Werner Syndrome Protein (WRN) Regulates Cell Proliferation and the Human Papillomavirus 16 Life Cycle during Epithelial Differentiation

ABSTRACT Human papillomaviruses recruit a host of DNA damage response factors to their viral genome to facilitate homologous recombination replication in association with the viral replication factors E1 and E2. We previously demonstrated that SIRT1 deacetylation of WRN promotes recruitment of WRN t...

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Autores principales: Claire D. James, Dipon Das, Ethan L. Morgan, Raymonde Otoa, Andrew Macdonald, Iain M. Morgan
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Publicado: American Society for Microbiology 2020
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spelling oai:doaj.org-article:108ae300650b4a188d515bf13ee2edca2021-11-15T15:30:58ZWerner Syndrome Protein (WRN) Regulates Cell Proliferation and the Human Papillomavirus 16 Life Cycle during Epithelial Differentiation10.1128/mSphere.00858-202379-5042https://doaj.org/article/108ae300650b4a188d515bf13ee2edca2020-10-01T00:00:00Zhttps://journals.asm.org/doi/10.1128/mSphere.00858-20https://doaj.org/toc/2379-5042ABSTRACT Human papillomaviruses recruit a host of DNA damage response factors to their viral genome to facilitate homologous recombination replication in association with the viral replication factors E1 and E2. We previously demonstrated that SIRT1 deacetylation of WRN promotes recruitment of WRN to E1-E2 replicating DNA and that WRN regulates both the levels and fidelity of E1-E2 replication. The deacetylation of WRN by SIRT1 results in an active protein able to complex with replicating DNA, but a protein that is less stable. Here, we demonstrate an inverse correlation between SIRT1 and WRN in CIN cervical lesions compared to normal control tissue, supporting our model of SIRT1 deacetylation destabilizing WRN protein. We CRISPR/Cas9 edited N/Tert-1 and N/Tert-1+HPV16 cells to knock out WRN protein expression and subjected the cells to organotypic raft cultures. In N/Tert-1 cells without WRN expression, there was enhanced basal cell proliferation, DNA damage, and thickening of the differentiated epithelium. In N/Tert-1+HPV16 cells, there was enhanced basal cell proliferation, increased DNA damage throughout the epithelium, and increased viral DNA replication. Overall, the results demonstrate that the expression of WRN is required to control the proliferation of N/Tert-1 cells and controls the HPV16 life cycle in these cells. This complements our previous data demonstrating that WRN controls the levels and fidelity of HPV16 E1-E2 DNA replication. The results describe a new role for WRN, a tumor suppressor, in controlling keratinocyte differentiation and the HPV16 life cycle. IMPORTANCE HPV16 is the major human viral carcinogen, responsible for around 3 to 4% of all cancers worldwide. Our understanding of how the viral replication machinery interacts with host factors to control/activate the DNA damage response to promote the viral life cycle remains incomplete. Recently, we demonstrated a SIRT1-WRN axis that controls HPV16 replication, and here we demonstrate that this axis persists in clinical cervical lesions induced by HPV16. Here, we describe the effects of WRN depletion on cellular differentiation with or without HPV16; WRN depletion results in enhanced proliferation and DNA damage irrespective of HPV16 status. Also, WRN is a restriction factor for the viral life cycle since replication is disrupted in the absence of WRN. Future studies will focus on enhancing our understanding of how WRN regulates viral replication. Our goal is to ultimately identify cellular factors essential for HPV16 replication that can be targeted for therapeutic gain.Claire D. JamesDipon DasEthan L. MorganRaymonde OtoaAndrew MacdonaldIain M. MorganAmerican Society for MicrobiologyarticleWRNWerner helicasehuman papillomavirus 16replicationlife cycleDNA damageMicrobiologyQR1-502ENmSphere, Vol 5, Iss 5 (2020)
institution DOAJ
collection DOAJ
language EN
topic WRN
Werner helicase
human papillomavirus 16
replication
life cycle
DNA damage
Microbiology
QR1-502
spellingShingle WRN
Werner helicase
