title: Impaired autophagy due to constitutive mTOR activation sensitizes TSC2-null cells to cell death under stress.
abstract: It has been well documented that cells deficient in either TSC1 or TSC2 are highly sensitive to various cell death stimuli. In this study, we utilized the TSC2 (-/-)  mouse embryonic fibroblasts (MEFs) to study the involvement of autophagy in the enhanced susceptibility of TSC2-null cells to cell death. We first confirmed that both TSC1-null and TSC2-null MEFs are more sensitive to apoptosis in response to amino acid starvation (EBSS) and hypoxia. Second, we found that both the basal and inducible autophagy in TSC2 (-/-)  MEFs is impaired, mainly due to constitutive activation of mTORC1. Third, suppression of autophagy by chloroquine and Atg7 knockdown sensitizes TSC2 (+/+)  cells, but not TSC2 (-/-)  cells, to EBSS-induced cell death. Conversely, the inhibition of mTORC1 by raptor knockdown and rapamycin activates autophagy and subsequently rescues TSC2 (-/-)  cells. Finally, in starved cells, nutrient supplementations (insulin-like growth factor-1 (IGF-1) and leucine) enhanced cell death in TSC2 (-/-)  cells, but reduced cell death in TSC2 (+/+)  cells. Taken together, these data indicate that constitutive activation of mTORC1 in TSC2 (-/-)  cells leads to suppression of autophagy and enhanced susceptibility to stress-mediated cell death. Our findings thus provide new insights into the complex relationships among mTOR, autophagy and cell death, and support the possible autophagy-targeted intervention strategies for the treatment of TSC-related pathologies.
authors.affiliation: Department of Epidemiology and Public Health, Yong Loo Lin School of Medicine, National University of Singapore, Singapore.
authors.forename: Shukie
authors.initials: S
authors.lastname: Ng
authors.forename: You-Tong
authors.initials: YT
authors.lastname: Wu
authors.forename: Bo
authors.initials: B
authors.lastname: Chen
authors.forename: Jing
authors.initials: J
authors.lastname: Zhou
authors.forename: Han-Ming
authors.initials: HM
authors.lastname: Shen
mesh_terms: D000818:Animals; D001343:Autophagy; D000071193:Autophagy-Related Protein 7; D016923:Cell Death; D002460:Cell Line; D002470:Cell Survival; D002738:Chloroquine / Q000494:pharmacology; D005347:Fibroblasts / Q000378:metabolism; D000076222:Mechanistic Target of Rapamycin Complex 1; D051379:Mice; D008869:Microtubule-Associated Proteins / Q000378:metabolism; D046912:Multiprotein Complexes; D011506:Proteins / Q000378:metabolism; D034741:RNA, Small Interfering / Q000378:metabolism; D015398:Signal Transduction; D058570:TOR Serine-Threonine Kinases / Q000378:metabolism*; D013997:Time Factors; D019076:Transgenes
chemical_list: C502661:Atg7 protein, mouse; D008869:Microtubule-Associated Proteins; D046912:Multiprotein Complexes; D011506:Proteins; D034741:RNA, Small Interfering; D002738:Chloroquine; C546843:mTOR protein, mouse; D000076222:Mechanistic Target of Rapamycin Complex 1; D058570:TOR Serine-Threonine Kinases; D000071193:Autophagy-Related Protein 7