title: Beclin-1 expression is retained in high-grade serous ovarian cancer yet is not essential for autophagy induction in vitro.
abstract: BACKGROUND
  Autophagy is a conserved cellular self-digestion mechanism that can either suppress or promote cancer in a context-dependent manner. In ovarian cancer, prevalent mono-allelic deletion of BECN1 (a canonical autophagy-inducer) suggests that autophagy is impaired to promote carcinogenesis and that Beclin-1 is a haploinsufficient tumor suppressor. Nonetheless, autophagy is known to be readily inducible in ovarian cancer cells. We sought to clarify whether Beclin-1 expression is in fact disrupted in ovarian cancer and whether this impacts autophagy regulation.
  
  
  METHODS
  BECN1 expression levels were assessed using The Cancer Genome Atlas (TCGA) datasets from 398 ovarian high-grade serous cystadenocarcinomas (HGSC) and protein immunoblot data from HGSC samples obtained at our institution. Knockdown of BECN1 and other autophagy-related gene expression was achieved using siRNA in established human ovarian cancer cell lines (CaOV3, OVCAR8, SKOV3, and HeyA8) and a novel early-passage, ascites-derived cell line (iOvCa147-E2). LC3 immunoblot, autophagic flux assays, transmission electron microscopy and fluorescence microscopy were used to assess autophagy.
  
  
  RESULTS
  We observed prevalent mono-allelic BECN1 gene deletion (76%) in TCGA tumors, yet demonstrate for the first time that Beclin-1 protein expression remains relatively unaltered in these and additional samples generated at our institution. Surprisingly, efficient siRNA-mediated Beclin-1 knockdown did not attenuate autophagy induction, whereas knockdown of other autophagy-related genes blocked the process. Beclin-1 knockdown instead decreased cell viability without inducing apoptosis.
  
  
  CONCLUSIONS
  Taken together, these data demonstrate that despite its sustained expression, Beclin-1 is dispensable for autophagy induction in ovarian tumor cells in vitro, yet may be retained to promote cell viability by a mechanism independent of autophagy or apoptosis regulation. Overall, this work makes novel observations about tumor expression of Beclin-1 and challenges the accepted understanding of its role in regulating autophagy in ovarian cancer.
authors.affiliation: Translational Ovarian Cancer Research Program, London Regional Cancer Program, London, Ontario, Canada. rcorrea2016@meds.uwo.ca.|Department of Biochemistry, Schulich School of Medicine & Dentistry, The University of Western Ontario, London, Ontario, Canada. rcorrea2016@meds.uwo.ca.
authors.forename: Rohann J M
authors.initials: RJ
authors.lastname: Correa
authors.affiliation: Translational Ovarian Cancer Research Program, London Regional Cancer Program, London, Ontario, Canada. yudithramos@yahoo.es.
authors.forename: Yudith Ramos
authors.initials: YR
authors.lastname: Valdes
authors.affiliation: Translational Ovarian Cancer Research Program, London Regional Cancer Program, London, Ontario, Canada. tshephe6@uwo.ca.|Department of Obstetrics & Gynaecology, Schulich School of Medicine & Dentistry, The University of Western Ontario, London, Ontario, Canada. tshephe6@uwo.ca.|Department of Oncology, Schulich School of Medicine & Dentistry, The University of Western Ontario, London, Ontario, Canada. tshephe6@uwo.ca.|Department of Anatomy & Cell Biology, Schulich School of Medicine & Dentistry, The University of Western Ontario, London, Ontario, Canada. tshephe6@uwo.ca.
authors.forename: Trevor G
authors.initials: TG
authors.lastname: Shepherd
authors.affiliation: Translational Ovarian Cancer Research Program, London Regional Cancer Program, London, Ontario, Canada. dimattia@uwo.ca.|Department of Biochemistry, Schulich School of Medicine & Dentistry, The University of Western Ontario, London, Ontario, Canada. dimattia@uwo.ca.|Department of Obstetrics & Gynaecology, Schulich School of Medicine & Dentistry, The University of Western Ontario, London, Ontario, Canada. dimattia@uwo.ca.|Department of Oncology, Schulich School of Medicine & Dentistry, The University of Western Ontario, London, Ontario, Canada. dimattia@uwo.ca.|London Regional Cancer Program, 790 Commissioners Road East, Room A4-919A, London, Ontario, Canada, N6A 4 L6. dimattia@uwo.ca.
authors.forename: Gabriel E
authors.initials: GE
authors.lastname: DiMattia
mesh_terms: D017209:Apoptosis / Q000235:genetics; D051017:Apoptosis Regulatory Proteins / Q000096:biosynthesis* / Q000235:genetics; D001343:Autophagy / Q000235:genetics*; D000071186:Beclin-1; D045744:Cell Line, Tumor; D002470:Cell Survival / Q000235:genetics; D018284:Cystadenocarcinoma, Serous / Q000235:genetics* / Q000473:pathology; D005260:Female; D015972:Gene Expression Regulation, Neoplastic; D006801:Humans; D008565:Membrane Proteins / Q000096:biosynthesis* / Q000235:genetics; D010051:Ovarian Neoplasms / Q000235:genetics* / Q000473:pathology; D018874:Spheroids, Cellular / Q000473:pathology
chemical_list: D051017:Apoptosis Regulatory Proteins; C491997:BECN1 protein, human; D000071186:Beclin-1; D008565:Membrane Proteins