Cancer cells are in continuous communication with the surrounding microenvironment and this communication can affect tumor evolution. In this work, we show that phospholipase D2 (PLD2) was overexpressed in colon tumors and is secreted by cancer cells, inducing senescence in neighboring fibroblasts. This occurs through its lipase domain. Senescence induced by its product, phosphatidic acid, leads to a senescence-associated secretory phenotype (SASP) able to increase the stem properties of cancer cells. This increase in stemness occurs by Wnt pathway activacion. This closes a feedback loop in which senescence acts as a crosspoint for the generation of CSCs mediated by phospholipid metabolism. We also demonstrate the connexion of both phenomena in mouse models in vivo showing that a high PLD2 expression increased stemness and tumorigenesis. Thus, the patients with colon cancer show high levels of PLD2 and SASP factor genes expression correlating with Wnt pathway activation. Therefore, we demonstrate that tumor cell-secreted PLD2 contributes to tumor development by modifying the microenvironment, making it a possible therapeutic target for cancer treatment. This mechanism may also explain the high levels of Wnt pathway activation in colon cancer.
Subscribe to Journal
Get full journal access for 1 year
only $2.38 per issue
All prices are NET prices.
VAT will be added later in the checkout.
Tax calculation will be finalised during checkout.
Rent or Buy article
Get time limited or full article access on ReadCube.
All prices are NET prices.
Joyce JA, Pollard JW. Microenvironmental regulation of metastasis. Nat Rev Cancer. 2009;9:239–52.
Valent P, Bonnet D, De Maria R, Lapidot T, Copland M, Melo JV, et al. Cancer stem cell definitions and terminology: the devil is in the details. Nat Rev Cancer. 2012;12:767–75.
Vermeulen L, De Sousa EMF, van der Heijden M, Cameron K, de Jong JH, Borovski T, et al. Wnt activity defines colon cancer stem cells and is regulated by the microenvironment. Nat Cell Biol. 2010;12:468–76.
Hanahan D, Weinberg RA. Hallmarks of cancer: the next generation. Cell. 2011;144:646–74.
Serrano M, Lin AW, McCurrach ME, Beach D, Lowe SW. Oncogenic ras provokes premature cell senescence associated with accumulation of p53 and p16INK4a. Cell. 1997;88:593–602.
Zhu J, Woods D, McMahon M, Bishop JM. Senescence of human fibroblasts induced by oncogenic Raf. Genes Dev. 1998;12:2997–3007.
Braig M, Lee S, Loddenkemper C, Rudolph C, Peters AH, Schlegelberger B, et al. Oncogene-induced senescence as an initial barrier in lymphoma development. Nature. 2005;436:660–5.
Chen Z, Trotman LC, Shaffer D, Lin HK, Dotan ZA, Niki M, et al. Crucial role of p53-dependent cellular senescence in suppression of Pten-deficient tumorigenesis. Nature. 2005;436:725–30.
Collado M, Gil J, Efeyan A, Guerra C, Schuhmacher AJ, Barradas M, et al. Tumour biology: senescence in premalignant tumours. Nature. 2005;436:642.
Michaloglou C, Vredeveld LC, Soengas MS, Denoyelle C, Kuilman T, van der Horst CM, et al. BRAFE600-associated senescence-like cell cycle arrest of human naevi. Nature. 2005;436:720–4.
Burd CE, Sorrentino JA, Clark KS, Darr DB, Krishnamurthy J, Deal AM, et al. Monitoring tumorigenesis and senescence in vivo with ap16(INK4a)-luciferase model. Cell. 2013;152:340–51.
Coppe JP, Desprez PY, Krtolica A, Campisi J. The senescence-associated secretory phenotype: the dark side of tumor suppression. Annu Rev Pathol. 2010;5:99–118.
Acosta JC, O’Loghlen A, Banito A, Guijarro MV, Augert A, Raguz S, et al. Chemokine signaling via the CXCR2 receptor reinforces senescence. Cell. 2008;133:1006–18.
Coppe JP, Patil CK, Rodier F, Sun Y, Munoz DP, Goldstein J, et al. Senescence-associated secretory phenotypes reveal cell-nonautonomous functions of oncogenic RAS and the p53 tumor suppressor. PLoS Biol. 2008;6:2853–68.
Herranz N, Gallage S, Mellone M, Wuestefeld T, Klotz S, Hanley CJ, et al. mTOR regulates MAPKAPK2 translation to control the senescence-associated secretory phenotype. Nat Cell Biol. 2015;17:1205–17.
Laberge RM, Sun Y, Orjalo AV, Patil CK, Freund A, Zhou L, et al. MTOR regulates the pro-tumorigenic senescence-associated secretory phenotype by promoting IL1A translation. Nat Cell Biol. 2015;17:1049–61.