human papillomavirus 16
replication
life cycle
DNA damage
Microbiology
QR1-502
Claire D. James
Dipon Das
Ethan L. Morgan
Raymonde Otoa
Andrew Macdonald
Iain M. Morgan
Werner Syndrome Protein (WRN) Regulates Cell Proliferation and the Human Papillomavirus 16 Life Cycle during Epithelial Differentiation
description ABSTRACT Human papillomaviruses recruit a host of DNA damage response factors to their viral genome to facilitate homologous recombination replication in association with the viral replication factors E1 and E2. We previously demonstrated that SIRT1 deacetylation of WRN promotes recruitment of WRN to E1-E2 replicating DNA and that WRN regulates both the levels and fidelity of E1-E2 replication. The deacetylation of WRN by SIRT1 results in an active protein able to complex with replicating DNA, but a protein that is less stable. Here, we demonstrate an inverse correlation between SIRT1 and WRN in CIN cervical lesions compared to normal control tissue, supporting our model of SIRT1 deacetylation destabilizing WRN protein. We CRISPR/Cas9 edited N/Tert-1 and N/Tert-1+HPV16 cells to knock out WRN protein expression and subjected the cells to organotypic raft cultures. In N/Tert-1 cells without WRN expression, there was enhanced basal cell proliferation, DNA damage, and thickening of the differentiated epithelium. In N/Tert-1+HPV16 cells, there was enhanced basal cell proliferation, increased DNA damage throughout the epithelium, and increased viral DNA replication. Overall, the results demonstrate that the expression of WRN is required to control the proliferation of N/Tert-1 cells and controls the HPV16 life cycle in these cells. This complements our previous data demonstrating that WRN controls the levels and fidelity of HPV16 E1-E2 DNA replication. The results describe a new role for WRN, a tumor suppressor, in controlling keratinocyte differentiation and the HPV16 life cycle. IMPORTANCE HPV16 is the major human viral carcinogen, responsible for around 3 to 4% of all cancers worldwide. Our understanding of how the viral replication machinery interacts with host factors to control/activate the DNA damage response to promote the viral life cycle remains incomplete. Recently, we demonstrated a SIRT1-WRN axis that controls HPV16 replication, and here we demonstrate that this axis persists in clinical cervical lesions induced by HPV16. Here, we describe the effects of WRN depletion on cellular differentiation with or without HPV16; WRN depletion results in enhanced proliferation and DNA damage irrespective of HPV16 status. Also, WRN is a restriction factor for the viral life cycle since replication is disrupted in the absence of WRN. Future studies will focus on enhancing our understanding of how WRN regulates viral replication. Our goal is to ultimately identify cellular factors essential for HPV16 replication that can be targeted for therapeutic gain.
format article
author Claire D. James
Dipon Das
Ethan L. Morgan
Raymonde Otoa
Andrew Macdonald
Iain M. Morgan
author_facet Claire D. James
Dipon Das
Ethan L. Morgan
Raymonde Otoa
Andrew Macdonald
Iain M. Morgan
author_sort Claire D. James
title Werner Syndrome Protein (WRN) Regulates Cell Proliferation and the Human Papillomavirus 16 Life Cycle during Epithelial Differentiation
title_short Werner Syndrome Protein (WRN) Regulates Cell Proliferation and the Human Papillomavirus 16 Life Cycle during Epithelial Differentiation
title_full Werner Syndrome Protein (WRN) Regulates Cell Proliferation and the Human Papillomavirus 16 Life Cycle during Epithelial Differentiation
title_fullStr Werner Syndrome Protein (WRN) Regulates Cell Proliferation and the Human Papillomavirus 16 Life Cycle during Epithelial Differentiation
title_full_unstemmed Werner Syndrome Protein (WRN) Regulates Cell Proliferation and the Human Papillomavirus 16 Life Cycle during Epithelial Differentiation
title_sort werner syndrome protein (wrn) regulates cell proliferation and the human papillomavirus 16 life cycle during epithelial differentiation
publisher American Society for Microbiology
publishDate 2020
url https://doaj.org/article/108ae300650b4a188d515bf13ee2edca
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