Dataset Information

Telomere dysfunction and DNA-PKcs deficiency: characterization and consequence.

ABSTRACT: The mechanisms by which cells accurately distinguish between DNA double-strand break (DSB) ends and telomeric DNA ends remain poorly defined. Recent investigations have revealed intriguing interactions between DNA repair and telomeres. We were the first to report a requirement for the nonhomologous end-joining (NHEJ) protein DNA-dependent protein kinase (DNA-PK) in the effective end-capping of mammalian telomeres. Here, we report our continued characterization of uncapped (as opposed to shortened) dysfunctional telomeres in cells deficient for the catalytic subunit of DNA-PK (DNA-PKcs) and shed light on their consequence. We present evidence in support of our model that uncapped telomeres in this repair-deficient background are inappropriately detected and processed as DSBs and thus participate not only in spontaneous telomere-telomere fusion but, importantly, also in ionizing radiation-induced telomere-DSB fusion events. We show that phosphorylation of DNA-PKcs itself (Thr-2609 cluster) is a critical event for proper telomere end-processing and that ligase IV (NHEJ) is required for uncapped telomere fusion. We also find uncapped telomeres in cells from the BALB/c mouse, which harbors two single-nucleotide polymorphisms that result in reduced DNA-PKcs abundance and activity, most markedly in mammary tissue, and are both radiosensitive and susceptible to radiogenic mammary cancer. Our results suggest mechanistic links between uncapped/dysfunctional telomeres in DNA-PKcs-deficient backgrounds, radiation-induced instability, and breast cancer. These studies provide the first direct evidence of genetic susceptibility and environmental insult interactions leading to a unique and ongoing form of genomic instability capable of driving carcinogenesis.

SUBMITTER: Williams ES

PROVIDER: S-EPMC3595006 | biostudies-literature | 2009 Mar

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

Telomere dysfunction and DNA-PKcs deficiency: characterization and consequence.

Williams Eli S ES Klingler Rebekah R Ponnaiya Brian B Hardt Tanja T Schrock Evelin E Lees-Miller Susan P SP Meek Katheryn K Ullrich Robert L RL Bailey Susan M SM

Cancer research 20090224 5

The mechanisms by which cells accurately distinguish between DNA double-strand break (DSB) ends and telomeric DNA ends remain poorly defined. Recent investigations have revealed intriguing interactions between DNA repair and telomeres. We were the first to report a requirement for the nonhomologous end-joining (NHEJ) protein DNA-dependent protein kinase (DNA-PK) in the effective end-capping of mammalian telomeres. Here, we report our continued characterization of uncapped (as opposed to shortene ...[more]

PMID: 19244120

Similar Datasets

Project description:Poly(ADP-ribose) polymerase 1 (PARP1) interacts genetically with the DNA-dependent protein kinase catalytic subunit (DNA-PKcs) to suppress early-onset T-lineage lymphomas in the mouse, but the underlying mechanisms have remained unknown. To address this question, we analyzed a series of lymphomas arising in PARP1(-/-)/DNA-PKcs(-/-) (P1(-/-)/D(-/-)) mice. We found that, despite defective V(D)J recombination, P1(-/-)/D(-/-) lymphomas lacked clonal reciprocal translocations involving antigen-receptor loci. Instead, tumor cells were characterized by aneuploidy driven by two main mechanisms: p53 inactivation and abnormal chromosome disjunction due to telomere fusions (TFs). Aberrant accumulation of p53 was observed in 13/19 (68.4%) lymphomas. Sequence analysis revealed five p53 mutations: three missense point mutations (one transition in exon 8 and two transversions in exons 5 and 8, respectively), one in-frame 5-11 microindel in exon 7 and a 410-bp deletion encompassing exons 5-8, resulting in a truncated protein. Analysis of tumor metaphases using sequential telomere fluorescent in-situ hybridization and spectral karyotyping revealed that nine out of nine lymphomas contained TFs. Mutant but not wild-type p53 status was associated with frequent clonal and nonclonal TFs, suggesting that p53 normally limits the extent of telomere dysfunction during transformation. Chromosomes involved in TFs were more likely to be aneuploid than chromosomes not involved in TFs in the same metaphases, regardless of the p53 status, indicating that TFs promote aneuploidy via a mechanism that is distinct from p53 loss. Finally, analysis of radiation responses in P1(-/-)/D(-/-), and control primary cells and tissues indicates that loss of PARP1 increases in vivo radiosensitivity and genomic instability in DNA-PKcs-deficient mice without impairing p53 stabilization and effector functions, suggesting a more severe defect in double-strand break (DSB) repair in double mutants. Together, our findings uncover defective DSB repair leading to tumor suppressor inactivation and abnormal segregation of fused chromosomes as two novel mechanisms promoting tumorigenesis in thymocytes lacking PARP1 and DNA-PKcs.

Project description:Clinical application of doxorubicin (DOX) is heavily hindered by DOX cardiotoxicity. Several theories were postulated for DOX cardiotoxicity including DNA damage and DNA damage response (DDR), although the mechanism(s) involved remains to be elucidated. This study evaluated the potential role of TBC domain family member 15 (TBC1D15) in DOX cardiotoxicity. Tamoxifen-induced cardiac-specific Tbc1d15 knockout (Tbc1d15CKO) or Tbc1d15 knockin (Tbc1d15CKI) male mice were challenged with a single dose of DOX prior to cardiac assessment 1 week or 4 weeks following DOX challenge. Adenoviruses encoding TBC1D15 or containing shRNA targeting Tbc1d15 were used for Tbc1d15 overexpression or knockdown in isolated primary mouse cardiomyocytes. Our results revealed that DOX evoked upregulation of TBC1D15 with compromised myocardial function and overt mortality, the effects of which were ameliorated and accentuated by Tbc1d15 deletion and Tbc1d15 overexpression, respectively. DOX overtly evoked apoptotic cell death, the effect of which was alleviated and exacerbated by Tbc1d15 knockout and overexpression, respectively. Meanwhile, DOX provoked mitochondrial membrane potential collapse, oxidative stress and DNA damage, the effects of which were mitigated and exacerbated by Tbc1d15 knockdown and overexpression, respectively. Further scrutiny revealed that TBC1D15 fostered cytosolic accumulation of the cardinal DDR element DNA-dependent protein kinase catalytic subunit (DNA-PKcs). Liquid chromatography-tandem mass spectrometry and co-immunoprecipitation denoted an interaction between TBC1D15 and DNA-PKcs at the segment 594-624 of TBC1D15. Moreover, overexpression of TBC1D15 mutant (∆594-624, deletion of segment 594-624) failed to elicit accentuation of DOX-induced cytosolic retention of DNA-PKcs, DNA damage and cardiomyocyte apoptosis by TBC1D15 wild type. However, Tbc1d15 deletion ameliorated DOX-induced cardiomyocyte contractile anomalies, apoptosis, mitochondrial anomalies, DNA damage and cytosolic DNA-PKcs accumulation, which were canceled off by DNA-PKcs inhibition or ATM activation. Taken together, our findings denoted a pivotal role for TBC1D15 in DOX-induced DNA damage, mitochondrial injury, and apoptosis possibly through binding with DNA-PKcs and thus gate-keeping its cytosolic retention, a route to accentuation of cardiac contractile dysfunction in DOX-induced cardiotoxicity.

Dataset Information

Telomere dysfunction and DNA-PKcs deficiency: characterization and consequence.

Publications

Telomere dysfunction and DNA-PKcs deficiency: characterization and consequence.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets