Abstract
Tumor suppressor complex TSC1/TSC2 represents a key negative regulator of mammalian target of rapamycin (mTOR)-S6 kinase 1 signaling. Mutational inactivation of TSC1 or TSC2, linked to a rare lung disease, lymphangioleiomyomatosis (LAM), manifests as neoplastic growth of smooth-muscle (SM)-like cells and cystic destruction of the lungs that induces loss of pulmonary function. However, the precise mechanisms of abnormal cell growth in LAM remain uncertain. Here, we demonstrate increased signal transducer and activator of transcription (STAT) 3 expression, phosphorylation, and nuclear localization in SM-like cells in LAM lungs and in TSC2-null xenographic tumors. Treatment of TSC2-null tumors with mTOR inhibitor rapamycin attenuated STAT3 expression and phosphorylation. Increased STAT3 level and activation were also observed in LAM-dissociated (LAMD) cell cultures compared with normal human bronchus fibroblasts (HBFs) from LAM patients. Although interferon (IFN)-γ inhibited proliferation of HBFs, IFN-γ treatment had little effect on proliferation of LAMD and TSC2-null cells. Re-expression of TSC2 or treatment with rapamycin inhibited IFN-γ-induced STAT3 phosphorylation and synergized with IFN-γ in inhibiting TSC2-null and LAMD cell proliferation. Reduction of STAT3 protein levels or activity using specific small interfering RNA or inhibitory peptide, respectively, decreased proliferation and induced apoptosis in TSC2-null and LAMD cells and sensitized cells to growth-inhibitory and proapoptotic effects of IFN-γ. Collectively, our data demonstrate that STAT3 activation is required for proliferation and survival of cells with TSC2 dysfunction, that STAT3 impedes growth-inhibitory and proapoptotic effects of IFN-γ, and that TSC2- and rapamycin-dependent inhibition of STAT3 restores antiproliferative effects of IFN-γ. Thus, STAT3 may provide a novel therapeutic target for diseases associated with TSC1/TSC2 dysfunction.
- TSC, tuberous sclerosis complex
- mTOR, mammalian target of rapamycin
- RAPA, rapamycin
- S6K1, S6 kinase 1
- LAM, lymphangioleiomyomatosis
- SM, smooth muscle
- IFN, interferon
- STAT, signal transducer and activator of transcription
- JAK, Janus tyrosine kinase
- MEF, mouse embryonic fibroblast
- LAMD, lymphangioleiomyomatosis-dissociated
- HBF, human bronchus fibroblast
- PBS, phosphate-buffered saline
- DAPI, 4,6-diamidino-2-phenylindole
- siRNA, small interfering RNA
- pEGFP, plasmid encoding for enhanced green fluorescent protein
- PDGF, platelet-derived growth factor
- BrdU, 5-bromo-2′-deoxyuridine
- GFP, green fluorescent protein
- ANOVA, analysis of variance
- CCI-779, temsirolimus
- NDRI, National Disease Research Interchange.
Footnotes
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The online version of this article (available at http://molpharm.aspetjournals.org) contains supplemental material.
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This work was supported by the National Institutes of Health National Heart, Lung, and Blood Institute [Grants 2R01-HL071106, 1R01-HL090829, HL55301, HL64063, R00-HL089409-03]; the Abramson Cancer Center Pilot Project Grant; the American Thoracic Society/LAM Foundation [Grant LAM-07-001]; the American Lung Association [Grant RG-49342-N]; and the Parker B. Francis Fellowship.
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Article, publication date, and citation information can be found at http://molpharm.aspetjournals.org.
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ABBREVIATIONS:
- Received April 21, 2009.
- Accepted July 13, 2009.
- © 2009 The American Society for Pharmacology and Experimental Therapeutics
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