Frohman MA, Sung TC, Morris AJ. Mammalian phospholipase D structure and regulation. Biochim Biophys Acta. 1999;1439:175–86.
Gomez-Cambronero J. Phospholipase D in cell signaling: from a myriad of cell functions to cancer growth and metastasis. J Biol Chem. 2014;289:22557–66.
Hammond SM, Jenco JM, Nakashima S, Cadwallader K, Gu Q, Cook S, et al. Characterization of two alternately spliced forms of phospholipase D1. Activation of the purified enzymes by phosphatidylinositol 4,5-bisphosphate, ADP-ribosylation factor, and Rho family monomeric GTP-binding proteins and protein kinase C-alpha. J Biol Chem. 1997;272:3860–8.
Lopez I, Arnold RS, Lambeth JD. Cloning and initial characterization of a human phospholipase D2 (hPLD2). ADP-ribosylation factor regulates hPLD2. J Biol Chem. 1998;273:12846–52.
Xu L, Frankel P, Jackson D, Rotunda T, Boshans RL, D’Souza-Schorey C, et al. Elevated phospholipase D activity in H-Ras- but not K-Ras-transformed cells by the synergistic action of RalA and ARF6. Mol Cell Biol. 2003;23:645–54.
Zheng Y, Rodrik V, Toschi A, Shi M, Hui L, Shen Y, et al. Phospholipase D couples survival and migration signals in stress response of human cancer cells. J Biol Chem. 2006;281:15862–8.
Foster DA, Xu L. Phospholipase D in cell proliferation and cancer. Mol Cancer Res. 2003;1:789–800.
Henkels KM, Boivin GP, Dudley ES, Berberich SJ, Gomez-Cambronero J. Phospholipase D (PLD) drives cell invasion, tumor growth and metastasis in a human breast cancer xenograph model. Oncogene. 2013;32:5551–62.
Song JG, Pfeffer LM, Foster DA. v-Src increases diacylglycerol levels via a type D phospholipase-mediated hydrolysis of phosphatidylcholine. Mol Cell Biol. 1991;11:4903–8.
Carnero A, Cuadrado A, del Peso L, Lacal JC. Activation of type D phospholipase by serum stimulation and ras-induced transformation in NIH3T3 cells. Oncogene. 1994;9:1387–95.
Carnero A, Dolfi F, Lacal JC. ras-p21 activates phospholipase D and A2, but not phospholipase C or PKC, in Xenopus laevis oocytes. J Cell Biochem. 1994;54:478–86.
Carnero A, Lacal JC. Activation of intracellular kinases in Xenopus oocytes by p21ras and phospholipases: a comparative study. Mol Cell Biol. 1995;15:1094–101.
Frankel P, Ramos M, Flom J, Bychenok S, Joseph T, Kerkhoff E, et al. Ral and Rho-dependent activation of phospholipase D in v-Raf-transformed cells. Biochem Biophys Res Commun. 1999;255:502–7.
Oshimoto H, Okamura S, Yoshida M, Mori M. Increased activity and expression of phospholipase D2 in human colorectal cancer. Oncol Res. 2003;14:31–37.
Saito M, Iwadate M, Higashimoto M, Ono K, Takebayashi Y, Takenoshita S. Expression of phospholipase D2 in human colorectal carcinoma. Oncol Rep. 2007;18:1329–34.
Jeon H, Kwak D, Noh J, Lee MN, Lee CS, Suh PG, et al. Phospholipase D2 induces stress fiber formation through mediating nucleotide exchange for RhoA. Cell Signal. 2011;23:1320–6.
Mahankali M, Peng HJ, Henkels KM, Dinauer MC, Gomez-Cambronero J. Phospholipase D2 (PLD2) is a guanine nucleotide exchange factor (GEF) for the GTPase Rac2. Proc Natl Acad Sci USA. 2011;108:19617–22.
Bach JP, Rinn B, Meyer B, Dodel R, Bacher M. Role of MIF in inflammation and tumorigenesis. Oncology. 2008;75:127–33.
Nakamura T, Matsumoto K, Kiritoshi A, Tano Y, Nakamura T. Induction of hepatocyte growth factor in fibroblasts by tumor-derived factors affects invasive growth of tumor cells: in vitro analysis of tumor-stromal interactions. Cancer Res. 1997;57:3305–13.
Soria G, Ben-Baruch A. The inflammatory chemokines CCL2 and CCL5 in breast cancer. Cancer Lett. 2008;267:271–85.
Tsuyada A, Chow A, Wu J, Somlo G, Chu P, Loera S, et al. CCL2 mediates cross-talk between cancer cells and stromal fibroblasts that regulates breast cancer stem cells. Cancer Res. 2012;72:2768–79.
Barrandon Y, Green H. Three clonal types of keratinocyte with different capacities for multiplication. Proc Natl Acad Sci USA. 1987;84:2302–6.
