Human cervical and NIH 3T3 cells were transfected with the XbaI-E-transforming fragment of human cytomegalovirus strain Towne. Southern blot hybridization showed that 3 of 4 transformed NIH 3T3 cell lines retained only the mtrII subfragment of Towne Xbal-E, but not the mtrlll subfragment. Even though mtrII was retained, no viral transcripts were detected. Analysis of genomic DNAs isolated from three independently derived lines of Towne XbaI-E-transfected human exocervical epithelial cells previously immortalized by human papillomavirus type 16 (CX16-2/Towne-E) revealed the retention of both mtrll and mtrlll subfragments of Towne Xba 30 subpassages. Southern blot hybridizations indicated the integration and rearrangement of mtrll as well as mtrlll. Poly (A)+RNA analysis of a CX16-2/Towne-E line revealed a 1.9-kb transcript which hybridized to mtr III. In contrast, no viral transcript from the mtrll region was detected in these cells. The pattern of HPV-16 DNA sequences and the profile of RNA transcripts were similar in the parental human exocervical cells (CX16-2) and in the CX16-2/Towne-E cells. Thus far, the CX16-2/Towne-E lines are nontumorigenic in nude mice. This study highlights not only differences in the ability of Towne Xbal-E to transform rodent cells and not human cells but also differences in the retention and expression of mtrll and mtrlll in these cells.