Detachment‐Based Equilibrium of Anoikic Cell Death and Autophagic Cell Survival Through Adaptor Protein p66^Shc

Abstract

Anoikis (detachment-induced cell death) confers a tumor-suppressive function in metastatic cancer cells. Autophagy, a conserved self-degradative process, enhances the anoikis resistance of detached cancer cells by maintaining cellular homeostasis. However, the mechanism of regulating cell fatedecision by balancing anoikis and autophagy has been poorly understood. Our previous studies have shown that the adaptor protein p66 Shc mediates anoikis through RhoA activation and inhibits tumor metastasis in vivo. We also found that p66 Shc depletion mitigates nutrient-deprivation-induced autophagy. These findings suggest p66 Shc may coordinately regulate these two processes. To verify this hypothesis, we investigated the effect of p66 Shc on the cell death of detached lung cancer cells, and measured autophagy markers and autophagic flux. Results showed that p66 Shc depletion significantly inhibited anoikis, and reduced the formation of LC3B-II and the degradation of Sequestosome 1 (SQSTM1, p62) in detachment-induced cells. Using monodansylcadaverine (MDC)-LysoTracker double staining and monomeric Cherry (mCherry)-GFP-LC3 assay, we found that the autophagic flux was also mitigated by p66 Shc depletion. In addition, p66 Shc knockdown increased the formation of full-length X-linked inhibitor of apoptosis (XIAP)-associated factor 1 (XAF1), which enhances anoikis sensitivity. In conclusion, p66 Shc plays an essential role in detachment-based equilibrium of anoikic cell death and autophagic cell survival. Anat Rec, 299:325-333, 2016. V C 2015 Wiley Periodicals, Inc.Abbreviations used: CH 5 collagen-homologous; ECM 5 extracellular matrix; ELISA 5 enzyme linked immunosorbent assay; Erk1/2 5 extracellular signaling-regulated kinase; GAPDH 5 glyceraldehyde 3-phosphate dehydrogenase; LC3B 5 microtubuleassociated protein 1A light chain protein 3B; LKB1 5 liver kinase B1; mCherry 5 monomeric Cherry; MDC 5 monodansylcadaverine; mTOR 5 mammalian target of rapamycin; PE 5 phosphatidylethanolamine; poly-HEMA 5 poly 2-hydroxyethyl methacrylate; PTEN 5 phosphatase and tensin homolog; qRT-PCR 5 quantitative reverse transcription polymerase chain reaction; s.e.m 5 standard error of mean; SH2 5 Src homology 2 5 ; shRNA 5 short hairpin RNA; XAF1 5 X-linked inhibitor of apoptosis (XIAP)-associated factor 1.

Anoikis-Associated Lung Cancer Metastasis: Mechanisms and Therapies

“…During cell detachment, p66 Shc translocated into mitochondria and bound to cytochrome c to release and cause death [55]. On the other hand, in the absence of adhesion conditions, p66 Shc induced autophagy and activated apoptosis-associated protein cleavage of caspase-7 and poly (ADP-ribose) polymerase (PARP) by inhibiting phosphorylation of ERK1/2 [57,58]. DNA methylation in promoter-mediated epigenetic repression of p66 Shc increased cancer cell survival and tumor progression [59].…”

Gq Signaling in Autophagy Control: Between Chemical and Mechanical Cues

“…Loss of cell attachment with the ECM usually results in programmed cell death via anoikis. In some cases, ECM-cell detachment can rapidly activate autophagy allowing for survival and re-attachment to the substrate [ 139 ]. However, it remains elusive the mechanism controlling this process with integrin-mediated adhesions emerging as a critical element [ 140 ].…”