Viral oncogenes accelerate conversion to immortality of cultured conditionally immortal human mammary epithelial cells.

Abstract

Our recent studies on the process of immortalization of cultured human mammary epithelial cells (HMEC) have uncovered a previously undescribed, apparently epigenetic step, termed conversion. When first isolated, clonally derived HMEC lines of indefinite lifespan showed little or no telomerase activity or ability to maintain growth in the presence of TGFbeta. Cell populations whose mean terminal restriction fragment length had declined to <3 kb also exhibited slow heterogeneous growth, and contained many non-proliferative cells. With continued passage, these conditionally immortal cell populations very gradually converted to a fully immortal phenotype of good growth+/-TGFbeta, expression of high levels of telomerase activity, and stabilization of telomere length. We now show that exposure of the early passage conditionally immortal 184A1 HMEC line to the viral oncogenes human papillomavirus type 16 (HPV16)-E6, -E7, or SV40T, results in either immediate (E6) or rapid (E7; SV40T) conversion of these telomerase negative, TGFbeta sensitive conditionally immortal cells to the fully immortal phenotype. Unlike conditional immortal 184A1, the HPV16-E7 and SV40T exposed cells were able to maintain growth in TGFbeta prior to expression of high levels of telomerase activity. A mutated HPV16-E6 oncogene, unable to inactivate p53, was still capable of rapidly converting conditional immortal 184A1. Our studies provide further evidence that the transforming potential of these viral oncogenes may involve activities beyond their inactivation of p53 and pRB functions. These additional activities may greatly accelerate a step in HMEC immortal transformation, conversion, that would be rate-limiting in the absence of viral oncogene exposure.