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Normal telomere length and chromosomal end capping in poly(ADP-ribose) polymerase-deficient mice and primary cells despite increased chromosomal instability.

Samper E, Goytisolo FA, Ménissier-de Murcia J, González-Suárez E, Cigudosa JC, de Murcia G, Blasco MA - J. Cell Biol. (2001)

Bottom Line: Similarly, there were no differences in the length of the G-strand overhang.The results presented here indicate that PARP-1 does not play a major role in regulating telomere length or in telomeric end capping, and the chromosomal instability of PARP-1(-/)- primary cells can be explained by the repair defect associated to PARP-1 deficiency.Finally, no interaction between PARP-1 and the telomerase reverse transcriptase subunit, Tert, was found using the two-hybrid assay.

View Article: PubMed Central - PubMed

Affiliation: Department of Immunology and Oncology, Centro Nacional de Biotecnología-CSIC, Campus Cantoblanco, E-28049 Madrid, Spain.

ABSTRACT
Poly(ADP-ribose) polymerase (PARP)-1, a detector of single-strand breaks, plays a key role in the cellular response to DNA damage. PARP-1-deficient mice are hypersensitive to genotoxic agents and display genomic instability due to a DNA repair defect in the base excision repair pathway. A previous report suggested that PARP-1-deficient mice also had a severe telomeric dysfunction consisting of telomere shortening and increased end-to-end fusions (d'Adda di Fagagna, F., M.P. Hande, W.-M. Tong, P.M. Lansdorp, Z.-Q. Wang, and S.P. Jackson. 1999. NAT: Genet. 23:76-80). In contrast to that, and using a panoply of techniques, including quantitative telomeric (Q)-FISH, we did not find significant differences in telomere length between wild-type and PARP-1(-/)- littermate mice or PARP-1(-/)- primary cells. Similarly, there were no differences in the length of the G-strand overhang. Q-FISH and spectral karyotyping analyses of primary PARP-1(-/)- cells showed a frequency of 2 end-to-end fusions per 100 metaphases, much lower than that described previously (d'Adda di Fagagna et al., 1999). This low frequency of end-to-end fusions in PARP-1(-/)- primary cells is accordant with the absence of severe proliferative defects in PARP-1(-/)- mice. The results presented here indicate that PARP-1 does not play a major role in regulating telomere length or in telomeric end capping, and the chromosomal instability of PARP-1(-/)- primary cells can be explained by the repair defect associated to PARP-1 deficiency. Finally, no interaction between PARP-1 and the telomerase reverse transcriptase subunit, Tert, was found using the two-hybrid assay.

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TRF analysis in wild-type, PARP-1+/−, and PARP-1−/− primary cells. TRF analysis of primary BM cells or splenocytes, as indicated, from four wild-type (V110, V64, V66, and V74), two PARP-1+/− (V107 and V67), and five PARP-1−/− (V106, V109, V65, V69, and V 75) littermate mice. TRF analysis of primary (passage 1) MEFs from littermate wild-type (D9) and PARP-1−/− (D7 and D11) embryos is also shown. Notice that TRF signals are similar in size in PARP-1−/−, PARP-1+/−, and wild-type.
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fig2: TRF analysis in wild-type, PARP-1+/−, and PARP-1−/− primary cells. TRF analysis of primary BM cells or splenocytes, as indicated, from four wild-type (V110, V64, V66, and V74), two PARP-1+/− (V107 and V67), and five PARP-1−/− (V106, V109, V65, V69, and V 75) littermate mice. TRF analysis of primary (passage 1) MEFs from littermate wild-type (D9) and PARP-1−/− (D7 and D11) embryos is also shown. Notice that TRF signals are similar in size in PARP-1−/−, PARP-1+/−, and wild-type.

Mentions: Finally, telomere length was also evaluated by Southern blot as an alternative technique to measure telomere length not based on fluorescence. Primary BM cells or splenocytes, as indicated, from four wild-type (V64, V66, V74, and V110), two PARP-1+/− (V67 and V107), and five PARP-1−/− (V65, V69, V75, V106, and V109) littermate mice were subjected to TRF analysis as described (Materials and methods; Fig. 2) . TRF analysis also showed a similar telomere length in primary (passage 1) MEFs from littermate wild-type (D9) and PARP-1−/− (D7 and D11) embryos (Fig. 2).


