Unravelling Betaretroviral Replication: The Pivotal Role of Host RNA Helicase DHX15

Kód výsledku v IS VaVaI

<a href="https://www.isvavai.cz/riv?ss=detail&h=RIV%2F60461373%3A22810%2F23%3A43932625" target="_blank" >RIV/60461373:22810/23:43932625 - isvavai.cz</a>

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RIV/60461373:22330/23:43932625

Výsledek na webu

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DOI - Digital Object Identifier

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Jazyk výsledku

angličtina

Název v původním jazyce

Unravelling Betaretroviral Replication: The Pivotal Role of Host RNA Helicase DHX15

Popis výsledku v původním jazyce

Viruses, characterized by their minimal genome size, heavily rely on host-cell protein machinery for effective replication. A prominent class of proteins in this context are helicases. The essential role of helicase activity in the replication cycle has been well-documented in various viruses, where helicases participate in reshaping viral ribonucleoproteins, predominantly during genome replication, splicing, nuclear export of unspliced viral RNA, transcription, translation, and viral packaging. Despite the pivotal role helicases play in viral propagation, many viruses, including well-known retroviruses like HIV, do not encode their own helicases, instead relying on those of the host cell.The Mason-Pfizer Monkey Virus (M-PMV) is a simple betaretrovirus, encoding its structural and enzymatic proteins in three genes - gag, pro, and pol - which are translated into three polyprotein precursors: Gag, Gag-Pro, and Gag-Pro-Pol. At the border of the Pro and Pol polyprotein precursors, a short glycine-rich peptide (G-patch domain, GPD) connects the protease (PR) and reverse transcriptase (RT). In eukaryotic cells, proteins containing GPD are often involved in the recruitment and subsequent activation of DEAH/RHA RNA helicases, such as DHX15.Through the MS analysis of the host-cell proteins in released M-PMV virions, we identified that RNA helicase DHX15 could be effectively packaged into M-PMV particles but not into ΔGPD M-PMV particles. To investigate the role of DHX15 in the M-PMV replication cycle, as well as the involvement of the M-PMV GPD in DHX15 recruitment, we performed a series of ex vivo and in vitro experiments, including mutagenesis, mRNA silencing, photoactivatable ribonucleoside-enhanced crosslinking, and immunoprecipitation assay (PAR-CLIP), thermophoresis, and isothermal titration calorimetry (ITC). Our results confirm the crucial involvement of DHX15 in the M-PMV replication cycle, but surprisingly, not only during genome replication but also in genomic RNA packaging.Acknowledgement: the research was funded by the The project National Institute of Virology and Bacteriology (Programme EXCELES, ID Project No. LX22NPO5103) - Funded by the European Union - Next Generation EU.

Název v anglickém jazyce

Unravelling Betaretroviral Replication: The Pivotal Role of Host RNA Helicase DHX15

Popis výsledku anglicky

Viruses, characterized by their minimal genome size, heavily rely on host-cell protein machinery for effective replication. A prominent class of proteins in this context are helicases. The essential role of helicase activity in the replication cycle has been well-documented in various viruses, where helicases participate in reshaping viral ribonucleoproteins, predominantly during genome replication, splicing, nuclear export of unspliced viral RNA, transcription, translation, and viral packaging. Despite the pivotal role helicases play in viral propagation, many viruses, including well-known retroviruses like HIV, do not encode their own helicases, instead relying on those of the host cell.The Mason-Pfizer Monkey Virus (M-PMV) is a simple betaretrovirus, encoding its structural and enzymatic proteins in three genes - gag, pro, and pol - which are translated into three polyprotein precursors: Gag, Gag-Pro, and Gag-Pro-Pol. At the border of the Pro and Pol polyprotein precursors, a short glycine-rich peptide (G-patch domain, GPD) connects the protease (PR) and reverse transcriptase (RT). In eukaryotic cells, proteins containing GPD are often involved in the recruitment and subsequent activation of DEAH/RHA RNA helicases, such as DHX15.Through the MS analysis of the host-cell proteins in released M-PMV virions, we identified that RNA helicase DHX15 could be effectively packaged into M-PMV particles but not into ΔGPD M-PMV particles. To investigate the role of DHX15 in the M-PMV replication cycle, as well as the involvement of the M-PMV GPD in DHX15 recruitment, we performed a series of ex vivo and in vitro experiments, including mutagenesis, mRNA silencing, photoactivatable ribonucleoside-enhanced crosslinking, and immunoprecipitation assay (PAR-CLIP), thermophoresis, and isothermal titration calorimetry (ITC). Our results confirm the crucial involvement of DHX15 in the M-PMV replication cycle, but surprisingly, not only during genome replication but also in genomic RNA packaging.Acknowledgement: the research was funded by the The project National Institute of Virology and Bacteriology (Programme EXCELES, ID Project No. LX22NPO5103) - Funded by the European Union - Next Generation EU.

Druh

O - Ostatní výsledky

CEP obor

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OECD FORD obor

10607 - Virology

Projekt

<a href="/cs/project/LX22NPO5103" target="_blank" >LX22NPO5103: Národní institut virologie a bakteriologie</a><br>

Návaznosti

P - Projekt vyzkumu a vyvoje financovany z verejnych zdroju (s odkazem do CEP)

Rok uplatnění

2023

Kód důvěrnosti údajů

S - Úplné a pravdivé údaje o projektu nepodléhají ochraně podle zvláštních právních předpisů