OBJECTIVES: This study is to evaluate the effect of separase depletion on cell cycle progression of irradiated and non-irradiated cells through the G(2)/M phases and consecutive cell survival. MATERIALS AND METHODS: Separase was depleted with siRNA in two human non-small cell lung carcinoma (NSCLC) cell lines. Cell cycle progression, mitotic fraction, DNA repair, apoptotic and clonogenic cell death were determined. RESULTS: By depletion of endogenous separase with siRNA in NSCLCs, we showed that separase affects progression through the G(2) phase. In non-irradiated exponentially growing cells, separase depletion led to an increased G(2) accumulation from 17.2% to 29.1% in H460 and from 15.7% to 30.9% in A549 cells and a decrease in mitotic cells. Depletion of separase significantly (P < 0.01) increased the fraction of radiation-induced G(2) arrested cells 30-56 h after irradiation and led to decrease in the mitotic fraction. This was associated with increased double-strand break repair as measured by gamma-H2AX foci kinetics in H460 cells and to a lesser extent in A549 cells. In addition, a decrease in the expression of mitotic linked cell death after irradiation was found. CONCLUSIONS: These results indicate that separase has additional targets involved in regulation of G(2) to M progression after DNA damage. Prolonged G(2) phase arrest in the absence of separase has consequences on repair of damaged DNA and cell death.
OBJECTIVES: This study is to evaluate the effect of separase depletion on cell cycle progression of irradiated and non-irradiated cells through the G(2)/M phases and consecutive cell survival. MATERIALS AND METHODS:Separase was depleted with siRNA in two human non-small cell lung carcinoma (NSCLC) cell lines. Cell cycle progression, mitotic fraction, DNA repair, apoptotic and clonogenic cell death were determined. RESULTS: By depletion of endogenous separase with siRNA in NSCLCs, we showed that separase affects progression through the G(2) phase. In non-irradiated exponentially growing cells, separase depletion led to an increased G(2) accumulation from 17.2% to 29.1% in H460 and from 15.7% to 30.9% in A549 cells and a decrease in mitotic cells. Depletion of separase significantly (P < 0.01) increased the fraction of radiation-induced G(2) arrested cells 30-56 h after irradiation and led to decrease in the mitotic fraction. This was associated with increased double-strand break repair as measured by gamma-H2AX foci kinetics in H460 cells and to a lesser extent in A549 cells. In addition, a decrease in the expression of mitotic linked cell death after irradiation was found. CONCLUSIONS: These results indicate that separase has additional targets involved in regulation of G(2) to M progression after DNA damage. Prolonged G(2) phase arrest in the absence of separase has consequences on repair of damaged DNA and cell death.
Authors: D P Cahill; C Lengauer; J Yu; G J Riggins; J K Willson; S D Markowitz; K W Kinzler; B Vogelstein Journal: Nature Date: 1998-03-19 Impact factor: 49.962
Authors: Juan A Bernal; Rosa Luna; Agueda Espina; Icíar Lázaro; Francisco Ramos-Morales; Francisco Romero; Carmen Arias; Augusto Silva; María Tortolero; José A Pintor-Toro Journal: Nat Genet Date: 2002-09-23 Impact factor: 38.330
Authors: Francisco Romero; Ana M Gil-Bernabé; Carmen Sáez; Miguel A Japón; José A Pintor-Toro; María Tortolero Journal: Mol Cell Biol Date: 2004-04 Impact factor: 4.272