Abstract
BackgroundAutophagy 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.MethodsBECN1 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.ResultsWe 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.ConclusionsTaken 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.Electronic supplementary materialThe online version of this article (doi:10.1186/s13048-015-0182-y) contains supplementary material, which is available to authorized users.
Highlights
Autophagy is a conserved cellular self-digestion mechanism that can either suppress or promote cancer in a context-dependent manner
Despite prevalent single-copy loss of the BECN1 gene, Beclin-1 protein expression is maintained in high-grade serous ovarian tumors and ascites-derived cells Heterozygous loss of the canonical autophagy gene BECN1 is well documented in epithelial ovarian cancer cells [13,14,15,16,17]
In order to assess the relationship between BECN1 copy-number and expression, we interrogated level 3 array comparative genomic hybridization and reverse-phase protein array (RPPA) data generated by The Cancer Genome Atlas (TCGA) from a large number of high-grade serous ovarian tumors (91 % of which are from metastatic, stage III-IV disease; Additional file 2: Table S2)
Summary
Autophagy is a conserved cellular self-digestion mechanism that can either suppress or promote cancer in a context-dependent manner. Macroautophagy (autophagy) is a conserved mechanism for the sequestration of cytoplasmic contents in membrane-bound vesicles and their subsequent lysosomemediated degradation. Correa et al Journal of Ovarian Research (2015) 8:52 is allowed to form and convey activating signals to downstream effectors [3] These effectors are mammalian homologues of autophagy-related (“atg”) proteins originally discovered in yeast [4]. Among the ATGs, BECN1 (mammalian homologue of yeast atg that encodes Beclin-1) was one of the earliest discovered and has been extensively studied It functions in a core complex with Class III PI3K (PI3K C3) [6] and p150 [7] as a canonical initiator of autophagy [8]. Beclin-1 – and autophagy by extension – are thought to be tumor suppressive
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