O’Brien CA, Pollett A, Gallinger S, Dick JE. A human colon cancer cell capable of initiating tumour growth in immunodeficient mice. Nature. 2007;445:106–10.
Ricci-Vitiani L, Lombardi DG, Pilozzi E, Biffoni M, Todaro M, Peschle C, et al. Identification and expansion of human colon-cancer-initiating cells. Nature. 2007;445:111–5.
Park JB, Lee CS, Jang JH, Ghim J, Kim YJ, You S, et al. Phospholipase signalling networks in cancer. Nat Rev Cancer. 2012;12:782–92.
Venable ME, Blobe GC, Obeid LM. Identification of a defect in the phospholipase D/diacylglycerol pathway in cellular senescence. J Biol Chem. 1994;269:26040–4.
Lee YH, Bae YS. Phospholipase D2 downregulation induces cellular senescence through a reactive oxygen species-p53-p21Cip1/WAF1 pathway. FEBS Lett. 2014;588:3251–8.
Perez-Mancera PA, Young AR, Narita M. Inside and out: the activities of senescence in cancer. Nat Rev Cancer. 2014;14:547–58.
Schwitalla S, Fingerle AA, Cammareri P, Nebelsiek T, Goktuna SI, Ziegler PK, et al. Intestinal tumorigenesis initiated by dedifferentiation and acquisition of stem-cell-like properties. Cell. 2013;152:25–38.
Gidekel Friedlander SY, Chu GC, Snyder EL, Girnius N, Dibelius G, Crowley D, et al. Context-dependent transformation of adult pancreatic cells by oncogenic K-Ras. Cancer Cell. 2009;16:379–89.
Knoepp SM, Chahal MS, Xie Y, Zhang Z, Brauner DJ, Hallman MA, et al. Effects of active and inactive phospholipase D2 on signal transduction, adhesion, migration, invasion, and metastasis in EL4 lymphoma cells. Mol Pharmacol. 2008;74:574–84.
Kang DW, Lee SH, Yoon JW, Park WS, Choi KY, Min do S. Phospholipase D1 drives a positive feedback loop to reinforce the Wnt/beta-catenin/TCF signaling axis. Cancer Res. 2010;70:4233–42.
Kang DW, Min do S. Positive feedback regulation between phospholipase D and Wnt signaling promotes Wnt-driven anchorage-independent growth of colorectal cancer cells. PLoS ONE. 2010;5:e12109.
Ferrer I, Verdugo-Sivianes EM, Castilla MA, Melendez R, Marin JJ, Munoz-Galvan S et al. Loss of the tumor suppressor spinophilin (PPP1R9B) increases the cancer stem cell population in breast tumors. Oncogene. 2015;35:2777–88.
Henkels KM, Farkaly T, Mahankali M, Segall JE, Gomez-Cambronero J. Cell invasion of highly metastatic MTLn3 cancer cells is dependent on phospholipase D2 (PLD2) and Janus kinase 3 (JAK3). J Mol Biol. 2011;408:850–62.
Mahankali M, Henkels KM, Alter G, Gomez-Cambronero J. Identification of the catalytic site of phospholipase D2 (PLD2) newly described guanine nucleotide exchange factor activity. J Biol Chem. 2012;287:41417–31.
Perez M, Lucena-Cacace A, Marin-Gomez LM, Padillo-Ruiz J, Robles-Frias MJ, Saez C, et al. Dasatinib, a Src inhibitor, sensitizes liver metastatic colorectal carcinoma to oxaliplatin in tumors with high levels of phospho-Src. Oncotarget. 2016;7:33111–24.
We thank the donors, the HUVR-IBiS Biobank (Andalusian Public Health System Biobank and ISCIII-Red de Biobancos PT13/0010/0056) for the human specimens used in this study, Carolina Castilla for the assistance with IHC assays and Carmen Saez for histological analyses. SM-G was supported by a Sara Borrell grant from ISCIII (CD16/00230). AC lab was supported by grants from the Spanish Ministry of Economy and Competitivity, Plan Estatal de I + D + I 2013–2016, ISCIII (Fis: PI15/00045), and CIBER de Cáncer (CD16/12/00275), co-funded by FEDER from Regional Development European Funds (European Union), Consejeria de Ciencia e Innovacion (CTS-1848), and Consejeria de Salud of the Junta de Andalucia (PI-0397–2017). This work was also supported by Fundacion BBVA. Especial thanks to the AECC Foundation for supporting this work. JG-C is supported by Grant HL056653–14 from the NIH.
Conflict of interest
The authors declare that they have no conflict of interest.
About this article
Cite this article
Muñoz-Galván, S., Lucena-Cacace, A., Perez, M. et al. Tumor cell-secreted PLD increases tumor stemness by senescence-mediated communication with microenvironment. Oncogene 38, 1309–1323 (2019). https://doi.org/10.1038/s41388-018-0527-2