title: Autophagy impairment induces premature senescence in primary human fibroblasts.
abstract: BACKGROUND
  Recent studies have demonstrated that activation of autophagy increases the lifespan of organisms from yeast to flies. In contrast to the lifespan extension effect in lower organisms, it has been reported that overexpression of unc-51-like kinase 3 (ULK3), the mammalian homolog of autophagy-specific gene 1 (ATG1), induces premature senescence in human fibroblasts. Therefore, we assessed whether the activation of autophagy would genuinely induce premature senescence in human cells.
  
  
  RESULTS
  Depletion of ATG7, ATG12, or lysosomal-associated membrane protein 2 (Lamp2) by transfecting siRNA or infecting cells with a virus containing gene-specific shRNA resulted in a senescence-like state in two strains of primary human fibroblasts. Prematurely senescent cells induced by autophagy impairment exhibited the senescent phenotypes, similar to the replicatively senescent cells, such as increased senescence associated β-galactosidase (SA-β-gal) activity, reactive oxygen species (ROS) generation, and accumulation of lipofuscin. In addition, expression levels of ribosomal protein S6 kinase1 (S6K1), p-S6K1, p-S6, and eukaryotic translation initiation factor 4E (eIF4E) binding protein 1 (4E-BP1) in the mammalian target of rapamycin (mTOR) pathway and beclin-1, ATG7, ATG12-ATG5 conjugate, and the sequestosome 1 (SQSTM1/p62) monomer in the autophagy pathway were decreased in both the replicatively and the autophagy impairment-induced prematurely senescent cells. Furthermore, it was found that ROS scavenging by N-acetylcysteine (NAC) and inhibition of p53 activation by pifithrin-α or knockdown of p53 using siRNA, respectively, delayed autophagy impairment-induced premature senescence and restored the expression levels of components in the mTOR and autophagy pathways.
  
  
  CONCLUSIONS
  Taken together, we concluded that autophagy impairment induces premature senescence through a ROS- and p53-dependent manner in primary human fibroblasts.
authors.affiliation: Institute on Aging, Seoul National University College of Medicine, Seoul, Korea.
authors.forename: Hyun Tae
authors.initials: HT
authors.lastname: Kang
authors.forename: Ki Baek
authors.initials: KB
authors.lastname: Lee
authors.forename: Sung Young
authors.initials: SY
authors.lastname: Kim
authors.forename: Hae Ri
authors.initials: HR
authors.lastname: Choi
authors.forename: Sang Chul
authors.initials: SC
authors.lastname: Park
mesh_terms: D000111:Acetylcysteine / Q000494:pharmacology; D048868:Adaptor Proteins, Signal Transducing / Q000378:metabolism; D001343:Autophagy / Q000235:genetics / Q000502:physiology*; D000071192:Autophagy-Related Protein 12; D000071193:Autophagy-Related Protein 7; D052160:Benzothiazoles / Q000494:pharmacology; D015153:Blotting, Western; D018797:Cell Cycle Proteins; D002478:Cells, Cultured; D016922:Cellular Senescence / Q000187:drug effects / Q000235:genetics / Q000502:physiology*; D002648:Child; D005347:Fibroblasts / Q000166:cytology / Q000378:metabolism* / Q000648:ultrastructure; D006801:Humans; D052119:Lysosomal-Associated Membrane Protein 2; D051907:Lysosomal Membrane Proteins / Q000235:genetics / Q000378:metabolism; D008297:Male; D008854:Microscopy, Electron; D010750:Phosphoproteins / Q000378:metabolism; D061251:Primary Cell Culture; D034622:RNA Interference; D017382:Reactive Oxygen Species / Q000378:metabolism; D038762:Ribosomal Protein S6 Kinases, 70-kDa / Q000378:metabolism; D000071456:Sequestosome-1 Protein; D015398:Signal Transduction / Q000235:genetics / Q000502:physiology*; D025841:Small Ubiquitin-Related Modifier Proteins / Q000235:genetics / Q000378:metabolism; D058570:TOR Serine-Threonine Kinases / Q000378:metabolism; D013997:Time Factors; D014050:Toluene / Q000031:analogs & derivatives / Q000494:pharmacology; D016159:Tumor Suppressor Protein p53 / Q000235:genetics / Q000378:metabolism; D044764:Ubiquitin-Activating Enzymes / Q000235:genetics / Q000378:metabolism; D001616:beta-Galactosidase / Q000378:metabolism
chemical_list: C466624:ATG12 protein, human; D048868:Adaptor Proteins, Signal Transducing; D000071192:Autophagy-Related Protein 12; D052160:Benzothiazoles; D018797:Cell Cycle Proteins; C087000:EIF4EBP1 protein, human; C497811:LAMP2 protein, human; D052119:Lysosomal-Associated Membrane Protein 2; D051907:Lysosomal Membrane Proteins; D010750:Phosphoproteins; D017382:Reactive Oxygen Species; C099718:SQSTM1 protein, human; D000071456:Sequestosome-1 Protein; D025841:Small Ubiquitin-Related Modifier Proteins; C495901:TP53 protein, human; D016159:Tumor Suppressor Protein p53; D014050:Toluene; C121565:pifithrin; C546842:MTOR protein, human; D038762:Ribosomal Protein S6 Kinases, 70-kDa; D058570:TOR Serine-Threonine Kinases; C448901:ribosomal protein S6 kinase, 70kD, polypeptide 1; D001616:beta-Galactosidase; C509349:ATG7 protein, human; D000071193:Autophagy-Related Protein 7; D044764:Ubiquitin-Activating Enzymes; D000111:Acetylcysteine