TGF-β/SMAD3 Pathway Stimulates Sphingosine-1 Phosphate Receptor 3 Expression: IMPLICATION OF SPHINGOSINE-1 PHOSPHATE RECEPTOR 3 IN LUNG ADENOCARCINOMA PROGRESSION

J Biol Chem. 2016 Dec 30;291(53):27343-27353. doi: 10.1074/jbc.M116.740084. Epub 2016 Nov 17.

Abstract

Previously, we showed that levels of sphingosine-1 phosphate receptor 3 (S1PR3) are increased in a panel of cultured human lung adenocarcinoma cell lines, and that S1PR3-mediated signaling pathways regulate proliferation, soft agar growth, and invasion of human lung adenocarcinoma cells in vitro In the present study, we examine S1PR3 levels in human lung adenocarcinoma specimens. cDNA array and tumor microarray analysis shows that mRNA and protein levels of S1PR3 are significantly increased in human lung adenocarcinomas when compared with normal lung epithelial cells. Promoter analysis shows 16 candidate SMAD3 binding sites in the promoter region of S1PR3. ChIP indicates that TGF-β treatment stimulates the binding of SMAD3 to the promoter region of S1PR3. Luciferase reporter assay demonstrates that SMAD3 transactivates S1PR3 promoter. TGF-β stimulation or ectopic expression of TGF-β up-regulates S1PR3 levels in vitro and ex vivo Pharmacologic inhibition of TGF-β receptor or SMAD3 abrogates the TGF-β-stimulated S1PR3 up-regulation. Moreover, S1PR3 knockdown dramatically inhibits tumor growth and lung metastasis, whereas ectopic expression of S1PR3 promotes the growth of human lung adenocarcinoma cells in animals. Pharmacological inhibition of S1PR3 profoundly inhibits the growth of lung carcinoma in mice. Our studies suggest that levels of S1PR3 are up-regulated in human lung adenocarcinomas, at least in part due to the TGF-β/SMAD3 signaling axis. Furthermore, S1PR3 activity promotes the progression of human lung adenocarcinomas. Therefore, S1PR3 may represent a novel therapeutic target for the treatment of deadly lung adenocarcinomas.

Keywords: S1PR3; SMAD transcription factor; TGF-β; lung cancer; sphingolipid; sphingosine-1 phosphate (S1P); transforming growth factor β (TGF-B).

MeSH terms

  • Adenocarcinoma / genetics
  • Adenocarcinoma / metabolism
  • Adenocarcinoma / secondary*
  • Animals
  • Apoptosis
  • Blotting, Western
  • Cell Proliferation
  • Cells, Cultured
  • Female
  • Humans
  • Lung / metabolism
  • Lung / pathology
  • Lung Neoplasms / genetics
  • Lung Neoplasms / metabolism
  • Lung Neoplasms / pathology*
  • Mice
  • Mice, Inbred C57BL
  • Mice, Inbred NOD
  • Mice, SCID
  • RNA, Messenger / genetics
  • Real-Time Polymerase Chain Reaction
  • Receptors, Lysosphingolipid / genetics
  • Receptors, Lysosphingolipid / metabolism*
  • Reverse Transcriptase Polymerase Chain Reaction
  • Smad3 Protein / genetics
  • Smad3 Protein / metabolism*
  • Sphingosine-1-Phosphate Receptors
  • Transforming Growth Factor beta / genetics
  • Transforming Growth Factor beta / metabolism*
  • Xenograft Model Antitumor Assays

Substances

  • RNA, Messenger
  • Receptors, Lysosphingolipid
  • SMAD3 protein, human
  • Smad3 Protein
  • Sphingosine-1-Phosphate Receptors
  • Transforming Growth Factor beta
  • sphingosine-1-phosphate receptor-3, human