Localization of Double-Strand Break Repair Proteins to Viral Replication Compartments following Lytic Reactivation of Kaposi's Sarcoma-Associated Herpesvirus

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@article{4376a00df54345a6aafb5ffc12d3986e,
title = "Localization of Double-Strand Break Repair Proteins to Viral Replication Compartments following Lytic Reactivation of Kaposi's Sarcoma-Associated Herpesvirus",
abstract = "Double-strand breaks (DSBs) in DNA are recognised by the Ku70/80 heterodimer and the MRE11-RAD50-NBS1 (MRN) complex and result in activation of the DNA-PK and ATM kinases that play key roles in regulating the cellular DNA damage response (DDR). DNA tumour viruses such as Kaposi{\textquoteright}s sarcoma-associated herpesvirus (KSHV) are known to interact extensively with the DDR during the course of their replicative cycles. Here we show that during lytic amplification of KSHV DNA, the Ku70/80 heterodimer and the MRN complex consistently co-localise with viral genomes in replication compartments (RCs) whereas other DSB repair proteins form foci outside of RCs. Depletion of MRE11 and abrogation of its exonuclease activity negatively impacts on viral replication, while in contrast, knockdown of Ku80 and inhibition of the DNA-PK enzyme, which are involved in non-homologous end joining repair (NHEJ), enhances amplification of viral DNA. Although the recruitment of DSB sensing proteins to KSHV RCs is a consistent occurrence across multiple cell types, activation of the ATM-CHK2 pathway during viral replication is a cell line-specific event indicating that recognition of viral DNA by the DDR does not necessarily result in activation of downstream signalling pathways. We have also observed that newly replicated viral DNA is not associated with cellular histones. Since the presence and modification of these DNA packaging proteins provides a scaffold for docking of multiple DNA repair factors, the absence of histone deposition may allow the virus to evade localisation of DSB repair proteins that would otherwise have a detrimental effect on viral replication.",
keywords = "herpesvirus Kaposi's sarcoma-associated herpesvirus , ATM , DDR , DNA repair , DNA-PK , lytic replication MRN complex NHEJ replication, compartments",
author = "Rob Hollingworth and Richard Horniblow and Calum Forrest and Grant Stewart and Roger Grand",
year = "2017",
month = oct,
day = "27",
doi = "10.1128/JVI.00930-17",
language = "English",
volume = "91",
journal = "Journal of virology",
issn = "0022-538X",
publisher = "American Society for Microbiology",

}

RIS

TY - JOUR

T1 - Localization of Double-Strand Break Repair Proteins to Viral Replication Compartments following Lytic Reactivation of Kaposi's Sarcoma-Associated Herpesvirus

AU - Hollingworth, Rob

AU - Horniblow, Richard

AU - Forrest, Calum

AU - Stewart, Grant

AU - Grand, Roger

PY - 2017/10/27

Y1 - 2017/10/27

N2 - Double-strand breaks (DSBs) in DNA are recognised by the Ku70/80 heterodimer and the MRE11-RAD50-NBS1 (MRN) complex and result in activation of the DNA-PK and ATM kinases that play key roles in regulating the cellular DNA damage response (DDR). DNA tumour viruses such as Kaposi’s sarcoma-associated herpesvirus (KSHV) are known to interact extensively with the DDR during the course of their replicative cycles. Here we show that during lytic amplification of KSHV DNA, the Ku70/80 heterodimer and the MRN complex consistently co-localise with viral genomes in replication compartments (RCs) whereas other DSB repair proteins form foci outside of RCs. Depletion of MRE11 and abrogation of its exonuclease activity negatively impacts on viral replication, while in contrast, knockdown of Ku80 and inhibition of the DNA-PK enzyme, which are involved in non-homologous end joining repair (NHEJ), enhances amplification of viral DNA. Although the recruitment of DSB sensing proteins to KSHV RCs is a consistent occurrence across multiple cell types, activation of the ATM-CHK2 pathway during viral replication is a cell line-specific event indicating that recognition of viral DNA by the DDR does not necessarily result in activation of downstream signalling pathways. We have also observed that newly replicated viral DNA is not associated with cellular histones. Since the presence and modification of these DNA packaging proteins provides a scaffold for docking of multiple DNA repair factors, the absence of histone deposition may allow the virus to evade localisation of DSB repair proteins that would otherwise have a detrimental effect on viral replication.

AB - Double-strand breaks (DSBs) in DNA are recognised by the Ku70/80 heterodimer and the MRE11-RAD50-NBS1 (MRN) complex and result in activation of the DNA-PK and ATM kinases that play key roles in regulating the cellular DNA damage response (DDR). DNA tumour viruses such as Kaposi’s sarcoma-associated herpesvirus (KSHV) are known to interact extensively with the DDR during the course of their replicative cycles. Here we show that during lytic amplification of KSHV DNA, the Ku70/80 heterodimer and the MRN complex consistently co-localise with viral genomes in replication compartments (RCs) whereas other DSB repair proteins form foci outside of RCs. Depletion of MRE11 and abrogation of its exonuclease activity negatively impacts on viral replication, while in contrast, knockdown of Ku80 and inhibition of the DNA-PK enzyme, which are involved in non-homologous end joining repair (NHEJ), enhances amplification of viral DNA. Although the recruitment of DSB sensing proteins to KSHV RCs is a consistent occurrence across multiple cell types, activation of the ATM-CHK2 pathway during viral replication is a cell line-specific event indicating that recognition of viral DNA by the DDR does not necessarily result in activation of downstream signalling pathways. We have also observed that newly replicated viral DNA is not associated with cellular histones. Since the presence and modification of these DNA packaging proteins provides a scaffold for docking of multiple DNA repair factors, the absence of histone deposition may allow the virus to evade localisation of DSB repair proteins that would otherwise have a detrimental effect on viral replication.

KW - herpesvirus Kaposi's sarcoma-associated herpesvirus

KW - ATM

KW - DDR

KW - DNA repair

KW - DNA-PK

KW - lytic replication MRN complex NHEJ replication

KW - compartments

U2 - 10.1128/JVI.00930-17

DO - 10.1128/JVI.00930-17

M3 - Article

C2 - 28855246

VL - 91

JO - Journal of virology

JF - Journal of virology

SN - 0022-538X

M1 - e00930-17

ER -