The TSC-mTOR pathway regulates macrophage polarization

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Abstract

Macrophages are able to polarize to proinflammatory M1 or alternative M2 states with distinct phenotypes and physiological functions. How metabolic status regulates macrophage polarization remains not well understood, and here we examine the role of mTOR (mechanistic target of rapamycin), a central metabolic pathway that couples nutrient sensing to regulation of metabolic processes. Using a mouse model in which myeloid lineage-specific deletion of Tsc1 (Tsc1Δ/Δ) leads to constitutive mTOR complex 1 (mTORC1) activation, we find that Tsc1Δ/Δ macrophages are refractory to IL-4-induced M2 polarization, but produce increased inflammatory responses to proinflammatory stimuli. Moreover, mTORC1-mediated downregulation of Akt signalling critically contributes to defective polarization. These findings highlight a key role for the mTOR pathway in regulating macrophage polarization, and suggest how nutrient sensing and metabolic status could be ‘hard-wired’ to control of macrophage function, with broad implications for regulation of type 2 immunity, inflammation and allergy.

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Figure 1: Tsc1Δ/Δ BMDMs have defective M2 polarization and enhanced responses to LPS stimulation.
Figure 2: STAT6 and PPARγ activity are normal in Tsc1Δ/Δ BMDMs.
Figure 3: Constitutive mTORC1 activity attenuates IL-4-induced Akt activation.
Figure 4: Akt signalling is critical for polarization in Tsc1Δ/Δ BMDMs.
Figure 5: M2 polarization in Tsc1Δ/Δ mice is impaired in vivo.
Figure 6: Proposed model for how mTORC1 activity controls macrophage polarization.

References

  1. 1

    Sica, A. & Mantovani, A. Macrophage plasticity and polarization: in vivo veritas. J. Clin. Invest. 122, 787–795 (2012).

  2. 2

    Gordon, S. & Martinez, F. O. Alternative activation of macrophages: mechanism and functions. Immunity 32, 593–604 (2010).

  3. 3

    Chawla, A. Control of macrophage activation and function by PPARs. Circ. Res. 106, 1559–1569 (2010).

  4. 4

    Odegaard, J. I. et al. Macrophage-specific PPARgamma controls alternative activation and improves insulin resistance. Nature 447, 1116–1120 (2007).

  5. 5

    Kang, K. et al. Adipocyte-derived Th2 cytokines and myeloid PPARdelta regulate macrophage polarization and insulin sensitivity. Cell Metab. 7, 485–495 (2008).

  6. 6

    Van den Bossche, J. et al. Pivotal advance: arginase-1-independent polyamine production stimulates the expression of IL-4-induced alternatively activated macrophage markers while inhibiting LPS-induced expression of inflammatory genes. J. Leukoc. Biol. 91, 685–699 (2012).

  7. 7

    Murray, P. J. & Wynn, T. A. Protective and pathogenic functions of macrophage subsets. Nat. Rev. Immunol. 11, 723–737 (2011).

  8. 8

    Cramer, T. et al. HIF-1alpha is essential for myeloid cell-mediated inflammation. Cell 112, 645–657 (2003).

  9. 9

    Vats, D. et al. Oxidative metabolism and PGC-1beta attenuate macrophage-mediated inflammation. Cell Metab. 4, 13–24 (2006).

  10. 10

    Howell, J. J. & Manning, B. D. mTOR couples cellular nutrient sensing to organismal metabolic homeostasis. Trends Endocrinol. Metab. 22, 94–102 (2011).

  11. 11

    Duvel, K. et al. Activation of a metabolic gene regulatory network downstream of mTOR complex 1. Mol. Cell 39, 171–183 (2010).

  12. 12

    Tee, A. R., Fingar, D. C., Manning, B. D., Kwiatkowski, D. J., Cantley, L. C. & Blenis, J. Tuberous sclerosis complex-1 and -2 gene products function together to inhibit mammalian target of rapamycin (mTOR)-mediated downstream signaling. Proc. Natl Acad. Sci. USA 99, 13571–13576 (2002).

  13. 13

    Huang, J. & Manning, B. D. A complex interplay between Akt, TSC2 and the two mTOR complexes. Biochem. Soc. Trans 37, 217–222 (2009).

  14. 14

    Chi, H. Regulation and function of mTOR signalling in T cell fate decisions. Nat. Rev. Immunol. 12, 325–338 (2012).