Normal telomere length and chromosomal end capping in poly(ADP-ribose) polymerase-deficient mice and primary cells despite increased chromosomal instability.

Samper E, Goytisolo FA, Ménissier-de Murcia J, González-Suárez E, Cigudosa JC, de Murcia G, Blasco MA - J. Cell Biol. (2001)

TRF analysis in wild-type, PARP-1+/−, and PARP-1−/− primary cells. TRF analysis of primary BM cells or splenocytes, as indicated, from four wild-type (V110, V64, V66, and V74), two PARP-1+/− (V107 and V67), and five PARP-1−/− (V106, V109, V65, V69, and V 75) littermate mice. TRF analysis of primary (passage 1) MEFs from littermate wild-type (D9) and PARP-1−/− (D7 and D11) embryos is also shown. Notice that TRF signals are similar in size in PARP-1−/−, PARP-1+/−, and wild-type.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2196874&req=5

fig2: TRF analysis in wild-type, PARP-1+/−, and PARP-1−/− primary cells. TRF analysis of primary BM cells or splenocytes, as indicated, from four wild-type (V110, V64, V66, and V74), two PARP-1+/− (V107 and V67), and five PARP-1−/− (V106, V109, V65, V69, and V 75) littermate mice. TRF analysis of primary (passage 1) MEFs from littermate wild-type (D9) and PARP-1−/− (D7 and D11) embryos is also shown. Notice that TRF signals are similar in size in PARP-1−/−, PARP-1+/−, and wild-type.
Mentions: Finally, telomere length was also evaluated by Southern blot as an alternative technique to measure telomere length not based on fluorescence. Primary BM cells or splenocytes, as indicated, from four wild-type (V64, V66, V74, and V110), two PARP-1+/− (V67 and V107), and five PARP-1−/− (V65, V69, V75, V106, and V109) littermate mice were subjected to TRF analysis as described (Materials and methods; Fig. 2) . TRF analysis also showed a similar telomere length in primary (passage 1) MEFs from littermate wild-type (D9) and PARP-1−/− (D7 and D11) embryos (Fig. 2).

Bottom Line: Similarly, there were no differences in the length of the G-strand overhang.The results presented here indicate that PARP-1 does not play a major role in regulating telomere length or in telomeric end capping, and the chromosomal instability of PARP-1(-/)- primary cells can be explained by the repair defect associated to PARP-1 deficiency.Finally, no interaction between PARP-1 and the telomerase reverse transcriptase subunit, Tert, was found using the two-hybrid assay.

View Article: PubMed Central - PubMed

Affiliation: Department of Immunology and Oncology, Centro Nacional de Biotecnología-CSIC, Campus Cantoblanco, E-28049 Madrid, Spain.

ABSTRACT
Poly(ADP-ribose) polymerase (PARP)-1, a detector of single-strand breaks, plays a key role in the cellular response to DNA damage. PARP-1-deficient mice are hypersensitive to genotoxic agents and display genomic instability due to a DNA repair defect in the base excision repair pathway. A previous report suggested that PARP-1-deficient mice also had a severe telomeric dysfunction consisting of telomere shortening and increased end-to-end fusions (d'Adda di Fagagna, F., M.P. Hande, W.-M. Tong, P.M. Lansdorp, Z.-Q. Wang, and S.P. Jackson. 1999. NAT: Genet. 23:76-80). In contrast to that, and using a panoply of techniques, including quantitative telomeric (Q)-FISH, we did not find significant differences in telomere length between wild-type and PARP-1(-/)- littermate mice or PARP-1(-/)- primary cells. Similarly, there were no differences in the length of the G-strand overhang. Q-FISH and spectral karyotyping analyses of primary PARP-1(-/)- cells showed a frequency of 2 end-to-end fusions per 100 metaphases, much lower than that described previously (d'Adda di Fagagna et al., 1999). This low frequency of end-to-end fusions in PARP-1(-/)- primary cells is accordant with the absence of severe proliferative defects in PARP-1(-/)- mice. The results presented here indicate that PARP-1 does not play a major role in regulating telomere length or in telomeric end capping, and the chromosomal instability of PARP-1(-/)- primary cells can be explained by the repair defect associated to PARP-1 deficiency. Finally, no interaction between PARP-1 and the telomerase reverse transcriptase subunit, Tert, was found using the two-hybrid assay.

Show MeSH
Related in: MedlinePlus