Takamitsu A Kato1, Ryuichi Okayasu, Penny A Jeggo, Akira Fujimori. 1. Heavy-Ion Radiobiology Research Group, Center for Charged Particle Therapy, National Institute of Radiological Sciences, 4-9-1 Anagawa, Inage-ku, Chiba-shi, Chiba, Japan.
Abstract
PURPOSE: In a previous study using HiCEP (High coverage expression profiling), we demonstrated that ASPM (abnormal spindle-like microcephaly-associated) or the most common-type microcephaly (MCPH5) gene was selectively down-regulated by IR (ionizing radiation). The roles of ASPM on radiosensitivity, however, have never been studied. MATERIALS AND METHODS: Using glioblastoma cell lines and normal human fibroblasts, we investigated how IR sensitivity (survived fraction, DNA repair and chromosome aberration) was affected by the reduction of ASPM by specific siRNA (small interfering RNA). RESULTS: Down-regulation of ASPM by siRNA enhanced radiosensitivity in three human cell lines examined. Constant-field gel electrophoreses and γ-H2AX (phosphorylated form of Histone H2A variant H2AX) foci analysis showed that ASPM-specific siRNA impaired DNA double-strand breaks (DSB) in irradiated cells. Elevated levels of abnormal chromosomes were also observed following ASPM siRNA. In addition IR-sensitization by ASPM knockdown was not enhanced in DNA-PK (DNA-dependent protein kinase) deficient glioblastoma cells suggesting that ASPM impacts upon a DNA-PK-dependent pathway. CONCLUSIONS: Our results show for the first time that ASPM is required for efficient non-homologous end-joining in mammalian cells. In clinical applications, ASPM could be a novel target for combination therapy with radiation as well as a useful biomarker for tumor prognosis as ever described.
PURPOSE: In a previous study using HiCEP (High coverage expression profiling), we demonstrated that ASPM (abnormal spindle-like microcephaly-associated) or the most common-type microcephaly (MCPH5) gene was selectively down-regulated by IR (ionizing radiation). The roles of ASPM on radiosensitivity, however, have never been studied. MATERIALS AND METHODS: Using glioblastoma cell lines and normal human fibroblasts, we investigated how IR sensitivity (survived fraction, DNA repair and chromosome aberration) was affected by the reduction of ASPM by specific siRNA (small interfering RNA). RESULTS: Down-regulation of ASPM by siRNA enhanced radiosensitivity in three human cell lines examined. Constant-field gel electrophoreses and γ-H2AX (phosphorylated form of Histone H2A variant H2AX) foci analysis showed that ASPM-specific siRNA impaired DNA double-strand breaks (DSB) in irradiated cells. Elevated levels of abnormal chromosomes were also observed following ASPM siRNA. In addition IR-sensitization by ASPM knockdown was not enhanced in DNA-PK (DNA-dependent protein kinase) deficient glioblastoma cells suggesting that ASPM impacts upon a DNA-PK-dependent pathway. CONCLUSIONS: Our results show for the first time that ASPM is required for efficient non-homologous end-joining in mammalian cells. In clinical applications, ASPM could be a novel target for combination therapy with radiation as well as a useful biomarker for tumor prognosis as ever described.
Authors: Junko Maeda; Allison J Allum; Jacob T Mussallem; Coral E Froning; Alexis H Haskins; Mark A Buckner; Chris D Miller; Takamitsu A Kato Journal: Genes (Basel) Date: 2020-02-25 Impact factor: 4.096
Authors: Scott E Williams; Idoia Garcia; Andrew J Crowther; Shiyi Li; Alyssa Stewart; Hedi Liu; Kendall J Lough; Sean O'Neill; Katherine Veleta; Esteban A Oyarzabal; Joseph R Merrill; Yen-Yu Ian Shih; Timothy R Gershon Journal: Development Date: 2015-10-08 Impact factor: 6.868