Abstract
Rejuvenation of telomeres with various lengths has been found in induced pluripotent stem cells (iPSCs). Mechanisms of telomere length regulation during induction and proliferation of iPSCs remain elusive. We show that telomere dynamics are variable in mouse iPSCs during reprogramming and passage, and suggest that these differences likely result from multiple potential factors, including the telomerase machinery, telomerase-independent mechanisms and clonal influences including reexpression of exogenous reprogramming factors. Using a genetic model of telomerase-deficient (Terc−/− and Terc+/−) cells for derivation and passages of iPSCs, we found that telomerase plays a critical role in reprogramming and self-renewal of iPSCs. Further, telomerase maintenance of telomeres is necessary for induction of true pluripotency while the alternative pathway of elongation and maintenance by recombination is also required, but not sufficient. Together, several aspects of telomere biology may account for the variable telomere dynamics in iPSCs. Notably, the mechanisms employed to maintain telomeres during iPSC reprogramming are very similar to those of embryonic stem cells. These findings may also relate to the cloning field where these mechanisms could be responsible for telomere heterogeneity after nuclear reprogramming by somatic cell nuclear transfer.
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Acknowledgements
We thank Dr Susan Bailey's lab for help and advice on CO-FISH experiments. This work was supported by MOST National Major Basic Research Program (2009CB941000), the Ministry of Agriculture of China Transgenic Special Program (2009ZX08006-010B) and NIH 2 R01 DK054369 (XP and HZ).
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( Supplementary information is linked to the online version of the paper on the Cell Research website.)
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Supplementary information, Figure S1
(A) Expression of Tert and Terc by qPCR analysis of N33 (ESCs), MEF(mouse embryonic fibroblasts), WTfb (Tail-tip fibroblasts (TTF) from adult wild-type mice at the age of 8 weeks, and Kofb (Tail-tips fibroblasts from adult telomerase Terc deficient mice). (PDF 53 kb)
Supplementary information, Figure S2
Dynamics of telomerase and telomeres during induction of iPSCs from telomerase Terc knockout (KO, Terc−/−) TTF. (PDF 91 kb)
Supplementary information, Figure S3
Telomerase activity and maintenance of telomeres during passages of telomerase haplo-insufficient heterozygous (Het., Terc+/−) iPSCs. (PDF 71 kb)
Supplementary information, Figure S4
Expression of endogenous and exogenous genes of iPSCs determined by real-time PCR analysis. (PDF 90 kb)
Supplementary information, Figure S5
Relative amount of exogenous transcript as a proportion of total transcripts in iPSCs determined by real-time PCR analysis. (PDF 87 kb)
Supplementary information, Figure S6
Differentiation tests in vitro of Terc-deficient iPSCs at early and late passages. (PDF 150 kb)
Supplementary information, Figure S7
Association of telomere shortening and dysfunction with reduced developmental pluripotency of iPSCs. (PDF 129 kb)
Supplementary information, Table S1
Karyotyping of iPSC lines (PDF 6 kb)
Supplementary information, Table S2
Telomere loss and chromosome fusion of iPSCs at early passage (P6) (PDF 6 kb)
Supplementary information, Table S3
Chimeras of iPSCs generated by injection into 8-cell albino ICR embryos (PDF 6 kb)
Supplementary information, Table S4
Comparison of telomere length and efficiency (%) of chimeras (PDF 6 kb)
Supplementary information, Table S5
Primers for PCR analysis (PDF 45 kb)
Supplementary information, Table S6
Ratio of contribution of iPSCs in the chimeras by microsatellite analysis (PDF 31 kb)
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Wang, F., Yin, Y., Ye, X. et al. Molecular insights into the heterogeneity of telomere reprogramming in induced pluripotent stem cells. Cell Res 22, 757–768 (2012). https://doi.org/10.1038/cr.2011.201
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DOI: https://doi.org/10.1038/cr.2011.201
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