Lymphangioleiomyomatosis (LAM) is a genetic disorder characterized by benign lesions of smooth muscle-like LAM cells in the airways and blood and lymph vessels of the lung. The molecular mechanisms triggering LAM cell proliferation remain largely unknown. Genetic studies demonstrated that somatic mutations in the tumor suppressor TSC2 gene are associated with pulmonary LAM. Our preliminary studies demonstrate, for the first time, that without extracellular stimuli loss of tuberin induces the constitutive activation of p70 S6 kinase (p70S6K) and the constitutive phosphorylation of ribosomal protein S6. In this application, we propose to dissect the intracellular molecular signaling mechanisms by which tuberin deficiency, in the absence of extracellular stimuli, leads to abnormal cell cycle progression and proliferation, focusing on defining the roles of the functional domains of tuberin as well as interactions with p70S6k, Rab5, and mTOR. The central hypothesis of the proposal is that loss of the tuberin function promotes LAM cell cycle progression and LAM cell proliferation by the constitutive activation of p70S6K. Rab5 and mTOR serve as downstream effectors of tuberin and upstream modulators of the constitutive activation of p70S6K. To test this hypothesis, in Aim 1, we will determine the role of the structural/functional domains of tuberin in the constitutive activation of p70S6K and cell proliferation.
In Aim 2, the role of Rab5 in modulating cell cycle progression and proliferation of tuberin-deficient cells by inducing the constitutive activation of p70S6K, will be examined.
In Aim 3, the role of mTOR in modulating the constitutive activation of p7036k will be established. Transfection, microinjection, flow cytometry, immunocytochemistry, immunoblot, DNA synthesis analysis, and Rab5, p70S6K, mTOR activity assays will be utilized to dissect these signaling pathways. These studies will provide insight into the molecular mechanisms that may be important in the pathogenesis of LAM; this understanding may lead to new therapeutic approaches to treat this devastating disease and to improve understanding of the molecular signaling pathways regulating mesenchymal cell proliferation.

Agency
National Institute of Health (NIH)
Institute
National Heart, Lung, and Blood Institute (NHLBI)
Type
Research Project (R01)
Project #
5R01HL071106-04
Application #
6901814
Study Section
Special Emphasis Panel (ZRG1-RESP (01))
Program Officer
Peavy, Hannah H
Project Start
2002-07-01
Project End
2007-02-28
Budget Start
2005-07-01
Budget End
2007-02-28
Support Year
4
Fiscal Year
2005
Total Cost
$277,375
Indirect Cost
Name
University of Pennsylvania
Department
Internal Medicine/Medicine
Type
Schools of Medicine
DUNS #
042250712
City
Philadelphia
State
PA
Country
United States
Zip Code
19104
Goncharova, Elena A; James, Melane L; Kudryashova, Tatiana V et al. (2014) Tumor suppressors TSC1 and TSC2 differentially modulate actin cytoskeleton and motility of mouse embryonic fibroblasts. PLoS One 9:e111476
Goncharova, Elena A; Goncharov, Dmitry A; Fehrenbach, Melane et al. (2012) Prevention of alveolar destruction and airspace enlargement in a mouse model of pulmonary lymphangioleiomyomatosis (LAM). Sci Transl Med 4:154ra134
Krymskaya, Vera P (2012) Treatment option(s) for pulmonary lymphangioleiomyomatosis: progress and current challenges. Am J Respir Cell Mol Biol 46:563-5
Goncharova, Elena A; Goncharov, Dmitry A; Li, Hua et al. (2011) mTORC2 is required for proliferation and survival of TSC2-null cells. Mol Cell Biol 31:2484-98
Krymskaya, Vera P; Snow, Jennifer; Cesarone, Gregory et al. (2011) mTOR is required for pulmonary arterial vascular smooth muscle cell proliferation under chronic hypoxia. FASEB J 25:1922-33
Goncharova, Elena A; Lim, Poay N; Chisolm, Amelia et al. (2010) Interferons modulate mitogen-induced protein synthesis in airway smooth muscle. Am J Physiol Lung Cell Mol Physiol 299:L25-35
Krymskaya, Vera P; Goncharova, Elena A (2009) PI3K/mTORC1 activation in hamartoma syndromes: therapeutic prospects. Cell Cycle 8:403-13
Goncharova, Elena A; Goncharov, Dmitry A; Damera, Gautam et al. (2009) Signal transducer and activator of transcription 3 is required for abnormal proliferation and survival of TSC2-deficient cells: relevance to pulmonary lymphangioleiomyomatosis. Mol Pharmacol 76:766-77
Goncharova, Elena A; Goncharov, Dmitry A; Chisolm, Amelia et al. (2008) Interferon beta augments tuberous sclerosis complex 2 (TSC2)-dependent inhibition of TSC2-null ELT3 and human lymphangioleiomyomatosis-derived cell proliferation. Mol Pharmacol 73:778-88
Krymskaya, Vera P (2008) Smooth muscle-like cells in pulmonary lymphangioleiomyomatosis. Proc Am Thorac Soc 5:119-26

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