Mapping and DNA sequence analysis of the cytomegalovirus transforming domain III (mtrIII)

The transforming activity of XbaI fragment E (map units 0.685-0.770) of human cytomegalovirus Towne strain was originally localized in two terminal segments, 3 kb XbaI-BamHI EM (mtrII) and 7.5 kb BamHI-XbaI EJ (mtrIII). Three immediate-early genes, IE1 to IE3, are located in the same region (map unit 0.685-0.770), and IE1 is located in the EJ fragment. IE1 protein increases chromatin transcriptional activity and causes alteration in chromatin conformation. In this study, we have investigated the potential transforming activity of IE1, IE1 plus its promoter-regulatory region (IE P/R), and the remaining fragment of EJ after deletion of IE1 plus IE P/R. Our results clearly demonstrate that IE1 or IE1 plus IE P/R is not involved in transformation, and that the transforming activity of mtrIII is located in a 2.1 kb SalI-XbaI subfragment of EJ. DNA sequence analysis reveals four putative open reading frames (ORFs) L1, L2, L3 and R1 in the 2.1 kb fragment. The data from the deletion clones of the 2.1 kb fragment and also by disruption of individual ORFs by restriction enzyme digestion, suggest that a complete ORF L3 or R1 may be essential but not sufficient for the transforming activity. Because significant reduction in transforming activity is obtained by interrupting ORFs LI and L2, these ORFs may be required for the full transforming activity.