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Impact of telomerase ablation on organismal viability, aging, and tumorigenesis in mice lacking the DNA repair proteins PARP-1, Ku86, or DNA-PKcs.

Espejel S, Klatt P, Ménissier-de Murcia J, Martín-Caballero J, Flores JM, Taccioli G, de Murcia G, Blasco MA - J. Cell Biol. (2004)

Bottom Line: First, we show that abrogation of PARP-1 in the absence of telomerase does not affect the rate of telomere shortening, telomere capping, or organismal viability compared with single telomerase-deficient controls.In contrast, mice doubly deficient for telomerase and either Ku86 or DNA-PKcs manifest accelerated loss of organismal viability compared with single telomerase-deficient mice.These results support the notion that absence of telomerase and short telomeres in combination with DNA repair deficiencies accelerate the aging process without impacting on tumorigenesis.

View Article: PubMed Central - PubMed

Affiliation: Molecular Oncology Program, Spanish National Cancer Center (CNIO), E-28029 Madrid, Spain.

ABSTRACT
The DNA repair proteins poly(ADP-ribose) polymerase-1 (PARP-1), Ku86, and catalytic subunit of DNA-PK (DNA-PKcs) have been involved in telomere metabolism. To genetically dissect the impact of these activities on telomere function, as well as organismal cancer and aging, we have generated mice doubly deficient for both telomerase and any of the mentioned DNA repair proteins, PARP-1, Ku86, or DNA-PKcs. First, we show that abrogation of PARP-1 in the absence of telomerase does not affect the rate of telomere shortening, telomere capping, or organismal viability compared with single telomerase-deficient controls. Thus, PARP-1 does not have a major role in telomere metabolism, not even in the context of telomerase deficiency. In contrast, mice doubly deficient for telomerase and either Ku86 or DNA-PKcs manifest accelerated loss of organismal viability compared with single telomerase-deficient mice. Interestingly, this loss of organismal viability correlates with proliferative defects and age-related pathologies, but not with increased incidence of cancer. These results support the notion that absence of telomerase and short telomeres in combination with DNA repair deficiencies accelerate the aging process without impacting on tumorigenesis.

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Effect of Ku86, DNA-PKcs, or PARP-1 deficiency on morphology, apoptosis, and proliferative potential of the intestinal epithelium in successive generations of telomerase-deficient mice. Representative photomicrographs of paraffin sections of large intestine from 1-yr-old mice of the indicated genotype stained with Harris hematoxylin and eosin (A), immunostained for active caspase-3 (B), and Ki67 (C). Arrowheads indicate examples of positive immunostaining for active caspase-3 and Ki67. ML, muscular layer; C, crypt; F, fold.
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fig4: Effect of Ku86, DNA-PKcs, or PARP-1 deficiency on morphology, apoptosis, and proliferative potential of the intestinal epithelium in successive generations of telomerase-deficient mice. Representative photomicrographs of paraffin sections of large intestine from 1-yr-old mice of the indicated genotype stained with Harris hematoxylin and eosin (A), immunostained for active caspase-3 (B), and Ki67 (C). Arrowheads indicate examples of positive immunostaining for active caspase-3 and Ki67. ML, muscular layer; C, crypt; F, fold.

Mentions: To further determine whether the severe degenerative lesions observed in double mutant mice were caused by an increase in apoptosis or, alternatively, by a loss of proliferative potential, we conducted a more detailed analysis of the intestinal epithelium (Fig. 4). Cross sections of large intestine from wild-type mice (Fig. 4 A) show an intact submucosa, muscular layer (ML), and mucosa with well-formed crypts (C) and transverse mucosal folds (F). Single deficiency of Ku86, DNA-PKcs, or PARP-1 occasionally caused mild atrophy of the mucosa. Similarly, ablation of telomerase in G2-Terc−/− mice resulted in detectable, albeit not severe, atrophy of the mucosal layer in the large intestine. Importantly, concomitant abrogation of telomerase and of Ku68, DNA-PKcs, or PARP further exacerbated the phenotype of intestinal atrophy (Fig. 3, B, E, and H). In many cases, signs of degeneration included severe glandular depletion with a loss of mucosal architecture and ulcers of the mucosa with submucosal inflammation and glandular cysts (see Fig. 4 A for a section from a Terc/DNA-PKcs double mouse). Staining for active caspase-3, a marker for apoptotic cells, suggested the virtual absence of apoptotic cells (<1 cell out of 100 cells in the intestinal epithelium) both in telomerase proficient wild-type and single mutant Ku86−/−, DNA-PKcs−/−, and PARP-1−/− tissues (Fig. 4 B). As shown in Fig. 4 B, positive staining for active caspase-3 was barely detectable in intestinal sections from Terc−/− single mutants or Terc−/−/PARP-1−/− double knockout mice. We have recently reported that deletion of Ku86 and DNA-PKcs abrogates apoptosis in telomerase-deficient male germ cells (Espejel et al., 2002a,b). Therefore, it is tempting to speculate that the combined loss of telomerase and Ku86 or DNA-PKcs will not provoke apoptosis in the intestine. In fact, we could not observe any significant amount of cells staining positive for caspase-3, neither in Terc−/−/Ku86−/− nor in Terc−/−/DNA-PKcs−/− double mutant mice.


