Reexpression of the major protein kinase C substrate, SSeCKS, suppresses v-src-induced morphological transformation and tumorigenesis.
SSeCKS (pronounced essex) encodes a major protein kinase C substrate, the expression of which is down-regulated in src- and ras-transformed rodent fibroblasts but not in raf-transformed rodent fibroblasts (X. Lin et al., Mol. Cell. Biol., 15: 2754-2762, 1995). Using a panel of ras-transformed or revertant Rat-6 cells that exhibit selective parameters of transformation, we show that down-regulation of SSeCKS correlates with anchorage-independent growth. Cotransfection of NIH3T3 fibroblasts with an SSeCKS expression plasmid decreased 6-30-fold the ability of a v-src expressor plasmid to induce colonies in soft agar. To differentiate between possible tumor suppressive or growth-inhibitory effects of SSeCKS, we developed conditionally transformed cell lines (expressing ts72v-src) with tetracycline-regulated SSeCKS expression. SSeCKS suppressed the ability of v-src to induce increased cellular refractility, focus formation, soft agar colony formation, in vitro invasiveness in Matrigel, and growth in low serum (0.5%) but did not inhibit cell proliferation in high serum (10%) at the permissive (35 degrees C) temperature for src kinase activity. However, at the nonpermissive (39.5 degrees C) temperature, SSeCKS induced growth arrest. SSeCKS expression did not affect: (a) the protein level, in vivo or in vitro kinase activity of ts72src; (b) the activity of jun NH2-terminal kinase; and (c) the level of mitogen-activated protein kinase (extracellular signal-regulated kinase 2) protein. However, extracellular signal-regulated kinase 2 activity was induced 5-10-fold by SSeCKS in the presence of active src. SSeCKS reversed the ability of v-src to decrease the formation of vinculin-associated adhesion plaques, actin-based stress fibers, and filopodia structures. These data suggest a tumor suppressive role for SSeCKS via the control of cytoskeletal architecture and cell signaling.
Lin
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Department of Microbiology, Mount Sinai School of Medicine, New York, New York 10029-6574, USA.
Gelman
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D016475:3T3 Cells; D054758:A Kinase Anchor Proteins; D000818:Animals; D017209:Apoptosis; D015153:Blotting, Western; D018797:Cell Cycle Proteins; D002455:Cell Division; D002471:Cell Transformation, Neoplastic; D015536:Down-Regulation; D019084:Fluorescent Antibody Technique, Indirect; D015972:Gene Expression Regulation, Neoplastic; D051379:Mice; D008934:Mitogens / Q000235:genetics / Q000378:metabolism*; D014411:Neoplastic Stem Cells; D015688:Oncogene Protein pp60(v-src) / Q000235:genetics / Q000378:metabolism*; D010590:Phalloidine / Q000378:metabolism; D011493:Protein Kinase C / Q000378:metabolism; D014158:Transcription, Genetic; D014407:Tumor Cells, Cultured; D016596:Vinculin / Q000378:metabolism
D054758:A Kinase Anchor Proteins; C491161:Akap12 protein, mouse; D018797:Cell Cycle Proteins; D008934:Mitogens; D016596:Vinculin; D010590:Phalloidine; D015688:Oncogene Protein pp60(v-src); D011493:Protein Kinase C