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An unbiased in vivo screen reveals multiple transcription factors that control HPV E6-regulated hTERT in keratinocytes.

ABSTRACT: Activation of telomerase by human papillomavirus 16 (HPV16) E6 is a critical step for cell immortalization and transformation in human foreskin keratinocytes (HFKs). Multiple transcription factors have been identified as being involved in E6-induced hTERT expression. Here, we adapted an unbiased in vivo screen using a LacO-LacI system in human cells to discover hTERT promoter-interacting regulators. This approach allowed us to identify a novel hTERT repressor, Maz, which bound the hTERT promoter. E6 expression reduced Maz binding and correspondingly increased Sp1 binding at the hTERT promoter. Knockdown of Maz further increased histone acetylation, as well as hTERT expression in the presence of E6. Overall, these data indicate the utility of a novel screen for promoter-interacting and transcription-regulating proteins. These data also highlight multiple factors that normally regulate hTERT repression in HFKs, and therefore are targeted by E6 for hTERT expression.

SUBMITTER: Xu M

PROVIDER: S-EPMC3787310 | biostudies-literature | 2013 Nov

REPOSITORIES: biostudies-literature

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An unbiased in vivo screen reveals multiple transcription factors that control HPV E6-regulated hTERT in keratinocytes.

Xu Mei M Katzenellenbogen Rachel A RA Grandori Carla C Galloway Denise A DA

Virology 20130808 1-2

Activation of telomerase by human papillomavirus 16 (HPV16) E6 is a critical step for cell immortalization and transformation in human foreskin keratinocytes (HFKs). Multiple transcription factors have been identified as being involved in E6-induced hTERT expression. Here, we adapted an unbiased in vivo screen using a LacO-LacI system in human cells to discover hTERT promoter-interacting regulators. This approach allowed us to identify a novel hTERT repressor, Maz, which bound the hTERT promoter ...[more]

PMID: 24074563

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Project description:PurposeHigh-risk human papillomaviruses (HR HPV) cause cervical cancer, and in these cancers, HPV type 16 is the most common HR type. The HR viral oncogenes E6 and E7 partner with cellular proteins to drive cancer and modulate immune pathways; previously, we demonstrated in keratinocytes that HPV 16 E6 and high expression of the endogenous host protein partner NFX1-123 led to the increased expression of multiple genes, including Notch1, secretory leukocyte peptidase inhibitor (SLPI), and retinoic acid early transcript 1G (RAET1G). The present study was conducted to determine if NFX1-123 was highly expressed in cervical cancer and if genes increased by NFX1-123 and 16E6 in keratinocytes were also increased in cervical cancers.Materials and methodsThe Cancer Genome Atlas (TCGA) database and The Human Protein Atlas database were used to compare relative mRNA and protein gene expression, respectively, in the normal cervix and cervical cancers. Formalin-fixed paraffin-embedded (FFPE) normal cervix and HPV 16 positive cervical cancer samples were analyzed for relative protein expression by immunohistochemical staining. Protein expression of a subset of regulated genes was quantified by Western blot of HPV positive and negative cell lines.ResultsImmunohistochemical staining of HPV 16 positive cervical dysplasias and cancers revealed high NFX1-123, Ki67, and Notch1 expression. NFX1 and NFX1L1 mRNA levels were increased in cervical cancers compared to normal cervix in the TCGA database. Fourteen genes previously identified as upregulated in keratinocytes with 16E6 and overexpressed NFX1-123 also had high mRNA expression and selected genes had high protein expression in cervical cancers and cell lines.ConclusionIn cervical cancer, NFX1-123 is highly expressed, and 16E6 and NFX1-123 together alter the expression of a wide set of genes. The involvement of these genes in cell proliferation, differentiation, invasion, and metastasis provides further insight into potential ways that HR HPVs promote cancer initiation and maintenance.

Dataset Information

An unbiased in vivo screen reveals multiple transcription factors that control HPV E6-regulated hTERT in keratinocytes.

Publications

An unbiased in vivo screen reveals multiple transcription factors that control HPV E6-regulated hTERT in keratinocytes.

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