  15. 15

    Sengupta, S., Peterson, T. R. & Sabatini, D. M. Regulation of the mTOR complex 1 pathway by nutrients, growth factors, and stress. Mol. Cell 40, 310–322 (2010).

  16. 16

    Chong-Kopera, H. et al. TSC1 stabilizes TSC2 by inhibiting the interaction between TSC2 and the HERC1 ubiquitin ligase. J. Biol. Chem. 281, 8313–8316 (2006).

  17. 17

    Pan, H., O'Brien, T. F., Zhang, P. & Zhong, X. P. The role of tuberous sclerosis complex 1 in regulating innate immunity. J. Immunol. 188, 3658–3666 (2012).

  18. 18

    Fingar, D. C., Salama, S., Tsou, C., Harlow, E. & Blenis, J. Mammalian cell size is controlled by mTOR and its downstream targets S6K1 and 4EBP1/eIF4E. Genes Dev. 16, 1472–1487 (2002).

  19. 19

    Mantovani, A., Sica, A., Sozzani, S., Allavena, P., Vecchi, A. & Locati, M. The chemokine system in diverse forms of macrophage activation and polarization. Trends Immunol. 25, 677–686 (2004).

  20. 20

    Filardy, A. A. et al. Proinflammatory clearance of apoptotic neutrophils induces an IL-12(low)IL-10(high) regulatory phenotype in macrophages. J. Immunol. 185, 2044–2050 (2010).

  21. 21

    Martinez, F. O., Sica, A., Mantovani, A. & Locati, M. Macrophage activation and polarization. Front. Biosci. 13, 453–461 (2008).

  22. 22

    Szanto, A. et al. STAT6 transcription factor is a facilitator of the nuclear receptor PPARgamma-regulated gene expression in macrophages and dendritic cells. Immunity 33, 699–712 (2010).

  23. 23

    Acacia de Sa Pinheiro, A. et al. IL-4 induces a wide-spectrum intracellular signaling cascade in CD8+ T cells. J. Leukoc. Biol. 81, 1102–1110 (2007).

  24. 24

    Wang, I. M., Lin, H., Goldman, S. J. & Kobayashi, M. STAT-1 is activated by IL-4 and IL-13 in multiple cell types. Mol. Immunol. 41, 873–884 (2004).

  25. 25

    Bhattacharjee, A. et al. IL-4 and IL-13 employ discrete signaling pathways for target gene expression in alternatively activated monocytes/macrophages. Free Radic. Biol. Med. 54, 1–16 (2013).

  26. 26

    Tontonoz, P. et al. Adipocyte-specific transcription factor ARF6 is a heterodimeric complex of two nuclear hormone receptors, PPAR gamma and RXR alpha. Nucleic. Acids. Res. 22, 5628–5634 (1994).

  27. 27

    Tontonoz, P., Nagy, L., Alvarez, J. G., Thomazy, V. A. & Evans, R. M. PPARgamma promotes monocyte/macrophage differentiation and uptake of oxidized LDL. Cell 93, 241–252 (1998).

  28. 28

    Reilly, S. M. & Lee, C. H. PPAR delta as a therapeutic target in metabolic disease. FEBS Lett. 582, 26–31 (2008).

  29. 29

    Wurster, A. L., Withers, D. J., Uchida, T., White, M. F. & Grusby, M. J. Stat6 and IRS-2 cooperate in interleukin 4 (IL-4)-induced proliferation and differentiation but are dispensable for IL-4-dependent rescue from apoptosis. Mol. Cell Biol. 22, 117–126 (2002).

  30. 30

    Heller, N. M. et al. Type I IL-4Rs selectively activate IRS-2 to induce target gene expression in macrophages. Sci. Signal 1, ra17 (2008).

  31. 31

    Manning, B. D. & Cantley, L. C. AKT/PKB signaling: navigating downstream. Cell 129, 1261–1274 (2007).

  32. 32

    Fang, X. et al. Convergence of multiple signaling cascades at glycogen synthase kinase 3: edg receptor-mediated phosphorylation and inactivation by lysophosphatidic acid through a protein kinase C-dependent intracellular pathway. Mol. Cell Biol. 22, 2099–2110 (2002).

