Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

PKCα is required for Akt-mTORC1 activation in non-small cell lung carcinoma (NSCLC) with EGFR mutation

Oncogene volume 38, pages 7311–7328 (2019)Cite this article

Subjects

A Correction to this article was published on 01 October 2019

This article has been updated

Abstract

Mutational activation of the epidermal growth factor receptor (EGFR) is a major player in the pathogenesis of non-small cell lung cancer (NSCLC). NSCLC patients with constitutively active EGFR mutations eventually develop drug resistance against EGFR tyrosine-kinase inhibitors; therefore, better understandings of key components of mutant EGFR (mtEGFR) signaling are required. Here, we initially observed aberrantly high expression of protein kinase Cα (PKCα) in lung adenocarcinomas, especially those with EGFR mutations, and proceeded to examine the role of PKCα in the regulation of the signaling pathways downstream of mtEGFR. The results showed that NSCLC cell lines with constitutively active EGFR mutations tend to have very or moderately high PKCα levels. Furthermore, PKCα was constitutively activated in HCC827 and H4006 cells which have an EGFR deletion mutation in exon 19. Interestingly, mtEGFR was not required for the induction of PKCα at protein and message levels suggesting that the increased levels of PKCα are due to independent selection. On the other hand, mtEGFR activity was required for robust activation of PKCα. Loss of functions studies revealed that the NSCLC cells rely heavily on PKCα for the activation of the mTORC1 signaling pathway. Unexpectedly, the results demonstrated that PKCα was required for activation of Akt upstream of mTOR but only in cells with the mtEGFR and with the increased expression of PKCα. Functionally, inhibition of PKCα in HCC827 led to caspase-3-dependent apoptosis and a significant decrease in cell survival in response to cellular stress induced by serum starvation. In summary, the results identified important roles of PKCα in regulating mTORC1 activity in lung cancer cells, whereby a primary switching occurs from PKCα-independent to PKCα-dependent signaling in the presence of EGFR mutations. The results present PKCα as a potential synergistic target of personalized treatment for NSCLC with constitutively active mutant forms of EGFR and constitutively active PKCα.

This is a preview of subscription content, access via your institution

Access options

Rent or buy this article

Prices vary by article type

from$1.95

to$39.95

Learn more

Prices may be subject to local taxes which are calculated during checkout

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5
Fig. 6
Fig. 7
Fig. 8
Fig. 9

Change history

  • 30 September 2019

    The original HTML version of this Article was updated after publication to include the Supplementary file, as it was previously omitted.

  • 01 October 2019

    An amendment to this paper has been published and can be accessed via a link at the top of the paper.

References

  1. Kanne JP. Screening for lung cancer: what have we learned? Am J Roentgenol. 2014;202:530–5.

    Google Scholar 

  2. Ferlay J, Soerjomataram I, Dikshit R, Eser S, Mathers C, Rebelo M, et al. Cancer incidence and mortality worldwide: sources, methods and major patterns in GLOBOCAN 2012. Int J Cancer. 2015;136:E359–86.

    CAS  Google Scholar 

  3. Molina JR, Yang P, Cassivi SD, Schild SE, Adjei AA (eds). Non-small cell lung cancer: epidemiology, risk factors, treatment, and survivorship. Mayo Clin Proc. 2008;83:584–94.

  4. Shames DS, Wistuba II. The evolving genomic classification of lung cancer. J Pathol. 2014;232:121–33.

    CAS  PubMed  Google Scholar 

  5. Hollstein M, Sidransky D, Vogelstein B, Harris CC. p53 mutations in human cancers. Science. 1991;253:49–53.

    CAS  PubMed  Google Scholar 

  6. Reissmann PT, Koga H, Takahashi R, Figlin RA, Holmes EC, Piantadosi S, et al. Inactivation of the retinoblastoma susceptibility gene in non-small-cell lung cancer. The Lung Cancer Study Group. Oncogene. 1993;8:1913–9.

