A subpopulation of adult skeletal muscle stem cells retains all template DNA strands after cell division.

Satellite cells are adult skeletal muscle stem cells that are quiescent and constitute a poorly defined heterogeneous population. Using transgenic Tg:Pax7-nGFP mice, we show that Pax7-nGFP(Hi) cells are less primed for commitment and have a lower metabolic status and delayed first mitosis compared to Pax7-nGFP(Lo) cells. Pax7-nGFP(Hi) can give rise to Pax7-nGFP(Lo) cells after serial transplantations. Proliferating Pax7-nGFP(Hi) cells exhibit lower metabolic activity, and the majority performs asymmetric DNA segregation during cell division, wherein daughter cells retaining template DNA strands express stem cell markers. Using chromosome orientation-fluorescence in situ hybridization, we demonstrate that all chromatids segregate asymmetrically, whereas Pax7-nGFP(Lo) cells perform random DNA segregation. Therefore, quiescent Pax7-nGFP(Hi) cells represent a reversible dormant stem cell state, and during muscle regeneration, Pax7-nGFP(Hi) cells generate distinct daughter cell fates by asymmetrically segregating template DNA strands to the stem cell. These findings provide major insights into the biology of stem cells that segregate DNA asymmetrically.

Telomerase reverses epidermal hair follicle stem cell defects and loss of long-term survival associated with critically short telomeres.

Organ homeostasis and organismal survival are related to the ability of stem cells to sustain tissue regeneration. As a consequence of accelerated telomerase shortening, telomerase-deficient mice show defective tissue regeneration and premature death. This suggests a direct impact of telomere length and telomerase activity on stem cell biology. We recently found that short telomeres impair the ability of epidermal stem cells to mobilize out of the hair follicle (HF) niche, resulting in impaired skin and hair growth and in the suppression of epidermal stem cell proliferative capacity in vitro. Here, we demonstrate that telomerase reintroduction in mice with critically short telomeres is sufficient to correct epidermal HF stem cell defects. Additionally, telomerase reintroduction into these mice results in a normal life span by preventing degenerative pathologies in the absence of increased tumorigenesis.

In vivo epigenetic reprogramming of primary human colon cancer cells enhances metastases.

How metastases develop is not well understood and no genetic mutations have been reported as specific metastatic drivers. Here we have addressed the idea that epigenetic reprogramming by GLI-regulated pluripotent stemness factors promotes metastases. Using primary human colon cancer cells engrafted in mice, we find that transient expression of OCT4, SOX2, KLF4 +/- cMYC establishes an enhanced pro-metastatic state in the primary tumor that is stable through sequential engraftments and is transmitted through clonogenic cancer stem cells. Metastatic reprogramming alters NANOG methylation and stably boosts NANOG and NANOGP8 expression. Metastases and reprogrammed EMT-like phenotypes require endogenous NANOG, but enhanced NANOG is not sufficient to induce these phenotypes. Finally, reprogrammed tumors enhance GLI2, and we show that GLI2(high) and AXIN2(low), which are markers of the metastatic transition of colon cancers, are prognostic of poor disease outcome in patients. We propose that metastases arise through epigenetic reprogramming of cancer stem cells within primary tumors.

Loss of WNT-TCF addiction and enhancement of HH-GLI1 signalling define the metastatic transition of human colon carcinomas.

Previous studies demonstrate the initiation of colon cancers through deregulation of WNT-TCF signalling. An accepted but untested extension of this finding is that incurable metastatic colon carcinomas (CCs) universally remain WNT-TCF-dependent, prompting the search for WNT-TCF inhibitors. CCs and their stem cells also require Hedgehog (HH)-GLI1 activity, but how these pathways interact is unclear. Here we define coincident high-to-low WNT-TCF and low-to-high HH-GLI transitions in patient CCs, most strikingly in their CD133(+) stem cells, that mark the development of metastases. We find that enhanced HH-GLI mimics this transition, driving also an embryonic stem (ES)-like stemness signature and that GLI1 can be regulated by multiple CC oncogenes. The data support a model in which the metastatic transition involves the acquisition or enhancement of a more primitive ES-like phenotype, and the downregulation of the early WNT-TCF programme, driven by oncogene-regulated high GLI1 activity. Consistently, TCF blockade does not generally inhibit tumour growth; instead, it, like enhanced HH-GLI, promotes metastatic growth in vivo. Treatments for metastatic disease should therefore block HH-GLI1 but not WNT-TCF activities.

MSH2 deficiency abolishes the anticancer and pro-aging activity of short telomeres.

Mutations in the mismatch repair (MMR) pathway occur in human colorectal cancers with microsatellite instability. Mounting evidence suggests that cell-cycle arrest in response to a number of cellular stresses, including telomere shortening, is a potent anticancer barrier. The telomerase-deficient mouse model illustrates the anticancer effect of cell-cycle arrest provoked by short telomeres. Here, we describe a role for the MMR protein, MSH2, in signaling cell-cycle arrest in a p21/p53-dependent manner in response to short telomeres in the context of telomerasedeficient mice. In particular, progressively shorter telomeres at successive generations of MSH2(-/-) Terc(-/--) mice did not suppress cancer in these mice, indicating that MSH2 deficiency abolishes the tumor suppressor activity of short telomeres. Interestingly, MSH2 deficiency prevented degenerative pathologies in the gastrointestinal tract of MSH2(-/-) Terc(-/-) mice concomitant with a rescue of proliferative defects. The abolishment of the anticancer and pro-aging effects of short telomeres provoked by MSH2 abrogation was independent of changes in telomere length. These results highlight a role for MSH2 in the organismal response to dysfunctional telomeres, which in turn may be important in the pathobiology of human cancers bearing mutations in the MMR pathway.

Deficient mismatch repair improves organismal fitness and survival of mice with dysfunctional telomeres.

Mismatch repair (MMR) has important roles in meiotic and mitotic recombination, DNA damage signaling, and various aspects of DNA metabolism including class-switch recombination, somatic hypermutation, and triplet-repeat expansion. Defects in MMR are responsible for human cancers characterized by microsatellite instability. Intriguingly, MMR deficiency has been shown to rescue survival and proliferation of telomerase-deficient yeast strains. A putative role for MMR at mammalian telomeres that could have an impact on cancer and aging is, however, unknown. Here, we studied the role of MMR in response to dysfunctional telomeres by generating mice doubly deficient for telomerase and the PMS2 MMR gene (Terc-/-/PMS2-/- mice). PMS2 deficiency prolonged the mean lifespan and median survival of telomerase-deficient mice concomitant with rescue of degenerative pathologies. This rescue of survival was independent of changes in telomere length, in sister telomere recombination, and in microsatellite instability. Importantly, PMS2 deficiency rescued cell proliferation defects but not apoptotic defects in vivo, concomitant with a decreased p21 induction in response to short telomeres. The proliferative advantage conferred to telomerase-deficient cells by the ablation of PMS2 did not produce increased tumors. Indeed, Terc-/-/PMS2-/- mice showed reduced tumors compared with PMS2-/- mice, in agreement with a tumor suppressor role for short telomeres in the context of MMR deficiencies. These results highlight an unprecedented role for MMR in mediating the cellular response to dysfunctional telomeres in vivo by attenuating p21 induction.