NRF2 regulates serine biosynthesis in non–small cell lung cancer

An Erratum to this article was published on 29 March 2016

This article has been updated

Abstract

Tumors have high energetic and anabolic needs for rapid cell growth and proliferation1, and the serine biosynthetic pathway was recently identified as an important source of metabolic intermediates for these processes2,3. We integrated metabolic tracing and transcriptional profiling of a large panel of non–small cell lung cancer (NSCLC) cell lines to characterize the activity and regulation of the serine/glycine biosynthetic pathway in NSCLC. Here we show that the activity of this pathway is highly heterogeneous and is regulated by NRF2, a transcription factor frequently deregulated in NSCLC. We found that NRF2 controls the expression of the key serine/glycine biosynthesis enzyme genes PHGDH, PSAT1 and SHMT2 via ATF4 to support glutathione and nucleotide production. Moreover, we show that expression of these genes confers poor prognosis in human NSCLC. Thus, a substantial fraction of human NSCLCs activates an NRF2-dependent transcriptional program that regulates serine and glycine metabolism and is linked to clinical aggressiveness.

Access options

Rent or Buy article

Get time limited or full article access on ReadCube.

from$8.99

All prices are NET prices.

Figure 1: Serine biosynthesis activity in lung cancer.
Figure 2: NRF2 regulates serine biosynthesis.
Figure 3: NRF2 regulates the expression of serine/glycine biosynthesis genes through ATF4.
Figure 4: PHGDH-derived serine supports the transsulfuration and folate cycles.
Figure 5: Activation of the serine biosynthesis pathway promotes tumorigenesis in NSCLC.
Figure 6: Model of the regulation of serine/glycine biosynthesis by NRF2.

Change history

  • 15 February 2016

    In the version of this article initially published, the colors of the lines in the key in the top right corner of Figure 5h were incorrect. The line labeled "High" should be red and the line labeled "Low" should be blue. The error has been corrected in the HTML and PDF versions of the article.

References

  1. 1

    Vander Heiden, M.G., Cantley, L.C. & Thompson, C.B. Understanding the Warburg effect: the metabolic requirements of cell proliferation. Science 324, 1029–1033 (2009).

    CAS  Article  Google Scholar 

  2. 2

    Locasale, J.W. et al. Phosphoglycerate dehydrogenase diverts glycolytic flux and contributes to oncogenesis. Nat. Genet. 43, 869–874 (2011).

    CAS  Article  Google Scholar 

  3. 3

    Possemato, R. et al. Functional genomics reveal that the serine synthesis pathway is essential in breast cancer. Nature 476, 346–350 (2011).

    CAS  Article  Google Scholar 

  4. 4

    Mullarky, E., Mattaini, K.R., Vander Heiden, M.G., Cantley, L.C. & Locasale, J.W. PHGDH amplification and altered glucose metabolism in human melanoma. Pigment Cell Melanoma Res. 24, 1112–1115 (2011).

    CAS  Article  Google Scholar 

  5. 5

    Barretina, J. et al. The Cancer Cell Line Encyclopedia enables predictive modelling of anticancer drug sensitivity. Nature 483, 603–607 (2012).

    CAS  Article  Google Scholar 

  6. 6

    Mootha, V.K. et al. PGC-1α–responsive genes involved in oxidative phosphorylation are coordinately downregulated in human diabetes. Nat. Genet. 34, 267–273 (2003).

    CAS  Article  Google Scholar 

  7. 7

    Subramanian, A. et al. Gene set enrichment analysis: a knowledge-based approach for interpreting genome-wide expression profiles. Proc. Natl. Acad. Sci. USA 102, 15545–15550 (2005).

    CAS  Article  Google Scholar 

  8. 8

    Ye, J. et al. Pyruvate kinase M2 promotes de novo serine synthesis to sustain mTORC1 activity and cell proliferation. Proc. Natl. Acad. Sci. USA 109, 6904–6909 (2012).

    CAS  Article  Google Scholar 

  9. 9

    Miyamoto, N. et al. Transcriptional regulation of activating transcription factor 4 under oxidative stress in retinal pigment epithelial ARPE-19/HPV-16 cells. Invest. Ophthalmol. Vis. Sci. 52, 1226–1234 (2011).