    CAS  PubMed  Google Scholar 

  7. Jin G, Kim MJ, Jeon H-S, Choi JE, Kim DS, Lee EB, et al. PTEN mutations and relationship to EGFR, ERBB2, KRAS, and TP53 mutations in non-small cell lung cancers. Lung Cancer. 2010;69:279–83.

    PubMed  Google Scholar 

  8. Sordella R, Bell DW, Haber DA, Settleman J. Gefitinib-sensitizing EGFR mutations in lung cancer activate anti-apoptotic pathways. Science. 2004;305:1163–7.

    CAS  PubMed  Google Scholar 

  9. Gazdar A. Activating and resistance mutations of EGFR in non-small-cell lung cancer: role in clinical response to EGFR tyrosine kinase inhibitors. Oncogene. 2009;28:S24–31.

    CAS  PubMed  PubMed Central  Google Scholar 

  10. Cooper WA, Lam DC, O'Toole SA, Minna JD. Molecular biology of lung cancer. J Thorac Dis. 2013;5:S479–90.

    PubMed  PubMed Central  Google Scholar 

  11. Antonicelli A, Cafarotti S, Indini A, Galli A, Russo A, Cesario A, et al. EGFR-targeted therapy for non-small cell lung cancer: focus on EGFR oncogenic mutation. Int J Med Sci. 2013;10:320–30.

    PubMed  PubMed Central  Google Scholar 

  12. Steins M, Thomas M.Geissler M, Recent results in cancer research Fortschritte der Krebsforschung Progres dans les recherches sur le cancer. Erlotinib. 2014;201:109–23.

  13. Steinberg SF. Structural basis of protein kinase C isoform function. Physiol Rev. 2008;88:1341–78.

    CAS  PubMed  PubMed Central  Google Scholar 

  14. Griner EM, Kazanietz MG. Protein kinase C and other diacylglycerol effectors in cancer. Nat Rev Cancer. 2007;7:281–94.

    CAS  PubMed  Google Scholar 

  15. Kong C, Zhu Y, Liu D, Yu M, Li S, Li Z, et al. Role of protein kinase C-alpha in superficial bladder carcinoma recurrence. Urology. 2005;65:1228–32.

    PubMed  Google Scholar 

  16. Gupta A, Galoforo S, Berns C, Martinez A, Corry P, Guan KL, et al. Elevated levels of ERK2 in human breast carcinoma MCF‐7 cells transfected with protein kinase Cα. Cell Prolif. 1996;29:655–63.

    CAS  PubMed  Google Scholar 

  17. Kim S, Han J, Lee SK, Choi M-Y, Kim J, Lee J, et al. Berberine suppresses the TPA-induced MMP-1 and MMP-9 expressions through the inhibition of PKC-α in breast cancer cells. J Surg Res. 2012;176:e21–9.

    CAS  PubMed  Google Scholar 

  18. Oster H, Leitges M. Protein kinase C α but not PKCζ suppresses intestinal tumor formation in ApcMin/+ mice. Cancer Res. 2006;66:6955–63.

    CAS  PubMed  Google Scholar 

  19. Jiang X-H, Tu S-P, Cui J-T, Lin MC, Xia HH, Wong WM, et al. Antisense targeting protein kinase C α and β1 inhibits gastric carcinogenesis. Cancer Res. 2004;64:5787–94.

    CAS  PubMed  Google Scholar 

  20. Fan Q-W, Cheng C, Knight ZA, Haas-Kogan D, Stokoe D, James CD, et al. EGFR signals to mTOR through PKC and independently of Akt in glioma. Sci Signal. 2009;2:ra4.

    PubMed  PubMed Central  Google Scholar 

  21. Smith SD, Enge M, Bao W, Thullberg M, Costa TD, Olofsson H, et al. Protein kinase Calpha (PKCalpha) regulates p53 localization and melanoma cell survival downstream of integrin alphav in three-dimensional collagen and in vivo. J Biol Chem. 2012;287:29336–47.