  33. 33

    Cross, D. A., Alessi, D. R., Cohen, P., Andjelkovich, M. & Hemmings, B. A. Inhibition of glycogen synthase kinase-3 by insulin mediated by protein kinase B. Nature 378, 785–789 (1995).

  34. 34

    Eldar-Finkelman, H., Seger, R., Vandenheede, J. R. & Krebs, E. G. Inactivation of glycogen synthase kinase-3 by epidermal growth factor is mediated by mitogen-activated protein kinase/p90 ribosomal protein S6 kinase signaling pathway in NIH/3T3 cells. J. Biol. Chem. 270, 987–990 (1995).

  35. 35

    Zhang, H. H., Lipovsky, A. I., Dibble, C. C., Sahin, M. & Manning, B. D. S6K1 regulates GSK3 under conditions of mTOR-dependent feedback inhibition of Akt. Mol. Cell 24, 185–197 (2006).

  36. 36

    Harrington, L. S. et al. The TSC1-2 tumor suppressor controls insulin-PI3K signaling via regulation of IRS proteins. J. Cell Biol. 166, 213–223 (2004).

  37. 37

    Shah, O. J., Wang, Z. & Hunter, T. Inappropriate activation of the TSC/Rheb/mTOR/S6K cassette induces IRS1/2 depletion, insulin resistance, and cell survival deficiencies. Curr. Biol. 14, 1650–1656 (2004).

  38. 38

    Yecies, J. L. et al. Akt stimulates hepatic SREBP1c and lipogenesis through parallel mTORC1-dependent and independent pathways. Cell Metab. 14, 21–32 (2011).

  39. 39

    Fritsche, L. et al. Insulin-induced serine phosphorylation of IRS-2 via ERK1/2 and mTOR: studies on the function of Ser675 and Ser907. Am. J. Physiol. Endocrinol. Metab. 300, E824–E836 (2011).

  40. 40

    Harrington, L. S., Findlay, G. M. & Lamb, R. F. Restraining PI3K: mTOR signalling goes back to the membrane. Trends. Biochem. Sci. 30, 35–42 (2005).

  41. 41

    Yu, Y. et al. Phosphoproteomic analysis identifies Grb10 as an mTORC1 substrate that negatively regulates insulin signaling. Science 332, 1322–1326 (2011).

  42. 42

    Hsu, P. P. et al. The mTOR-regulated phosphoproteome reveals a mechanism of mTORC1-mediated inhibition of growth factor signaling. Science 332, 1317–1322 (2011).

  43. 43

    Holt, L. J. & Siddle, K. Grb10 and Grb14: enigmatic regulators of insulin action--and more? Biochem. J. 388, 393–406 (2005).

  44. 44

    Wick, K. R. et al. Grb10 inhibits insulin-stimulated insulin receptor substrate (IRS)-phosphatidylinositol 3-kinase/Akt signaling pathway by disrupting the association of IRS-1/IRS-2 with the insulin receptor. J. Biol. Chem. 278, 8460–8467 (2003).

  45. 45

    Vecchione, A., Marchese, A., Henry, P., Rotin, D. & Morrione, A. The Grb10/Nedd4 complex regulates ligand-induced ubiquitination and stability of the insulin-like growth factor I receptor. Mol. Cell Biol. 23, 3363–3372 (2003).

  46. 46

    Mora, A., Komander, D., van Aalten, D. M. & Alessi, D. R. PDK1, the master regulator of AGC kinase signal transduction. Semin. Cell Dev. Biol. 15, 161–170 (2004).

  47. 47

    Jenkins, S. J. et al. Local macrophage proliferation, rather than recruitment from the blood, is a signature of TH2 inflammation. Science 332, 1284–1288 (2011).

  48. 48

    Satoh, T. et al. The Jmjd3-Irf4 axis regulates M2 macrophage polarization and host responses against helminth infection. Nat. Immunol. 11, 936–944 (2010).

  49. 49

    Reese, T. A. et al. Chitin induces accumulation in tissue of innate immune cells associated with allergy. Nature 447, 92–96 (2007).

  50. 50

    Weichhart, T. et al. The TSC-mTOR signaling pathway regulates the innate inflammatory response. Immunity 29, 565–577 (2008).

  51. 51

    Chen, W. et al. Macrophage-induced tumor angiogenesis is regulated by the TSC2-mTOR pathway. Cancer Res. 72, 1363–1372 (2012).

