The Differentiation-Specific Factor CDP/Cut Represses Transcription and Replication of Human Papillomaviruses through a Conserved Silencing Element

Abstract

ABSTRACT The life cycles of human papillomaviruses (HPVs) are intimately linked to the differentiation program of infected stratified epithelia, with both viral gene expression and replication being maintained at low levels in undifferentiated basal cells and increased upon host cell differentiation. We recently identified, in HPV-16, a negative regulatory element between the epithelial-cell-specific enhancer and the E6 promoter that is capable of silencing E6 promoter activity, and we termed this element a papillomavirus silencing motif (PSM) and the unknown cellular factor that bound to it PSM binding protein (PSM-BP). Here we show that the homologous genomic segments of six other distantly related genital HPV types contain a PSM that binds PSM-BP and is capable of repressing transcription. Conservation of the PSM suggests that it is indispensable for the HPV life cycle. Purification, electrophoretic mobility shift assay experiments, and the use of specific antibodies proved that the cellular factor PSM-BP is identical to a previously described transcriptional repressor, the CCAAT displacement protein (CDP), also referred to as the human Cut protein (Cut). CDP/Cut repression of HPV-16 may stem from the modification of specifically positioned nucleosomes, as suggested by transcriptional stimulation under the influence of the histone deacetylase inhibitor trichostatin A. CDP/Cut is an important developmental regulator in several different tissues. It was recently shown that CDP/Cut is expressed in basal epithelial cells but not in differentiated primary keratinocytes. This suggests the possibility that repression by PSM couples HPV transcription to the stratification of epithelia. In each of the studied HPV types, the two CDP/Cut binding sites of PSM overlap with the known or presumed binding sites of the replication initiator protein E1. Transfection of CDP/Cut expression vectors into cells that support HPV-16 or HPV-31 replication leads to the elimination of viral episomes. Similarly, two PSM-like motifs overlapping the E1 binding site of bovine papillomavirus type 1 bind CDP/Cut, and CDP/Cut overexpression reduces the copy number of episomally replicating BPV-1 genomes in mouse fibroblasts. CDP/Cut appears to be a master regulator of HPV transcription and replication during epithelial differentiation, and PSMs are important cis -responsive targets of this repressor.