Different telomere-length dynamics at the inner cell mass versus established embryonic stem (ES) cells

Abstract

Murine embryonic stem (ES) cells have unusually long telomeres, much longer than those in embryonic tissues. Here we address whether hyper-long telomeres are a natural property of pluripotent stem cells, such as those present at the blastocyst inner cell mass (ICM), or whether it is a characteristic acquired by the in vitro expansion of ES cells. We find that ICM cells undergo telomere elongation during the in vitro derivation of ES-cell lines. In vivo analysis shows that the hyper-long telomeres of morula-injected ES cells remain hyper-long at the blastocyst stage and longer than telomeres of the blastocyst ICM. Telomere lengthening during derivation of ES-cell lines is concomitant with a decrease in heterochromatic marks at telomeres. We also found increased levels of the telomere repeat binding factor 1 (TRF1) telomere-capping protein in cultured ICM cells before telomere elongation occurs, coinciding with expression of pluripotency markers. These results suggest that high TRF1 levels are present in pluripotent cells, most likely to ensure proficient capping of the newly synthesized telomeres. These results highlight a previously unnoticed difference between ICM cells at the blastocyst and ES cells, and suggest that abnormally long telomeres in ES cells are likely to result from continuous telomere lengthening of proliferating ICM cells locked at an epigenetic state associated to pluripotency.ouse embryonic stem (ES) cells are pluripotent, proliferate indefinitely, and bear very long telomeres (1-3). ES cells emerge from preimplantation blastocyst-stage embryos (4), but how this process takes place is largely unknown. In previous studies, we observed that telomeres of mouse ES cells were much longer than those of mouse embryonic fibroblasts (MEFs) of the same genetic background (5), which are typically obtained at embryonic day 13.5 (E13.5). This observation raised the issue of whether blastocyst inner cell mass (ICM) cells, which are the natural equivalents of ES cells, also have hyper-long telomeres. If this is the case, then telomeres must shorten during fetal development, despite high telomerase activity (6-8). An alternative explanation emerges, however, that hyper-long telomeres in ES cell are aberrant and may result from the in vitro establishment and expansion of ES cells.ES-like pluripotent stem cells can be generated from differentiated cells (i.e., MEFs) by using defined factors, giving rise the so-called induced pluripotent stem (iPS) cells, which are considered functional equivalents of ES cells (9-16). We recently showed that iPS telomeres increase in length during and after nuclear reprogramming until reaching ES cell hyper-long telomeres. This elongation process occurs concomitantly to lower density of trimethylated histones H3K9 and H4K20 at the telomeric chromatin (5). Furthermore, hyper-long telomeres were not observed in iPS cells derived from first-generation telomerasedeficient MEFs, indicating that they do not originate from a selective reprogramming of a subset of parental cells with v...

Therapeutic effect of androgen therapy in a mouse model of aplastic anemia produced by short telomeres

“…Telomerase activity is high at the blastocyst stage during which telomere length is determined for a given individual. 6,7 Thereafter telomeres progressively shorten throughout life due to the down-regulation of telomerase and the loss of telomeric sequences associated with the replication of DNA ends with every cell division. Consequently, cells reaching a critical short telomere length enter into cellular senescence.…”

Telomerase-mediated telomere elongation from human blastocysts to embryonic stem cells

“…High telomerase activity is an important characteristic of ESCs. Our current knowledge of telomere maintenance in ESCs is largely derived from studies in mouse ESCs (mESCs): telomere elongation has been observed during in vitro derivation of mESCs (Varela et al, 2011); loss of telomerase activity in mESCs resulted in progressive telomere loss, genomic instability, aneuploidy and telomere fusion, eventually leading to reduced growth rates (Niida et al, 1998); telomerase overexpression enhanced self-renewal, improved resistance to apoptosis and increased proliferation in mESCs (Armstrong et al, 2005). Tetraploid embryo complementation experiments have shown that telomerase-deficient mESCs with short telomeres lose their ability to generate complete ESC-pups (Huang et al, 2011).…”

Totipotency: What It Is and What It Is Not

“…Similarly, there have been no reports of generating embryos from a combination of mES and trophoblast stem cells [84]. The failure of aggregated stem cell lines derived thus far to spontaneously form embryos is likely to reflect the fact that stem cell lines have a range of properties [85][86][87] that are not identical either to zygotes [88,89] or to cells that have been freshly isolated from embryos [43][44][45][46][47]. While these negative findings do not strictly rule out the possibility that later-stage blastomeres or pluripotent stem cells ''could'' combine in some way to generate an embryo (it is logically impossible to prove something from a negative finding), they strongly suggest that collective reconstitution of an embryo from cells of a reaggregated morula-stage embryo (Fig.…”

“…However, there is no scientific evidence to date that twinning at the blastocyst stage involves totipotent cells, and multiple studies have concluded exactly the opposite; that is, that blastocyst cells are quite unlike totipotent zygotes on multiple parameters [43][44][45][46][47][85][86][87] and that blastocyst cells are completely incapable of producing a whole embryo when isolated from each other [145][146][147], instead producing only stem cell lines. These observations argue strongly that twinning does not involve totipotent cells, but rather relies on some other developmental mechanism.…”

“…3A) and are clearly not equivalent to stem cells [43][44][45][46][47]. Zygotic transcription commences during the first cell cycle in both mouse [48][49][50][51] and human [52][53][54][55][56][57][58][59] embryos.…”