus. An icosahedral capsid encloses a single stranded DNA genome of ~5 kb in length. CPV internalisation into a host cell occurs via endocytosis followed by transport into the nucleus. Viral genome is replicated and packaged into newly\r\nassembled capsids in the nucleoplasm. Following maturation progeny viruses egress from the nucleus and are\r\nready to infect other cells. Parvoviruses are dependent on the host cell cycle regulation in order to promote\r\ntheir replication or the capsid assembly resulting in S/G2 cell cycle arrest. However, it has remained unknown\r\nwhether they require further cell cycle regulation during their egress from the nucleus.\r\nThe aim of this study was to analyse the role of cell cycle regulation and nuclear dynamics in parvoviral\r\nnuclear egress. The nuclear egress of parvoviruses is known to occur after S/G2. To study if the cell cycle\r\nproceeds towards G2/M during the egress, the intracellular distribution and phosphorylation levels of cyclin\r\nB1, a key regulator of the G2/M transition, was investigated. Recent studies with CPV have indicated that the\r\nlate-infection causes temporal interruptions in the host cell nuclear import. In this study, the relation of these\r\nphenomena on the virus nuclear egress or G2/M-phase transition was evaluated.\r\nThis study shows for the first time that the nuclear egress of CPV occurs in the G2/M \u2013phase of the cell cycle.\r\nThe results indicate that late CPV-infection causes nuclear accumulation of not only importin \u03b2 and cyclin\r\nB1 but also of the serine 126 and 147 phosphorylated forms of cyclin B1. Virus marginalization to the\r\nnucleus periphery near to the inner nuclear membrane prior to nuclear escape was accompanied with cyclin\r\nB1 phosphorylation at S126 and S147 and nuclear accumulation. Moreover, the intranuclear levels of these\r\nphosphorylated cyclins were increased during the progression of the infection showing gradual cell cycle\r\nprogression into G2/M. 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spellingShingle Järvensivu, Jani Solusyklin G2/M-vaiheen säätelyn vaikutus koiran parvoviruksen tumasta vapautumiseen solusykli sykliini B1 tumasta poistuminen canine parvovirus cell cycle regulation tuma Solu- ja molekyylibiologia Cell and molecular biology 4013 parvovirukset koira infektiot sykliinit solubiologia
title Solusyklin G2/M-vaiheen säätelyn vaikutus koiran parvoviruksen tumasta vapautumiseen
title_full Solusyklin G2/M-vaiheen säätelyn vaikutus koiran parvoviruksen tumasta vapautumiseen
title_fullStr Solusyklin G2/M-vaiheen säätelyn vaikutus koiran parvoviruksen tumasta vapautumiseen Solusyklin G2/M-vaiheen säätelyn vaikutus koiran parvoviruksen tumasta vapautumiseen
title_full_unstemmed Solusyklin G2/M-vaiheen säätelyn vaikutus koiran parvoviruksen tumasta vapautumiseen Solusyklin G2/M-vaiheen säätelyn vaikutus koiran parvoviruksen tumasta vapautumiseen
title_short Solusyklin G2/M-vaiheen säätelyn vaikutus koiran parvoviruksen tumasta vapautumiseen
title_sort solusyklin g2 m vaiheen säätelyn vaikutus koiran parvoviruksen tumasta vapautumiseen
title_txtP Solusyklin G2/M-vaiheen säätelyn vaikutus koiran parvoviruksen tumasta vapautumiseen
topic solusykli sykliini B1 tumasta poistuminen canine parvovirus cell cycle regulation tuma Solu- ja molekyylibiologia Cell and molecular biology 4013 parvovirukset koira infektiot sykliinit solubiologia
topic_facet 4013 Cell and molecular biology Solu- ja molekyylibiologia canine parvovirus cell cycle regulation infektiot koira parvovirukset solubiologia solusykli sykliini B1 sykliinit tuma tumasta poistuminen
url https://jyx.jyu.fi/handle/123456789/54879 http://www.urn.fi/URN:NBN:fi:jyu-201707063248
work_keys_str_mv AT järvensivujani solusykling2mvaiheensäätelynvaikutuskoiranparvoviruksentumastavapautumiseen