  52. 52

    O'Neill, L. A. & Hardie, D. G. Metabolism of inflammation limited by AMPK and pseudo-starvation. Nature 493, 346–355 (2013).

  53. 53

    Biswas, S. K. & Mantovani, A. Macrophage plasticity and interaction with lymphocyte subsets: cancer as a paradigm. Nat. Immunol. 11, 889–896 (2010).

  54. 54

    Wills-Karp, M. & Finkelman, F. D. Untangling the complex web of IL-4- and IL-13-mediated signaling pathways. Sci. Signal. 1, pe55 (2008).

  55. 55

    Song, M. S., Salmena, L. & Pandolfi, P. P. The functions and regulation of the PTEN tumour suppressor. Nat. Rev. Mol. Cell Biol. 13, 283–296 (2012).

  56. 56

    Luyendyk, J. P. et al. Genetic analysis of the role of the PI3K-Akt pathway in lipopolysaccharide-induced cytokine and tissue factor gene expression in monocytes/macrophages. J. Immunol. 180, 4218–4226 (2008).

  57. 57

    Fan, W. et al. FoxO1 regulates Tlr4 inflammatory pathway signalling in macrophages. EMBO J. 29, 4223–4236 (2010).

  58. 58

    Arranz, A. et al. Akt1 and Akt2 protein kinases differentially contribute to macrophage polarization. Proc. Natl. Acad. Sci. USA 109, 9517–9522 (2012).

  59. 59

    Guertin, D. A. et al. Ablation in mice of the mTORC components raptor, rictor, or mLST8 reveals that mTORC2 is required for signaling to Akt-FOXO and PKCalpha, but not S6K1. Dev. Cell 11, 859–871 (2006).

  60. 60

    Jacinto, E. et al. SIN1/MIP1 maintains rictor-mTOR complex integrity and regulates Akt phosphorylation and substrate specificity. Cell 127, 125–137 (2006).

  61. 61

    Laplante, M. & Sabatini, D. M. mTOR signaling in growth control and disease. Cell 149, 274–293 (2012).

  62. 62

    Chawla, A., Nguyen, K. D. & Goh, Y. P. Macrophage-mediated inflammation in metabolic disease. Nat. Rev. Immunol. 11, 738–749 (2011).

  63. 63

    Clausen, B. E. Conditional gene targeting in macrophages and granulocytes using LysMcre mice. Transgenic Res. 8, 265–277 (1999).

  64. 64

    Kwiatkowski, D. J. et al. A mouse model of TSC1 reveals sex-dependent lethality from liver hemangiomas, and up-regulation of p70S6 kinase activity in Tsc1 null cells. Hum. Mol. Genet. 11, 525–534 (2002).

  65. 65

    Lamming, D. W. et al. Rapamycin-induced insulin resistance is mediated by mTORC2 loss and uncoupled from longevity. Science 335, 1638–1643 (2012).

  66. 66

    Sinha, P., Clements, V. K. & Ostrand-Rosenberg, S. Reduction of myeloid-derived suppressor cells and induction of M1 macrophages facilitate the rejection of established metastatic disease. J. Immunol. 174, 636–645 (2005).

  67. 67

    Liu, S. et al. Role of peroxisome proliferator-activated receptor {delta}/{beta} in hepatic metabolic regulation. J. Biol. Chem. 286, 1237–1247 (2011).

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Acknowledgements

This project was supported by a NIH grant R01AI102964 (to T.H.). A.J.C. is a recipient of a Ford Foundation Predoctoral Fellowship. B.D.M. was supported by a NIH grant R01-CA122617 and I.B.-S. by a LAM Foundation Fellowship. D.M.S. was funded in part by a Julie Martin Mid-Career Award in Aging Research from the American Federation of Aging Research (AFAR) and is an Investigator of the Howard Hughes Medical Institute. D.W.L. is supported by a K99/R00 award from the NIH/NIA (1K99AG041765-01A1). We also thank C.H. Lee for critical reading of the manuscript and S.H. Liu for technical advice.

Author information

V.B. and A.J.C. designed and performed the experiments, analysed the data and wrote the paper. T.H. supervised the project, including experimental design and data analysis, and edited the paper. I.B.-S. and D.W.L. contributed technical expertise. D.M.S. and B.D.M. provided reagents and mice.

Correspondence to Tiffany Horng.

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The authors declare no competing financial interests.

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