Telomeres are repetitive DNA structures at the chromosome en

Telomeres are repetitive DNA structures at the chromosome ends that safeguard them from DNA repair and degradation activities. This protective function requires a proper telomere length and the binding of a six-protein complex known as shelterin, including TRF1, which is essential for telomere protection (Martinez and Blasco, 2010). Telomeres are elongated by telomerase (Greider and Blackburn, 1985), a reverse transcriptase present during early embryonic development and in adult stem cell compartments (Flores et al., 2005, 2006; Liu et al., 2007). However, telomerase activity in adult stem purchase Nocodazole can only partially counterbalance the telomere shortening associated with cell division, so telomeres shorten in all cell types with aging (Flores et al., 2006). Telomere shortening in adult stem cells causes severe impairment of the ability of stem cells to regenerate tissues (Flores et al., 2005). Thus, pathological conditions that require acquisition of cellular immortality, such as many cancer types, show high telomerase activity to allow indefinite proliferation (Martinez and Blasco, 2011). Interestingly, short telomeres in adult differentiated cells can regrow to the long telomeres of embryonic stem cells during in vitro reprogramming to induced pluripotent stem cells (iPSCs) (Marion et al., 2009b; Varela et al., 2011). Telomerase expression and telomere elongation are essential for the efficient generation and pluripotency of the iPSCs, as both the reprogramming efficiency and pluripotency features of these cells are impaired in telomerase-deficient cells with short telomeres (Marion et al., 2009a, 2009b). The shelterin component TRF1 is highly upregulated during iPSC generation, which in turn is essential both for initiation and maintenance of pluripotency in iPSCs (Schneider et al., 2013). Finally, in vitro reprogramming induces epigenetic changes at the telomeric chromatin, including decreased trimethylation of H3K9 and H4K20, indicative of a more “open” chromatin at telomeres (Marion et al., 2009b). These findings suggest substantial changes at telomeres during in vitro iPSC generation. However, whether changes at telomeres occur in association with tissue dedifferentiation induced by in vivo reprogramming or other pathological processes, such as cancer, remains unknown to date. In view of the importance of telomere biology in tissue regeneration, aging, and cancer, here we study telomere changes during tissue dedifferentiation induced by reprogramming in vivo. We find that in vivo reprogrammed areas present longer telomeres and increased expression of the telomerase Terc RNA component than non-reprogrammed tissue, and this telomere elongation is telomerase dependent as it is abolished in Terc-deficient mice. In vivo reprogrammed cells highly overexpress TRF1 in a manner that coincides with OCT4 expression. Chemical inhibition of TRF1 decreases in vivo reprogramming, suggesting an important role of TRF1 upregulation for tissue reprogramming. These telomere-related changes are accompanied by drastic chromatin changes, including loss of histone trimethylation marks. Finally, we extend these findings to pathological tissue dedifferentiation during cancer development. We found elevated TRF1 expression during pancreatic acinar-to-ductal metaplasia (ADM), which involves transdifferentiation of adult acinar cells into ductal-like cells as a result of K-Ras oncogene expression, which can subsequently progress to malignant adenocarcinoma. Telomeres were also elongated in a percentage of the lesions, in a manner uncoupled from TRF1 expression, also mimicking telomere changes during tissue dedifferentiation induced by reprogramming in vivo.
Results
Discussion Here we describe dramatic changes at telomeres during dedifferentiation of mouse adult tissues by induction of in vivo reprogramming, including upregulation of telomerase RNA expression, telomerase-dependent telomere elongation, and upregulation of the TRF1 telomere protein. In addition, we describe dramatic changes in the global structure of chromatin, including decreased heterochromatic marks and decreased expression of the SA1 telomeric cohesin. As increased cellular plasticity and dedifferentiation are also proposed to be important during tissue regeneration as well as in pathological conditions such as cancer, our findings suggest that similar changes at telomeres and global chromatin could also underlie these processes. In support of this notion, we extend the finding of TRF1 upregulation to tissue dedifferentiation during early neoplastic lesions, in a mouse model of pancreatic cancer induced by expression of oncogenic K-Ras. Interestingly, we find increased expression of TRF1 uncoupled from telomere elongation during the acinar-to-ductal transdifferentiation, a process that leads to the initiation of PDAC. Moreover, although a majority of the lesions showed shorter telomeres than the normal surrounding tissue, the remaining 28% showed telomere elongation, in a manner similar to that of in vivo reprogramming-induced dedifferentiation, anticipating the known telomerase upregulation associated with tumorigenesis (Martinez and Blasco, 2011). Thus, here we show analogous changes at telomeres associated with dedifferentiation induced by either in vivo reprogramming or initiation of cancer.