SAMHD1 Regulates Human Papillomavirus 16-Induced Cell Proliferation and Viral Replication during Differentiation of Keratinocytes

ABSTRACT Human papillomaviruses induce a host of anogenital cancers, as well as oropharyngeal cancer (HPV+OPC); human papillomavirus 16 (HPV16) is causative in around 90% of HPV+OPC cases. Using telomerase reverse transcriptase (TERT) immortalized foreskin keratinocytes (N/Tert-1), we have identifie...

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Autores principales:	Claire D. James, Apurva T. Prabhakar, Raymonde Otoa, Michael R. Evans, Xu Wang, Molly L. Bristol, Kun Zhang, Renfeng Li, Iain M. Morgan
Formato:	article
Lenguaje:	EN
Publicado:	American Society for Microbiology 2019
Materias:	SAMHD1 differentiation host-pathogen interactions life cycle papillomavirus viral replication Microbiology QR1-502
Acceso en línea:	https://doaj.org/article/e19dd57447874452a7dbf5de97248421
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spelling	oai:doaj.org-article:e19dd57447874452a7dbf5de972484212021-11-15T15:22:27ZSAMHD1 Regulates Human Papillomavirus 16-Induced Cell Proliferation and Viral Replication during Differentiation of Keratinocytes10.1128/mSphere.00448-192379-5042https://doaj.org/article/e19dd57447874452a7dbf5de972484212019-08-01T00:00:00Zhttps://journals.asm.org/doi/10.1128/mSphere.00448-19https://doaj.org/toc/2379-5042ABSTRACT Human papillomaviruses induce a host of anogenital cancers, as well as oropharyngeal cancer (HPV+OPC); human papillomavirus 16 (HPV16) is causative in around 90% of HPV+OPC cases. Using telomerase reverse transcriptase (TERT) immortalized foreskin keratinocytes (N/Tert-1), we have identified significant host gene reprogramming by HPV16 (N/Tert-1+HPV16) and demonstrated that N/Tert-1+HPV16 support late stages of the viral life cycle. Expression of the cellular dNTPase and homologous recombination factor sterile alpha motif and histidine-aspartic domain HD-containing protein 1 (SAMHD1) is transcriptionally regulated by HPV16 in N/Tert-1. CRISPR/Cas9 removal of SAMHD1 from N/Tert-1 and N/Tert-1+HPV16 demonstrates that SAMHD1 controls cell proliferation of N/Tert-1 only in the presence of HPV16; the deletion of SAMHD1 promotes hyperproliferation of N/Tert-1+HPV16 cells in organotypic raft cultures but has no effect on N/Tert-1. Viral replication is also elevated in the absence of SAMHD1. This new system has allowed us to identify a specific interaction between SAMHD1 and HPV16 that regulates host cell proliferation and viral replication; such studies are problematic in nonimmortalized primary keratinocytes due to their limited life span. To confirm the relevance of our results, we repeated the analysis with human tonsil keratinocytes (HTK) immortalized by HPV16 (HTK+HPV16) and observed the same hyperproliferative phenotype following CRISPR/Cas9 editing of SAMHD1. Identical results were obtained with three independent CRISPR/Cas9 guide RNAs. The isogenic pairing of N/Tert-1 with N/Tert-1+HPV16, combined with HTK+HPV16, presents a unique system to identify host genes whose products functionally interact with HPV16 to regulate host cellular growth in keratinocytes. IMPORTANCE HPVs are causative agents in human cancers and are responsible for around of 5% of all cancers. A better understanding of the viral life cycle in keratinocytes will facilitate the development of novel therapeutics to combat HPV-positive cancers. Here, we present a unique keratinocyte model to identify host proteins that specifically interact with HPV16. Using this system, we report that a cellular gene, SAMHD1, is regulated by HPV16 at the RNA and protein levels in keratinocytes. Elimination of SAMHD1 from these cells using CRISPR/Cas9 editing promotes enhanced cellular proliferation by HPV16 in keratinocytes and elevated viral replication but not in keratinocytes that do not have HPV16. Our study demonstrates a specific intricate interplay between HPV16 and SAMHD1 during the viral life cycle and establishes a unique model system to assist exploring host factors critical for HPV pathogenesis.Claire D. JamesApurva T. PrabhakarRaymonde OtoaMichael R. EvansXu WangMolly L. BristolKun ZhangRenfeng LiIain M. MorganAmerican Society for MicrobiologyarticleSAMHD1differentiationhost-pathogen interactionslife cyclepapillomavirusviral replicationMicrobiologyQR1-502ENmSphere, Vol 4, Iss 4 (2019)
institution	DOAJ
collection	DOAJ
language	EN
topic	SAMHD1 differentiation host-pathogen interactions life cycle papillomavirus viral replication Microbiology QR1-502
