title: p53-Dependent accelerated senescence induced by ionizing radiation in breast tumour cells.
abstract: Ionizing radiation has been reported to promote accelerated or premature senescence in both normal and tumour cell lines. The current studies were designed to characterize the accelerated senescence response to radiation in the breast tumour cell in terms of its dependence on functional p53 and its relationship to telomerase activity, telomere lengths, expression of human telomerase reverse transcriptase (hTERT, the catalytic subunit of telomerase) and human telomerase RNA (hTR, the RNA subunit of telomerase), as well as the induction of cytogenetic aberrations. Studies were performed in p53 wild-type MCF-7 cells, MCF-7/E6 cells with attenuated p53 function, MDA-MB231 cells with mutant p53 and MCF-7/hTERT cells with constitutive expression of hTERT. Telomerase activity was measured by the telomeric repeat amplification protocol (TRAP assay), telomere lengths by the terminal restriction fragment (TRF) assay, hTR and hTERT expression by reverse transcriptase-polymerase chain reaction (RT-PCR), senescence by beta-galactosidase staining, and apoptosis by TdT-mediated d-UTP-X nick-end labelling (TUNEL assay). Widespread and extensive expression of beta-galactosidase, a marker of cellular senescence, was evident in MCF-7 breast tumour cells following exposure to 10 Gy of ionizing radiation. Radiation did not suppress expression of either hTERT or hTR, alter telomerase activity or induce telomere shortening. Senescence arrest was also observed in irradiated MCF-7/hTERT cells, which have elongated telomeres due to the ectopic expression of the catalytic component of telomerase. In contrast to MCF-7 cells, irradiated MDA-MB231 breast tumour cells and MCF-7/E6 cells failed to senesce and instead demonstrated a delayed apoptotic cell death. Irradiation produced chromosome end associated abnormalities, including end-to-end fusions (an indicator of telomere dysfunction) in MCF-7 cells, MCF-7/hTERT cells, as well as in MCF-7/E6 cells. When cells were maintained in culture following irradiation, proliferative recovery was evident exclusively after senescence while the cell lines which responded to radiation by apoptosis continued to decline in cell number. Accelerated senescence in response to ionizing radiation is p53 dependent and associated with telomer dysfunction but is unrelated to changes in telomerase activity or telomere lengths, expression of hTERT and hTR. In the absence of functional p53, cells are unable to arrest for an extended period, resulting in apoptotic cell death while accelerated senescence in cells expressing p53 is succeeded by proliferative recovery.
authors.affiliation: Department of Pharmacology and Toxicology, Virginia Commonwealth University, Richmond 23298, USA.
authors.forename: K R
authors.initials: KR
authors.lastname: Jones
authors.forename: L W
authors.initials: LW
authors.lastname: Elmore
authors.forename: C
authors.initials: C
authors.lastname: Jackson-Cook
authors.forename: G
authors.initials: G
authors.lastname: Demasters
authors.forename: L F
authors.initials: LF
authors.lastname: Povirk
authors.forename: S E
authors.initials: SE
authors.lastname: Holt
authors.forename: D A
authors.initials: DA
authors.lastname: Gewirtz
mesh_terms: D017209:Apoptosis / Q000528:radiation effects; D001943:Breast Neoplasms / Q000235:genetics / Q000473:pathology / Q000532:radiotherapy*; D002453:Cell Cycle / Q000528:radiation effects; D045744:Cell Line, Tumor; D016922:Cellular Senescence / Q000528:radiation effects*; D002869:Chromosome Aberrations; D004268:DNA-Binding Proteins / Q000378:metabolism; D005260:Female; D006801:Humans; D012313:RNA / Q000032:analysis; D019098:Telomerase / Q000032:analysis / Q000378:metabolism; D016615:Telomere; D016159:Tumor Suppressor Protein p53 / Q000502:physiology*
chemical_list: D004268:DNA-Binding Proteins; D016159:Tumor Suppressor Protein p53; C422952:telomerase RNA; D012313:RNA; D019098:Telomerase