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Suppression of STAT-1 expression by human papillomaviruses is necessary for differentiation-dependent genome amplification and plasmid maintenance.

ABSTRACT: High-risk human papillomaviruses (HPVs) infect stratified epithelia to establish persistent infections that maintain low-copy-number episomes in infected basal cells. Amplification of viral genomes occurs upon keratinocyte differentiation, followed by virion synthesis. During persistent HPV infections, viral proteins act to evade surveillance by both innate and adaptive immune responses. One of the primary pathways regulating the innate immune response is the JAK/STAT pathway. Our studies indicate that the expression of STAT-1, but not other members of interferon (IFN)-stimulated gene factor 3 (ISGF-3) complex such as STAT-2 and IFN regulatory factor 9 (IRF9), is selectively suppressed by HPV proteins at the level of transcription. Both E6 and E7 oncoproteins independently suppress the expression of STAT-1, and mutational analyses indicate that the E6 targeting E6-associated protein (E6AP) is responsible for suppression. The levels of STAT-1 proteins increase upon differentiation of both normal and HPV-positive cells but are still significantly reduced in the latter cells. Transient restoration of STAT-1 levels in HPV-positive cells using recombinant retroviruses significantly impaired viral amplification upon differentiation while long-term increases abrogated maintenance of episomes. Similarly, increased levels of STAT-1 induced by gamma interferon treatment inhibited HPV genome amplification upon differentiation. Overall, our findings demonstrate that suppression of STAT-1 expression by HPV proteins is necessary for genome amplification and maintenance of episomes, suggesting an important role for this activity in viral pathogenesis.

SUBMITTER: Hong S

PROVIDER: S-EPMC3165741 | biostudies-literature | 2011 Sep

REPOSITORIES: biostudies-literature

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Suppression of STAT-1 expression by human papillomaviruses is necessary for differentiation-dependent genome amplification and plasmid maintenance.

Hong Shiyuan S Mehta Kavi P KP Laimins Laimonis A LA

Journal of virology 20110706 18

High-risk human papillomaviruses (HPVs) infect stratified epithelia to establish persistent infections that maintain low-copy-number episomes in infected basal cells. Amplification of viral genomes occurs upon keratinocyte differentiation, followed by virion synthesis. During persistent HPV infections, viral proteins act to evade surveillance by both innate and adaptive immune responses. One of the primary pathways regulating the innate immune response is the JAK/STAT pathway. Our studies indica ...[more]

PMID: 21734056

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Project description:Human papillomaviruses (HPVs) are epithelial tropic viruses that link their productive life cycles to the differentiation of infected host keratinocytes. A subset of the over 200 HPV types, referred to as high-risk, are the causative agents of most anogenital malignancies. HPVs infect cells in the basal layer, but restrict viral genome amplification, late gene expression, and capsid assembly to highly differentiated cells that are active in the cell cycle. In this study, we demonstrate that HPV proteins regulate the expression and activities of a critical cellular transcription factor, KLF4, through post-transcriptional and post-translational mechanisms. Our studies show that KLF4 regulates differentiation as well as cell cycle progression, and binds to sequences in the upstream regulatory region (URR) to regulate viral transcription in cooperation with Blimp1. KLF4 levels are increased in HPV-positive cells through a post-transcriptional mechanism involving E7-mediated suppression of cellular miR-145, as well as at the post-translational level by E6-directed inhibition of its sumoylation and phosphorylation. The alterations in KLF4 levels and functions results in activation and suppression of a subset of KLF4 target genes, including TCHHL1, VIM, ACTN1, and POT1, that is distinct from that seen in normal keratinocytes. Knockdown of KLF4 with shRNAs in cells that maintain HPV episomes blocked genome amplification and abolished late gene expression upon differentiation. While KLF4 is indispensable for the proliferation and differentiation of normal keratinocytes, it is necessary only for differentiation-associated functions of HPV-positive keratinocytes. Increases in KLF4 levels alone do not appear to be sufficient to explain the effects on proliferation and differentiation of HPV-positive cells indicating that additional modifications are important. KLF4 has also been shown to be a critical regulator of lytic Epstein Barr virus (EBV) replication underscoring the importance of this cellular transcription factor in the life cycles of multiple human cancer viruses.

Project description:Human papillomaviruses infect stratified epithelia and link their productive life cycle to the differentiation state of the host cell. Productive viral replication or amplification is restricted to highly differentiated suprabasal cells and is dependent on the activation of the ATM DNA damage pathway. The ATM pathway has three arms that can act independently of one another. One arm is centered on p53, another on CHK2 and a third on SMC1/NBS1 proteins. A role for CHK2 in HPV genome amplification has been demonstrated but it was unclear what other factors provided important activities. The cohesin protein, SMC1, is necessary for sister chromatid association prior to mitosis. In addition the phosphorylated form of SMC1 plays a critical role together with NBS1 in the ATM DNA damage response. In normal cells, SMC1 becomes phosphorylated in response to radiation, however, in HPV positive cells our studies demonstrate that it is constitutively activated. Furthermore, pSMC1 is found localized in distinct nuclear foci in complexes with γ-H2AX, and CHK2 and bound to HPV DNA. Importantly, knockdown of SMC1 blocks differentiation-dependent genome amplification. pSMC1 forms complexes with the insulator transcription factor CTCF and our studies show that these factors bind to conserved sequence motifs in the L2 late region of HPV 31. Similar motifs are found in most HPV types. Knockdown of CTCF with shRNAs blocks genome amplification and mutation of the CTCF binding motifs in the L2 open reading frame inhibits stable maintenance of viral episomes in undifferentiated cells as well as amplification of genomes upon differentiation. These findings suggest a model in which SMC1 factors are constitutively activated in HPV positive cells and recruited to viral genomes through complex formation with CTCF to facilitate genome amplification. Our findings identify both SMC1 and CTCF as critical regulators of the differentiation-dependent life cycle of high-risk human papillomaviruses.

Dataset Information

Suppression of STAT-1 expression by human papillomaviruses is necessary for differentiation-dependent genome amplification and plasmid maintenance.

Publications

Suppression of STAT-1 expression by human papillomaviruses is necessary for differentiation-dependent genome amplification and plasmid maintenance.

Similar Datasets

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