spellingShingle	SAMHD1 differentiation host-pathogen interactions life cycle papillomavirus viral replication Microbiology QR1-502 Claire D. James Apurva T. Prabhakar Raymonde Otoa Michael R. Evans Xu Wang Molly L. Bristol Kun Zhang Renfeng Li Iain M. Morgan SAMHD1 Regulates Human Papillomavirus 16-Induced Cell Proliferation and Viral Replication during Differentiation of Keratinocytes
description	ABSTRACT Human papillomaviruses induce a host of anogenital cancers, as well as oropharyngeal cancer (HPV+OPC); human papillomavirus 16 (HPV16) is causative in around 90% of HPV+OPC cases. Using telomerase reverse transcriptase (TERT) immortalized foreskin keratinocytes (N/Tert-1), we have identified significant host gene reprogramming by HPV16 (N/Tert-1+HPV16) and demonstrated that N/Tert-1+HPV16 support late stages of the viral life cycle. Expression of the cellular dNTPase and homologous recombination factor sterile alpha motif and histidine-aspartic domain HD-containing protein 1 (SAMHD1) is transcriptionally regulated by HPV16 in N/Tert-1. CRISPR/Cas9 removal of SAMHD1 from N/Tert-1 and N/Tert-1+HPV16 demonstrates that SAMHD1 controls cell proliferation of N/Tert-1 only in the presence of HPV16; the deletion of SAMHD1 promotes hyperproliferation of N/Tert-1+HPV16 cells in organotypic raft cultures but has no effect on N/Tert-1. Viral replication is also elevated in the absence of SAMHD1. This new system has allowed us to identify a specific interaction between SAMHD1 and HPV16 that regulates host cell proliferation and viral replication; such studies are problematic in nonimmortalized primary keratinocytes due to their limited life span. To confirm the relevance of our results, we repeated the analysis with human tonsil keratinocytes (HTK) immortalized by HPV16 (HTK+HPV16) and observed the same hyperproliferative phenotype following CRISPR/Cas9 editing of SAMHD1. Identical results were obtained with three independent CRISPR/Cas9 guide RNAs. The isogenic pairing of N/Tert-1 with N/Tert-1+HPV16, combined with HTK+HPV16, presents a unique system to identify host genes whose products functionally interact with HPV16 to regulate host cellular growth in keratinocytes. IMPORTANCE HPVs are causative agents in human cancers and are responsible for around of 5% of all cancers. A better understanding of the viral life cycle in keratinocytes will facilitate the development of novel therapeutics to combat HPV-positive cancers. Here, we present a unique keratinocyte model to identify host proteins that specifically interact with HPV16. Using this system, we report that a cellular gene, SAMHD1, is regulated by HPV16 at the RNA and protein levels in keratinocytes. Elimination of SAMHD1 from these cells using CRISPR/Cas9 editing promotes enhanced cellular proliferation by HPV16 in keratinocytes and elevated viral replication but not in keratinocytes that do not have HPV16. Our study demonstrates a specific intricate interplay between HPV16 and SAMHD1 during the viral life cycle and establishes a unique model system to assist exploring host factors critical for HPV pathogenesis.
format	article
author	Claire D. James Apurva T. Prabhakar Raymonde Otoa Michael R. Evans Xu Wang Molly L. Bristol Kun Zhang Renfeng Li Iain M. Morgan
author_facet	Claire D. James Apurva T. Prabhakar Raymonde Otoa Michael R. Evans Xu Wang Molly L. Bristol Kun Zhang Renfeng Li Iain M. Morgan
author_sort	Claire D. James
title	SAMHD1 Regulates Human Papillomavirus 16-Induced Cell Proliferation and Viral Replication during Differentiation of Keratinocytes
title_short	SAMHD1 Regulates Human Papillomavirus 16-Induced Cell Proliferation and Viral Replication during Differentiation of Keratinocytes
title_full	SAMHD1 Regulates Human Papillomavirus 16-Induced Cell Proliferation and Viral Replication during Differentiation of Keratinocytes
title_fullStr	SAMHD1 Regulates Human Papillomavirus 16-Induced Cell Proliferation and Viral Replication during Differentiation of Keratinocytes
title_full_unstemmed	SAMHD1 Regulates Human Papillomavirus 16-Induced Cell Proliferation and Viral Replication during Differentiation of Keratinocytes
title_sort	samhd1 regulates human papillomavirus 16-induced cell proliferation and viral replication during differentiation of keratinocytes
publisher	American Society for Microbiology
publishDate	2019
url	https://doaj.org/article/e19dd57447874452a7dbf5de97248421
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SAMHD1 Regulates Human Papillomavirus 16-Induced Cell Proliferation and Viral Replication during Differentiation of Keratinocytes

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