    CAS  PubMed  PubMed Central  Google Scholar 

  22. Hsieh Y-H, Wu T-T, Huang C-Y, Hsieh Y-S, Hwang J-M, Liu J-Y. p38 mitogen-activated protein kinase pathway is involved in protein kinase Cα–regulated invasion in human hepatocellular carcinoma cells. Cancer Res. 2007;67:4320–7.

    CAS  PubMed  Google Scholar 

  23. Singhal SS, Yadav S, Singhal J, Drake K, Awasthi YC, Awasthi S. The role of PKCα and RLIP76 in transport‐mediated doxorubicin‐resistance in lung cancer. FEBS Lett. 2005;579:4635–41.

    CAS  PubMed  Google Scholar 

  24. O'Neill AK, Gallegos LL, Justilien V, Garcia EL, Leitges M, Fields AP, et al. Protein kinase Cα promotes cell migration through a PDZ-dependent interaction with its novel substrate discs large homolog 1 (DLG1). J Biol Chem. 2011;286:43559–68.

    CAS  PubMed  PubMed Central  Google Scholar 

  25. La Porta CA, Tessitore L, Comolli R. Changes in protein kinase C alpha, delta and in nuclear beta isoform expression in tumour and lung metastatic nodules induced by diethylnitrosamine in the rat. Carcinogenesis. 1997;18:715–9.

    CAS  PubMed  Google Scholar 

  26. Cheng CY, Hsieh HL, Sun CC, Lin CC, Luo SF, Yang CM. IL‐1β induces urokinse‐plasminogen activator expression and cell migration through PKCα, JNK1/2, and NF‐κB in A549 cells. J Cell Physiol. 2009;219:183–93.

    CAS  PubMed  Google Scholar 

  27. Wang C, Wang X, Liang H, Wang T, Yan X, Cao M, et al. miR-203 inhibits cell proliferation and migration of lung cancer cells by targeting PKCα. PLoS ONE. 2013;8:e73985.

    CAS  PubMed  PubMed Central  Google Scholar 

  28. Abera MB, Kazanietz MG. Protein kinase Cα mediates erlotinib resistance in lung cancer cells. Mol Pharm. 2015;87:832–41.

    CAS  Google Scholar 

  29. Lahn M, Su C, Li S, Chedid M, Hanna KR, Graff JR, et al. Expression levels of protein kinase C-α in non–small-cell lung cancer. Clin Lung Cancer. 2004;6:184–9.

    CAS  PubMed  Google Scholar 

  30. Barretina J, Caponigro G, Stransky N, Venkatesan K, Margolin AA, Kim S, et al. The Cancer Cell Line Encyclopedia enables predictive modelling of anticancer drug sensitivity. Nature. 2012;483:603–7.

    CAS  PubMed  PubMed Central  Google Scholar 

  31. El Osta M, Liu M, Adada M, Senkal CE, Idkowiak-Baldys J, Obeid LM, et al. Sustained PKCβII activity confers oncogenic properties in a phospholipase D-and mTOR-dependent manner. FASEB J. 2014;28:495–505.

    CAS  PubMed  PubMed Central  Google Scholar 

  32. Vincent E, Elder D, Thomas E, Phillips L, Morgan C, Pawade J, et al. Akt phosphorylation on Thr308 but not on Ser473 correlates with Akt protein kinase activity in human non-small cell lung cancer. Br J Cancer. 2011;104:1755–61.

    CAS  PubMed  PubMed Central  Google Scholar 

  33. Vansteenkiste J, Ramlau R, Von Pawel J, San Antonio B, Eschbach C, Szczesna A, et al. A phase II randomized study of cisplatin-pemetrexed plus either enzastaurin or placebo in chemonaive patients with advanced non-small cell lung cancer. Oncology. 2012;82:25–9.

