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A large-scale RNA interference screen identifies genes that regulate autophagy at different stages.
Guo, Sujuan; Pridham, Kevin J; Virbasius, Ching-Man; He, Bin; Zhang, Liqing; Varmark, Hanne; Green, Michael R; Sheng, Zhi.
Afiliación
  • Guo S; Virginia Tech Carilion Research Institute, Roanoke, VA, 24016, United States.
  • Pridham KJ; Virginia Tech Carilion Research Institute, Roanoke, VA, 24016, United States.
  • Virbasius CM; Graduate Program in Translational Biology, Medicine, and Health, Virginia Tech, Blacksburg, VA, 24061, United States.
  • He B; Howard Hughes Medical Institute and Department of Molecular, Cell and Cancer Biology, University of Massachusetts Medical School, Worcester, MA, 01605, United States.
  • Zhang L; Department of Computer Science, Virginia Tech, Blacksburg, VA, 24061, United States.
  • Varmark H; Department of Computer Science, Virginia Tech, Blacksburg, VA, 24061, United States.
  • Green MR; The Novo Nordisk Foundation Center for Basic Metabolic Research, University of Copenhagen, Copenhagen, Denmark.
  • Sheng Z; Howard Hughes Medical Institute and Department of Molecular, Cell and Cancer Biology, University of Massachusetts Medical School, Worcester, MA, 01605, United States. Michael.green@umassmed.edu.
Sci Rep ; 8(1): 2822, 2018 02 12.
Article en En | MEDLINE | ID: mdl-29434216
Dysregulated autophagy is central to the pathogenesis and therapeutic development of cancer. However, how autophagy is regulated in cancer is not well understood and genes that modulate cancer autophagy are not fully defined. To gain more insights into autophagy regulation in cancer, we performed a large-scale RNA interference screen in K562 human chronic myeloid leukemia cells using monodansylcadaverine staining, an autophagy-detecting approach equivalent to immunoblotting of the autophagy marker LC3B or fluorescence microscopy of GFP-LC3B. By coupling monodansylcadaverine staining with fluorescence-activated cell sorting, we successfully isolated autophagic K562 cells where we identified 336 short hairpin RNAs. After candidate validation using Cyto-ID fluorescence spectrophotometry, LC3B immunoblotting, and quantitative RT-PCR, 82 genes were identified as autophagy-regulating genes. 20 genes have been reported previously and the remaining 62 candidates are novel autophagy mediators. Bioinformatic analyses revealed that most candidate genes were involved in molecular pathways regulating autophagy, rather than directly participating in the autophagy process. Further autophagy flux assays revealed that 57 autophagy-regulating genes suppressed autophagy initiation, whereas 21 candidates promoted autophagy maturation. Our RNA interference screen identifies identified genes that regulate autophagy at different stages, which helps decode autophagy regulation in cancer and offers novel avenues to develop autophagy-related therapies for cancer.
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Texto completo: 1 Colección: 01-internacional Base de datos: MEDLINE Asunto principal: Autofagia / Leucemia Mielógena Crónica BCR-ABL Positiva Tipo de estudio: Prognostic_studies Límite: Humans Idioma: En Revista: Sci Rep Año: 2018 Tipo del documento: Article País de afiliación: Estados Unidos Pais de publicación: Reino Unido
Texto completo
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Texto completo: 1 Colección: 01-internacional Base de datos: MEDLINE Asunto principal: Autofagia / Leucemia Mielógena Crónica BCR-ABL Positiva Tipo de estudio: Prognostic_studies Límite: Humans Idioma: En Revista: Sci Rep Año: 2018 Tipo del documento: Article País de afiliación: Estados Unidos Pais de publicación: Reino Unido