Factors Influencing Physiological Variations in the Activity of the Rous Sarcoma Virus Long Terminal Repeat

Abstract

Previous reports have shown that Rous sarcoma virus (RSV) transcript levels in mammalian cells can be elevated by serum treatment and cellular transformation. To understand this, we have examined how the RSV long terminal repeat (LTR) enhancer is affected by cellular growth state in clonally related normal and RSV-infected Rat-1 cell lines. Functional assays with enhancer mutants have shown that two LTR CArG motifs and a CCAAT box have individual and combinatorial effects on basal LTR activity, but only the CArG elements contribute to serum responses in phenotypically normal cells. Augmented enhancer activity in transformed cells is mediated in part by these CArG motifs, which under these conditions are not further stimulable by serum. Protein binding to the CArG and CCAAT elements corresponds with functional variations, binding proteins being scarce in serum-deprived normal cells and enhanced by serum stimulation, cellular transformation and, in part, cellular density. These findings provide an explanation for physiologically dependent fluctuations in RSV expression, which may include the initiation of the proviral transcriptional repression that is frequently observed in mammalian hosts. However, we also show that transcript levels of some integrated proviruses are independent of variations in the cell's ability to support LTR activity, showing that the site of insertion can be of overriding importance in determining proviral expression.