    CAS  PubMed  Google Scholar 

  34. Willey CD, Xiao D, Tu T, Kim KW, Moretti L, Niermann KJ, et al. Enzastaurin (LY317615), a protein kinase C beta selective inhibitor, enhances antiangiogenic effect of radiation. Int J Radiat Oncol* Biol* Phys. 2010;77:1518–26.

    CAS  Google Scholar 

  35. Caino MC, Lopez-Haber C, Kim J, Mochly-Rosen D, Kazanietz MG. Proteins kinase Cϵ is required for non-small cell lung carcinoma growth and regulates the expression of apoptotic genes. Oncogene. 2012;31:2593–2600.

    CAS  PubMed  Google Scholar 

  36. Hill K, Erdogan E, Khoor A, Walsh M, Leitges M, Murray NR, et al. Protein kinase Cα suppresses Kras-mediated lung tumor formation through activation of a p38 MAPK-TGFβ signaling axis. Oncogene. 2014;33:2134–44.

    CAS  PubMed  Google Scholar 

  37. Antal CE, Hudson AM, Kang E, Zanca C, Wirth C, Stephenson NL, et al. Cancer-associated protein kinase C mutations reveal kinase’s role as tumor suppressor. Cell. 2015;160:489–502.

    CAS  PubMed  PubMed Central  Google Scholar 

  38. Alessi DR, James SR, Downes CP, Holmes AB, Gaffney PR, Reese CB, et al. Characterization of a 3-phosphoinositide-dependent protein kinase which phosphorylates and activates protein kinase Bα. Curr Biol. 1997;7:261–9.

    CAS  PubMed  Google Scholar 

  39. Vanhaesebroeck B, Alessi DR. The PI3K–PDK1 connection: more than just a road to PKB. Biochem J. 2000;346:561–76.

    CAS  PubMed  PubMed Central  Google Scholar 

  40. Sarbassov DD, Guertin DA, Ali SM, Sabatini DM. Phosphorylation and regulation of Akt/PKB by the rictor-mTOR complex. Science. 2005;307:1098–101.

    CAS  PubMed  Google Scholar 

  41. Hanada M, Feng J, Hemmings BA. Structure, regulation and function of PKB/AKT—a major therapeutic target. Biochim Biophys Acta (BBA)-Proteins Proteom. 2004;1697:3–16.

    CAS  Google Scholar 

  42. Tsurutani J, Fukuoka J, Tsurutani H, Shih JH, Hewitt SM, Travis WD, et al. Evaluation of two phosphorylation sites improves the prognostic significance of Akt activation in non-small-cell lung cancer tumors. J Clin Oncol. 2006;24:306–14.

    CAS  PubMed  Google Scholar 

  43. Gallay N, Dos Santos C, Cuzin L, Bousquet M, Gouy VS, Chaussade C, et al. The level of AKT phosphorylation on threonine 308 but not on serine 473 is associated with high-risk cytogenetics and predicts poor overall survival in acute myeloid leukaemia. Leukemia. 2009;23:1029–38.

    CAS  PubMed  Google Scholar 

  44. Rodrigues GA, Falasca M, Zhang Z, Ong SH, Schlessinger J. A novel positive feedback loop mediated by the docking protein Gab1 and phosphatidylinositol 3-kinase in epidermal growth factor receptor signaling. Mol Cell Biol. 2000;20:1448–59.

    CAS  PubMed  PubMed Central  Google Scholar 

  45. Mattoon DR, Lamothe B, Lax I, Schlessinger J. The docking protein Gab1 is the primary mediator of EGF-stimulated activation of the PI-3K/Akt cell survival pathway. BMC Biol. 2004;2:24.

    PubMed  PubMed Central  Google Scholar 

  46. Song X, Fan P-D, Bantikassegn A, Guha U, Threadgill DW, Varmus H, et al. ERBB3-independent activation of the PI3K pathway in EGFR-mutant lung adenocarcinomas. Cancer Res. 2015;75:1035–45.

    CAS  PubMed  PubMed Central  Google Scholar 

  47. Feng X, Hannun YA. An essential role for autophosphorylation in the dissociation of activated protein kinase C from the plasma membrane. J Biol Chem. 1998;273:26870–4.

