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Forschungsschwerpunkt Angewandte Tumorvirologie, Deutsches Krebsforschungszentrum, Im Neuenheimer Feld 242, D-6900 Heidelberg, Germany
Gene expression of the cancer-associated human papillomavirus (HPV) type 18 is modulated by cis-regulatory elements located within the viral upstream regulatory region (URR). All cellular factors identified so far involved in viral gene control bind to the 3' portion of the HPV-18 URR. In contrast, very little is known about regulatory elements within the 5' portion of the URR. We therefore analysed this region of unknown function to delineate potential cis-regulatory elements contained therein. By utilizing transient expression assays, an 84 bp fragment could be identified in the 5' portion of the URR that exhibits orientation-independent cis-stimulatory activity in HeLa cervical carcinoma cells and primary human fibroblasts. Gel retardation assays and competition experiments indicated specific binding of cellular proteins to the 84 bp fragment. By functional dissection, a regulatory element with intrinsic cis-stimulatory activity could be mapped within the 84 bp fragment. Binding studies indicate that this cis-stimulatory element contains a sequence-aberrant Sp1 recognition site. Transient luciferase assays performed with mutated templates demonstrate that this Sp1 binding site behaves as a functional Sp1 element in vivo and is a main determinant of the cis-stimulatory activity exerted by the 84 bp fragment. These data show that the 5' portion of the HPV-18 URR contains cis-activating elements and indicate an important functional role for the cellular transcription factor Sp1 in their regulation.
Received 5 August 1992;
accepted 5 October 1992.
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