    CAS  Article  Google Scholar 

  10. 10

    Afonyushkin, T. et al. Oxidized phospholipids regulate expression of ATF4 and VEGF in endothelial cells via NRF2-dependent mechanism: novel point of convergence between electrophilic and unfolded protein stress pathways. Arterioscler. Thromb. Vasc. Biol. 30, 1007–1013 (2010).

    CAS  Article  Google Scholar 

  11. 11

    Ye, P. et al. Nrf2- and ATF4-dependent upregulation of xCT modulates the sensitivity of T24 bladder carcinoma cells to proteasome inhibition. Mol. Cell. Biol. 34, 3421–3434 (2014).

    Article  Google Scholar 

  12. 12

    He, C.H. et al. Identification of activating transcription factor 4 (ATF4) as an Nrf2-interacting protein. Implication for heme oxygenase-1 gene regulation. J. Biol. Chem. 276, 20858–20865 (2001).

    CAS  Article  Google Scholar 

  13. 13

    Harding, H.P. et al. Regulated translation initiation controls stress-induced gene expression in mammalian cells. Mol. Cell 6, 1099–1108 (2000).

    CAS  Article  Google Scholar 

  14. 14

    Hayes, J.D. & McMahon, M. NRF2 and KEAP1 mutations: permanent activation of an adaptive response in cancer. Trends Biochem. Sci. 34, 176–188 (2009).

    CAS  Article  Google Scholar 

  15. 15

    Kim, Y.R. et al. Oncogenic NRF2 mutations in squamous cell carcinomas of oesophagus and skin. J. Pathol. 220, 446–451 (2010).

    CAS  Article  Google Scholar 

  16. 16

    Konstantinopoulos, P.A. et al. Keap1 mutations and Nrf2 pathway activation in epithelial ovarian cancer. Cancer Res. 71, 5081–5089 (2011).

    CAS  Article  Google Scholar 

  17. 17

    Seng, S. et al. NRP/B mutations impair Nrf2-dependent NQO1 induction in human primary brain tumors. Oncogene 28, 378–389 (2009).

    CAS  Article  Google Scholar 

  18. 18

    Zhang, P. et al. Loss of Kelch-like ECH-associated protein 1 function in prostate cancer cells causes chemoresistance and radioresistance and promotes tumor growth. Mol. Cancer Ther. 9, 336–346 (2010).

    CAS  Article  Google Scholar 

  19. 19

    Shibata, T. et al. Cancer related mutations in NRF2 impair its recognition by Keap1-Cul3 E3 ligase and promote malignancy. Proc. Natl. Acad. Sci. USA 105, 13568–13573 (2008).

    CAS  Article  Google Scholar 

  20. 20

    Solis, L.M. et al. Nrf2 and Keap1 abnormalities in non–small cell lung carcinoma and association with clinicopathologic features. Clin. Cancer Res. 16, 3743–3753 (2010).

    CAS  Article  Google Scholar 

  21. 21

    Singh, A. et al. RNAi-mediated silencing of nuclear factor erythroid-2–related factor 2 gene expression in non–small cell lung cancer inhibits tumor growth and increases efficacy of chemotherapy. Cancer Res. 68, 7975–7984 (2008).

    CAS  Article  Google Scholar 

  22. 22

    DeNicola, G.M. et al. Oncogene-induced Nrf2 transcription promotes ROS detoxification and tumorigenesis. Nature 475, 106–109 (2011).

    CAS  Article  Google Scholar 

  23. 23

    Ohta, T. et al. Loss of Keap1 function activates Nrf2 and provides advantages for lung cancer cell growth. Cancer Res. 68, 1303–1309 (2008).

    CAS  Article  Google Scholar 

  24. 24

    Mitsuishi, Y. et al. Nrf2 redirects glucose and glutamine into anabolic pathways in metabolic reprogramming. Cancer Cell 22, 66–79 (2012).

    CAS  Article  Google Scholar 

  25. 25

    Singh, A. et al. Transcription factor NRF2 regulates miR-1 and miR-206 to drive tumorigenesis. J. Clin. Invest. 123, 2921–2934 (2013).

    CAS  Article  Google Scholar 

  26. 26

    Cheng, T. et al. Pyruvate carboxylase is required for glutamine-independent growth of tumor cells. Proc. Natl. Acad. Sci. USA 108, 8674–8679 (2011).