    CAS  PubMed  Google Scholar 

  48. Idkowiak-Baldys J, Becker KP, Kitatani K, Hannun YA. Dynamic sequestration of the recycling compartment by classical protein kinase C. J Biol Chem. 2006;281:22321–31.

    CAS  PubMed  Google Scholar 

  49. Liu M, Idkowiak-Baldys J, Roddy PL, Baldys A, Raymond J, Clarke CJ, et al. Sustained activation of protein kinase C induces delayed phosphorylation and regulates the fate of epidermal growth factor receptor. PLoS ONE. 2013;8:e80721.

    CAS  PubMed  PubMed Central  Google Scholar 

  50. Sanjana NE, Shalem O, Zhang F. Improved vectors and genome-wide libraries for CRISPR screening. Nat Methods. 2014;11:783.

    CAS  PubMed  PubMed Central  Google Scholar 

  51. Pulkoski-Gross MJ, Jenkins ML, Truman J-P, Salama MF, Clarke CJ, Burke JE, et al. An intrinsic lipid-binding interface controls sphingosine kinase 1 function. J Lipid Res. 2018;59:462–74.

    CAS  PubMed  PubMed Central  Google Scholar 

  52. Idkowiak-Baldys J, Baldys A, Raymond JR, Hannun YA. Sustained receptor stimulation leads to sequestration of recycling endosomes in a classical protein kinase C- and phospholipase D-dependent manner. J Biol Chem. 2009;284:22322–31.

    CAS  PubMed  PubMed Central  Google Scholar 

Download references

Acknowledgements

This work was supported by NCI grant CA97132.

Author information

Authors and Affiliations

  1. Department of Medicine, Stony Brook University, Stony Brook, NY, 11794, USA

    Mohamed F. Salama, Mengling Liu, Christopher J. Clarke, Mel Pilar Espaillat, Lina M. Obeid & Yusuf A. Hannun

  2. Stony Brook Cancer Center, Stony Brook University Hospital, Stony Brook, NY, 11794, USA

    Mohamed F. Salama, Mengling Liu, Christopher J. Clarke, Mel Pilar Espaillat, Lina M. Obeid & Yusuf A. Hannun

  3. Department of Biochemistry, Faculty of Veterinary Medicine, Mansoura University, Mansoura, Egypt

    Mohamed F. Salama

  4. Department of Immuno-Oncology, HD Biosciences Inc, San Diego, CA, 92121, USA

    Mengling Liu

  5. Departments of Biochemistry and Pathology, Stony Brook University, Stony Brook, NY, 11794, USA

    John D. Haley & Yusuf A. Hannun

  6. Department of Biomedical Informatics, Stony Brook University, Stony Brook, NY, 11794, USA

    Ting Jin & Daifeng Wang

  7. Department of Biostatistics and Medical Informatics and Waisman Center, University of Wisconsin-Madison, Madison, WI, 53792, USA

    Daifeng Wang

Authors
  1. Mohamed F. Salama
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Mengling Liu
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Christopher J. Clarke
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Mel Pilar Espaillat
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. John D. Haley
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Ting Jin
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Daifeng Wang
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. Lina M. Obeid
    View author publications

    You can also search for this author in PubMed Google Scholar

  9. Yusuf A. Hannun
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Yusuf A. Hannun.

Ethics declarations

Conflict of interest

The authors declare that they have no conflict of interest.

Additional information

Publisher’s note: Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Supplementary information

Rights and permissions

Reprints and Permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Salama, M.F., Liu, M., Clarke, C.J. et al. PKCα is required for Akt-mTORC1 activation in non-small cell lung carcinoma (NSCLC) with EGFR mutation. Oncogene 38, 7311–7328 (2019). https://doi.org/10.1038/s41388-019-0950-z

Download citation

  • Received:

  • Revised:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/s41388-019-0950-z

Search

Advanced search

Quick links