Integration of the SV40 promoter/enhancer sequences in an anchorage-independent clonal subline of SV40-infected human keratinocytes

Abstract

Long-term cultures of SV40-infected human keratinocytes contain integrated, but structurally altered copies of the viral sequences. The presence of these sequences is required for expression of properties associated with the transformed phenotype including immortalization. The integrated viral sequences in an anchorage-independent line of viral-transformed human keratinocytes have been found to be contained on two BamHI fragments of about 6.9 and 5.2 kb. In the larger fragment the viral sequences were present as two tandemly repeated subgenomic fragments containing the viral origin/promoter region with nucleotide alterations that affect enhancer function, the origin of replication and T antigen binding site 1. The viral early gene promoter in one integrant gave rise to an unspliced fusion transcript comprised of a short portion of the viral early gene leader sequence and the flanking human sequences. The smaller fragment consisted of full-length SV40 containing a nine-nucleotide insertion in the C-terminal portion of the SV40 T antigen, a region involved in the regulation of viral host range.