A tumor-promoting mechanism mediated by retrotransposon-encoded reverse transcriptase is active in human transformed cell lines
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Ilaria Sciamanna1, Alberto Gualtieri1,5, Cristina Cossetti1,5, Emanuele Felice Osimo1, Manuela Ferracin2, Gianfranco Macchia1, Eleonora Aricò1, Gianni Prosseda3, Patrizia Vitullo1, Tom Misteli4 and Corrado Spadafora1
1 Istituto Superiore di Sanità, Viale Regina Elena 299, Rome, Italy.
2 Laboratory for Technologies of Advanced Therapies (LTTA) and Department of Morphology, Surgery and Experimental Medicine, University of Ferrara, Ferrara, Italy.
3 Department of Biology and Biotechnology “Charles Darwin”, Sapienza University, Rome, Italy.
4 National Cancer Institute, NIH, Bethesda MD, USA.
5 Department of Experimental Medicine and Surgery - University of Rome “Tor Vergata”, Rome, Italy.
Corrado Spadafora, email:
Keywords: LINE-1 retrotransposons, reverse transcriptase, transcriptome, miRNAs, DNA:RNA hybrids, cancer genome, reverse transcriptase inhibitor.
Received: September 11, 2013 Accepted: October 12, 2013 Published: October 14, 2013
LINE-1 elements make up the most abundant retrotransposon family in the human genome. Full-length LINE-1 elements encode a reverse transcriptase (RT) activity required for their own retrotranpsosition as well as that of non-autonomous Alu elements. LINE-1 are poorly expressed in normal cells and abundantly in cancer cells. Decreasing RT activity in cancer cells, by either LINE-1-specific RNA interference, or by RT inhibitory drugs, was previously found to reduce proliferation and promote differentiation and to antagonize tumor growth in animal models. Here we have investigated how RT exerts these global regulatory functions.
We report that the RT inhibitor efavirenz (EFV) selectively downregulates proliferation of transformed cell lines, while exerting only mild effects on non-transformed cells; this differential sensitivity matches a differential RT abundance, which is high in the former and undetectable in the latter. Using CsCl density gradients, we selectively identify Alu and LINE-1 containing DNA:RNA hybrid molecules in cancer but not in normal cells. Remarkably, hybrid molecules fail to form in tumor cells treated with EFV under the same conditions that repress proliferation and induce the reprogramming of expression profiles of coding genes, microRNAs (miRNAs) and ultraconserved regions (UCRs). The RT-sensitive miRNAs and UCRs are significantly associated with Alu sequences.
The results suggest that LINE-1-encoded RT governs the balance between single-stranded and double-stranded RNA production. In cancer cells the abundant RT reverse-transcribes retroelement-derived mRNAs forming RNA:DNA hybrids. We propose that this impairs the formation of double-stranded RNAs and the ensuing production of small regulatory RNAs, with a direct impact on gene expression. RT inhibition restores the ‘normal’ small RNA profile and the regulatory networks that depend on them. Thus, the retrotransposon-encoded RT drives a previously unrecognized mechanism crucial to the transformed state in tumor cells.
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