Tumor Cell-mediated Induction of the Stromal Factor Stromelysin-3 Requires Heterotypic Cell Contact-dependent Activation of Specific Protein Kinase C Isoforms

doi:10.1074/jbc.M405482200

Tumor Cell-mediated Induction of the Stromal Factor Stromelysin-3 Requires Heterotypic Cell Contact-dependent Activation of Specific Protein Kinase C Isoforms*

^‡INSERM U526, IFR50, Faculté de Médecine Pasteur, 06107 Nice, France, the ^¶INSERM U469, CCIPE, 34094 Montpellier, France, the ^**INSERM U575, Université Louis Pasteur, 67084 Strasbourg, France, and the ^‡‡Dana-Farber Cancer Institute, Harvard Medical School, Boston, Massachusetts 02115

§§ To whom correspondence should be addressed: INSERM U526, Faculté deMédecine Pasteur, 06107 Nice, France. Tel.: 33-493-377-017; Fax: 33-493-817-852; E-mail: bernard.mari{at}unice.fr.

Abstract

Stromelysin-3 (ST3, MMP-11) has been shown to be strongly overexpressed in stromal fibroblasts of most invasive human carcinomas. However, the molecular mechanisms leading to ST3 expression in nonmalignant fibroblasts remain unknown. The aim of the present study was to analyze the signaling pathways activated in normal pulmonary fibroblasts after their interaction with non-small cell lung cancer (NSCLC) cells and leading to ST3 expression. The use of selective signaling pathway inhibitors showed that conventional and novel protein kinase Cs (PKC) were required for ST3 induction, whereas Src kinases exerted a negative control. We observed by both conventional and real time confocal microscopy that green fluorescent protein-tagged PKCα and PKCϵ, but not PKCδ, transfected in fibroblasts, accumulate selectively at the cell-cell contacts between fibroblasts and tumor cells. In agreement, RNAi-mediated depletion of PKCα and PKCϵ, but not PKCδ significantly decreased co-culture-dependent ST3 production. Finally, a tetracycline-inducible expression model allowed us to confirm the central role of these PKC isoforms and the negative regulatory function of c-Src in the control of ST3 expression. Altogether, our data emphasize signaling changes occurring in the tumor microenvironment that may define new stromal targets for therapeutic intervention.

Footnotes

↵1 The abbreviations used are: MMP, matrix metalloproteinases; CM, conditioned medium; EGF, epithelial growth factor; MAPK, mitogen-activated protein kinase; MMP, matrix metalloproteinase; NSCLC, non-small cell lung cancer; PKC, protein kinase C; PMA, phorbol 12-myristate 13-acetate; ST3, stromelysin-3; Tet, tetracycline; TIMP, tissue inhibitor of matrix metalloproteinases; C/EBP, CCAAT/enhancer-binding protein; Ab, antibody; siRNA, small interfering RNA; cPKC, conventional PKC; nPKC, novel PKC; JNK, c-Jun N-terminal kinase; CA, constitutively active; DN, dominant negative; GFP green fluorescent protein.
↵* This work was supported in part by INSERM, University of Nice-Sophia Antipolis, the Fondation de France, and the Association pour la Recherche Contre le Cancer Contract 3355. The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
↵§ Supported by a fellowship from the Ligue Nationale contre Le Cancer.
↵∥ Supported by a fellowship from the Fondation de France. Present address: INSERM U606, Hôpital Lariboisière, rue Ambroise Paré, 75475 Paris, France.
- Received May 17, 2004.
- Revision received September 27, 2004.
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