Abstract
Suppression of anoikis after detachment of cancer cells from the extracellular matrix is a key step during metastasis. Here we show that, after detachment, mouse embryonic fibroblasts (MEFs) transformed by K-Ras(V12) or ETV6-NTRK3 (EN) activate a transcriptional response overrepresented by genes related to bioenergetic stress and the AMP-activated protein kinase (AMPK) energy-sensing pathway. Accordingly, AMPK is activated in both transformed and non-transformed cells after detachment, and AMPK deficiency restores anoikis to transformed MEFs. However, AMPK activation represses the mTOR complex-1 (mTORC1) pathway only in transformed cells, suggesting a key role for AMPK-mediated mTORC1 inhibition in the suppression of anoikis. Consistent with this, AMPK−/− MEFs transformed by EN or K-Ras show sustained mTORC1 activation after detachment and fail to suppress anoikis. Transformed TSC1−/− MEFs, which are incapable of suppressing mTORC1, also undergo anoikis after detachment, which is reversed by mTORC1 inhibitors. Furthermore, transformed AMPK−/− and TSC1−/− MEFs both have higher total protein synthesis rates than wild-type controls, and translation inhibition using cycloheximide partially restores their anoikis resistance, indicating a mechanism whereby mTORC1 inhibition suppresses anoikis. Finally, breast carcinoma cell lines show similar detachment-induced AMPK/mTORC1 activation and restoration of anoikis by AMPK inhibition. Our data implicate AMPK-mediated mTORC1 inhibition and suppression of protein synthesis as a means for bioenergetic conservation during detachment, thus promoting anoikis resistance.
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Abbreviations
- AMPK:
-
AMP-activated protein kinase
- CHX:
-
cycloheximide
- DN:
-
dominant negative
- EN:
-
ETV6-NTRK3
- FACS:
-
fluorescence-activated cell sorting
- GEP:
-
gene expression profile
- GO:
-
gene ontology
- GSEA:
-
Gene Set Enrichment Analysis
- LDH:
-
lactate dehydrogenase
- MEF:
-
mouse embryonic fibroblast
- mTORC1:
-
mTOR complex-1
- TSC:
-
tuberous sclerosis complex
- wt :
-
wild type.
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Acknowledgements
We thank B Rotblat and C Tognon helpful discussions, and T Tang and A Barokas for technical assistance. This study was funded by the US Department of Defense Grant W81XWH-07-1-0580 (to PHBS and TJT). This work was also supported by funds from the British Columbia Cancer Foundation through generous donations from Team Finn and other generous riders in the Ride to Conquer Cancer. TLN is supported by training fellowships from the Clinician Investigator Program of the Royal College of Physicians of Canada, the Canadian Institutes of Health Research (CIHR), and the Michael Smith Foundation for Health Research. We thank D Kwiatkowski, N Sonenberg, N Mizushima, and R Baserga for MEF cell lines, and W Hahn and N Hay for plasmid constructs.
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Ng, T., Leprivier, G., Robertson, M. et al. The AMPK stress response pathway mediates anoikis resistance through inhibition of mTOR and suppression of protein synthesis. Cell Death Differ 19, 501–510 (2012). https://doi.org/10.1038/cdd.2011.119
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DOI: https://doi.org/10.1038/cdd.2011.119
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