Impact of telomerase ablation on organismal viability, aging, and tumorigenesis in mice lacking the DNA repair proteins PARP-1, Ku86, or DNA-PKcs.

Espejel S, Klatt P, Ménissier-de Murcia J, Martín-Caballero J, Flores JM, Taccioli G, de Murcia G, Blasco MA - J. Cell Biol. (2004)

Effect of Ku86, DNA-PKcs, or PARP-1 deficiency on morphology, apoptosis, and proliferative potential of the intestinal epithelium in successive generations of telomerase-deficient mice. Representative photomicrographs of paraffin sections of large intestine from 1-yr-old mice of the indicated genotype stained with Harris hematoxylin and eosin (A), immunostained for active caspase-3 (B), and Ki67 (C). Arrowheads indicate examples of positive immunostaining for active caspase-3 and Ki67. ML, muscular layer; C, crypt; F, fold.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2172587&req=5

fig4: Effect of Ku86, DNA-PKcs, or PARP-1 deficiency on morphology, apoptosis, and proliferative potential of the intestinal epithelium in successive generations of telomerase-deficient mice. Representative photomicrographs of paraffin sections of large intestine from 1-yr-old mice of the indicated genotype stained with Harris hematoxylin and eosin (A), immunostained for active caspase-3 (B), and Ki67 (C). Arrowheads indicate examples of positive immunostaining for active caspase-3 and Ki67. ML, muscular layer; C, crypt; F, fold.
Mentions: To further determine whether the severe degenerative lesions observed in double mutant mice were caused by an increase in apoptosis or, alternatively, by a loss of proliferative potential, we conducted a more detailed analysis of the intestinal epithelium (Fig. 4). Cross sections of large intestine from wild-type mice (Fig. 4 A) show an intact submucosa, muscular layer (ML), and mucosa with well-formed crypts (C) and transverse mucosal folds (F). Single deficiency of Ku86, DNA-PKcs, or PARP-1 occasionally caused mild atrophy of the mucosa. Similarly, ablation of telomerase in G2-Terc−/− mice resulted in detectable, albeit not severe, atrophy of the mucosal layer in the large intestine. Importantly, concomitant abrogation of telomerase and of Ku68, DNA-PKcs, or PARP further exacerbated the phenotype of intestinal atrophy (Fig. 3, B, E, and H). In many cases, signs of degeneration included severe glandular depletion with a loss of mucosal architecture and ulcers of the mucosa with submucosal inflammation and glandular cysts (see Fig. 4 A for a section from a Terc/DNA-PKcs double mouse). Staining for active caspase-3, a marker for apoptotic cells, suggested the virtual absence of apoptotic cells (<1 cell out of 100 cells in the intestinal epithelium) both in telomerase proficient wild-type and single mutant Ku86−/−, DNA-PKcs−/−, and PARP-1−/− tissues (Fig. 4 B). As shown in Fig. 4 B, positive staining for active caspase-3 was barely detectable in intestinal sections from Terc−/− single mutants or Terc−/−/PARP-1−/− double knockout mice. We have recently reported that deletion of Ku86 and DNA-PKcs abrogates apoptosis in telomerase-deficient male germ cells (Espejel et al., 2002a,b). Therefore, it is tempting to speculate that the combined loss of telomerase and Ku86 or DNA-PKcs will not provoke apoptosis in the intestine. In fact, we could not observe any significant amount of cells staining positive for caspase-3, neither in Terc−/−/Ku86−/− nor in Terc−/−/DNA-PKcs−/− double mutant mice.

Bottom Line: First, we show that abrogation of PARP-1 in the absence of telomerase does not affect the rate of telomere shortening, telomere capping, or organismal viability compared with single telomerase-deficient controls.In contrast, mice doubly deficient for telomerase and either Ku86 or DNA-PKcs manifest accelerated loss of organismal viability compared with single telomerase-deficient mice.These results support the notion that absence of telomerase and short telomeres in combination with DNA repair deficiencies accelerate the aging process without impacting on tumorigenesis.

View Article: PubMed Central - PubMed

Affiliation: Molecular Oncology Program, Spanish National Cancer Center (CNIO), E-28029 Madrid, Spain.

ABSTRACT
The DNA repair proteins poly(ADP-ribose) polymerase-1 (PARP-1), Ku86, and catalytic subunit of DNA-PK (DNA-PKcs) have been involved in telomere metabolism. To genetically dissect the impact of these activities on telomere function, as well as organismal cancer and aging, we have generated mice doubly deficient for both telomerase and any of the mentioned DNA repair proteins, PARP-1, Ku86, or DNA-PKcs. First, we show that abrogation of PARP-1 in the absence of telomerase does not affect the rate of telomere shortening, telomere capping, or organismal viability compared with single telomerase-deficient controls. Thus, PARP-1 does not have a major role in telomere metabolism, not even in the context of telomerase deficiency. In contrast, mice doubly deficient for telomerase and either Ku86 or DNA-PKcs manifest accelerated loss of organismal viability compared with single telomerase-deficient mice. Interestingly, this loss of organismal viability correlates with proliferative defects and age-related pathologies, but not with increased incidence of cancer. These results support the notion that absence of telomerase and short telomeres in combination with DNA repair deficiencies accelerate the aging process without impacting on tumorigenesis.

Show MeSH
Related in: MedlinePlus