    CAS  Article  Google Scholar 

  27. 27

    Mullen, A.R. et al. Reductive carboxylation supports growth in tumour cells with defective mitochondria. Nature 481, 385–388 (2012).

    CAS  Article  Google Scholar 

  28. 28

    Yuan, M., Breitkopf, S.B., Yang, X. & Asara, J.M. A positive/negative ion-switching, targeted mass spectrometry–based metabolomics platform for bodily fluids, cells, and fresh and fixed tissue. Nat. Protoc. 7, 872–881 (2012).

    CAS  Article  Google Scholar 

  29. 29

    Shedden, K. et al. Gene expression–based survival prediction in lung adenocarcinoma: a multi-site, blinded validation study. Nat. Med. 14, 822–827 (2008).

    CAS  Article  Google Scholar 

  30. 30

    Rädle, B. et al. Metabolic labeling of newly transcribed RNA for high resolution gene expression profiling of RNA synthesis, processing and decay in cell culture. J. Vis. Exp. 10.3791/50195 (8 August 2013).

  31. 31

    Meylan, E. et al. Requirement for NF-κB signalling in a mouse model of lung adenocarcinoma. Nature 462, 104–107 (2009).

    CAS  Article  Google Scholar 

Download references

Acknowledgements

We thank G. Poulogiannis for bioinformatics advice and H. Abbasi, C. Klimko and M. Yuan for technical support with mass spectrometry experiments. This work was supported by US National Institutes of Health grants P01 CA117969 and R01 GM041890 (L.C.C.), R01 CA157996-01 (R.J.D.), 5R01 CA152301 (Y.X.) and P50 CA70907 (J.D.M., I.I.W., Y.X. and K.E.H.) and by Cancer Prevention Research Institute of Texas (CPRIT) funding to J.D.M., Y.X., I.I.W. and K.E.H. (RP110708 and RP120732) and R.J.D. (RP130272). P.-H.C. was supported by a grant from the Welch Foundation to R.J.D. (I-1733). The mass spectrometry work was partially supported by US National Institutes of Health grants 5P30 CA006516 and 5 P01 CA120964 (J.M.A.). G.M.D. was the Malcolm A.S. Moore Hope Funds for Cancer Research Fellow and is supported by the PanCAN/AACR Pathway to Leadership grant.

Author information

Affiliations

Authors

Contributions

G.M.D., R.J.D. and L.C.C. designed the study. G.M.D. and E.M. performed molecular biology experiments. G.M.D., P.-H.C., E.M., J.A.S., Z.H. and J.M.A. performed metabolomics and isotope labeling and analyzed the data. D.W. performed xenograft experiments. H.T. and Y.X. performed bioinformatics analysis. K.E.H., I.I.W. and J.D.M. contributed highly annotated lung cancer cell lines. G.M.D., E.M. and L.C.C. wrote the manuscript. All authors commented on the manuscript.

Corresponding author

Correspondence to Lewis C Cantley.

Ethics declarations

Competing interests

L.C.C. owns equity in, receives compensation from and serves on the Board of Directors and Scientific Advisory Board of Agios Pharmaceuticals. Agios Pharmaceuticals is identifying metabolic pathways in cancer cells and developing drugs to inhibit such enzymes to disrupt tumor cell growth and survival. R.J.D. is on the scientific advisory boards of Agios Pharmaceuticals and Peloton Therapeutics. Peloton Therapeutics is developing drugs to target altered molecular pathways in cancer, including altered metabolism.

Supplementary information

Supplementary Text and Figures

Supplementary Figures 1–22, Supplementary Tables 2–5 and Supplementary Note. (PDF 6832 kb)

Supplementary Table 1

Gene expression correlations with [13C]serine and [13C]glycine labeling at 6 and 24 h. (XLSX 161 kb)

Source data

Rights and permissions

Reprints and Permissions

About this article

Verify currency and authenticity via CrossMark

Cite this article

DeNicola, G., Chen, PH., Mullarky, E. et al. NRF2 regulates serine biosynthesis in non–small cell lung cancer. Nat Genet 47, 1475–1481 (2015). https://doi.org/10.1038/ng.3421

Download citation

Further reading

Search

Quick links

Nature Briefing

Sign up for the Nature Briefing newsletter — what matters in science, free to your inbox daily.

Get the most important science stories of the day, free in your inbox. Sign up for Nature Briefing