Purification and characterization of the simian virus 40 transcription elongation complex

The transcriptional regulatory region of the simian virus 40 minichromosome that is being transcribed in the cell is nucleosome-free, while that of the non-transcribed minichromosome is nucleosome covered. Although additional studies have shown that the two structures are otherwise similar, the precision of these indirect studies has not been sufficient to determine if the transition between the two involves nucleosome displacement or nucleosome sliding. In order to address this question directly, we have developed a new function-based affinity isolation method that is capable of purifying the native transcription elongation complex of a single gene from mammalian cells. The simian virus 40 transcription elongation complex was purified by this method and the topological linking number of its DNA was compared directly to that of the bulk, non-transcribed minichromosome. The results show that the two types of minichromosome contain the same number of nucleosomes as well as nucleosomal structure. These findings indicate that interconversion between the non-transcribing and transcribing states is accomplished by a remodeling event involving nucleosome sliding rather than nucleosome displacement.