Antioxidant and oncogene rescue of metabolic defects caused by loss of matrix attachment

Abstract

Normal epithelial cells require matrix attachment for survival and the ability of tumour cells to survive outside their natural extracellular matrix (ECM) niches is dependent on acquisition of anchorage independence1. While apoptosis is the most rapid mechanism for eliminating cells lacking appropriate ECM attachment2, recent reports suggest that non-apoptotic death processes prevent survival when apoptosis is inhibited in matrix-deprived cells3 , 4. Here we demonstrate that detachment of mammary epithelial cells from ECM causes an ATP deficiency due to loss of glucose transport. Overexpression of ErbB2 rescues the ATP deficiency by restoring glucose uptake through stabilization of EGFR and PI(3)K activation and this rescue is dependent on glucose-stimulated flux through the antioxidant-generating pentose phosphate pathway (PPP). Interestingly, we found that the ATP deficiency could be rescued by antioxidant treatment without rescue of glucose uptake. This rescue was found to be dependent on stimulation of fatty acid oxidation (FAO), which is inhibited by detachment-induced reactive oxygen species (ROS). The significance of these findings was supported by evidence of an elevation in ROS in matrix-deprived cells in the luminal space of mammary acini and that antioxidants facilitate the survival of these cells and enhance anchorageindependent colony formation. These results reveal both the importance of matrix attachment in regulating metabolic activity and an unanticipated mechanism for cell survival in altered matrix environments through antioxidant restoration of ATP generation.Epithelial cells are dependent on interactions with specific extracellular matrix (ECM) components for survival, proliferation, and differentiation functions 5 . Loss of matrix attachment of cultured epithelial cells activates a caspase-mediated apoptotic program known as anoikis2. In glandular cancers, like breast cancer, tumour cells are displaced from their * To whom correspondence should be addressed: Department of Cell Biology, Harvard Medical School, 240 Longwood Avenue, Boston, MA 02115, Phone: 617-432-3974, Fax: 617-432-3969, joan_brugge@hms.harvard.edu. 4 Present address: Department of Biological Sciences, University of Notre Dame, Notre Dame, IN 46556 † These authors contributed equally to this work.Supplementary information is linked to the online version of the paper at www.nature.com/nature. A figure summarising the main result of this paper ( Supplementary Fig. 1) is included in the supplementary information.Author Contributions Z.T.S. and J.S.B. were responsible for the overall study design. Z.T.S., A.R.G., H.Y.I., and S.G. conducted experiments. L.S. and Z.J. conducted the experiments measuring native fluorescence of NAD(P)H in 3D cell culture. Z.G.-H. and P.P. designed the fatty acid oxidation studies and Z.T.S. and Z.G.-H. conducted the fatty acid oxidation assays. Z.T.S. and J.S.B. drafted the manuscript and all other authors made revisions. Author InformationReprints and permissions information is available...

Anoikis evasion in inflammatory breast cancer cells is mediated by Bim-EL sequestration

“…Previous studies have revealed a role for the ERK/MAPK pathway in the evasion of anoikis. 21,24,27,32 In order to assess the activation of ERK signaling in IBC cells, we examined phospho-ERK levels in KPL-4 cells deficient in ErbB2 or SUM149 cells deficient in EGFR. In each case, we discovered that the deficiency in receptor tyrosine kinase expression (i.e., ErbB2 in KPL-4 and EGFR in SUM149) resulted in a concomitant loss of phospho-ERK (Figure 3a).…”

“…Perhaps this defect could be overcome through changes in glucose or fatty acid metabolism that have previously been reported in ECM-detached cells. 24,48 Thus it will be especially important to examine metabolic changes in IBC cells to understand how these cells deal with metabolic needs during ECM detachment. Nonetheless, the data presented here provide significant and novel insight into the molecular mechanisms utilized by IBC cells to survive in the absence of ECM attachment.…”

“…[18][19][20] Interestingly, previous studies have shown that ErbB2 and EGFR, which are hyperactivated in a substantial percentage of IBC patients, 21 can effectively antagonize the anoikis program to facilitate anchorage-independent growth. [22][23][24][25][26][27][28] However, a detailed examination of the molecular mechanisms underlying anoikis inhibition in IBC cells has yet to be completed. In this study, we demonstrate that signaling from EGFR and ErbB2 through ERK/MAPK has a major role in the ability of IBC cells to survive in the absence of ECM attachment.…”

Regulatory crosstalk of the metabolic network

“…Recent results, however, indicate that the underlying metabolic transition (the Warburg transition) could be essential for cancer cells to survive outside their natural extracellular matrix (ECM) niches. Rerouting of the carbohydrate flux from glycolysis to the pentose phosphate pathway (PPP) is essential for survival of detached cells and is independent of their energy status [56].…”

Flavonoid diosmetin increases ATP levels in kidney cells and relieves ATP depleting effect of ochratoxin A

“…The impact is so effective that 15 lM DIOS restore completely the intracellular ATP levels. Since ATP synthesis can be influenced by the oxidative status of the cells [46] antioxidant effect of DIOS was also tested applying the same conditions. 15 lM OTA caused an about 60% increase of ROS production (Fig.…”