Galectin-3 regulates RasGRP4-mediated activation of N-Ras and H-Ras.
Galectin-3 (Gal-3) is a pleiotropic beta-galactoside-binding protein expressed at relatively high levels in human neoplasms. Its carbohydrate recognition domain (CRD) contains a hydrophobic pocket that can accommodate the farnesyl moiety of K-Ras. Binding of K-Ras to Gal-3 stabilizes K-Ras in its active (GTP-bound) state. Gal-3, which does not interact with N-Ras, was nevertheless shown to reduce N-Ras-GTP in BT-549 cells by an unknown mechanism that we explored here. First, comparative analysis of various cancer cell lines (glioblastomas, breast cancer cells and ovarian carcinomas) showed a positive correlation between low N-Ras-GTP/high K-Ras-GTP phenotype and Gal-3 expression levels. Next we found that epidermal growth factor-stimulated GTP loading of N-Ras, but not of K-Ras, is blocked in cells expressing high levels of Gal-3. Activation of Ras guanine nucleotide releasing proteins (RasGRPs) by phorbol 12-myristate 13-acetate (PMA) or downregulation of Gal-3 by Gal-3 shRNA increased the levels of N-Ras-GTP in Gal-3 expressing cells. We further show that the N-terminal domain of Gal-3 interacts with and inhibits RasGRP4-mediated GTP loading on N-Ras and H-Ras proteins. Growth of BT-549 cells stably expressing the Gal-3 N-terminal domain was strongly attenuated. Overall, these experiments demonstrate a new control mechanism of Ras activation in cancer cells whereby the Gal-3 N-terminal domain inhibits activation of N-Ras and H-Ras proteins.
Shalom-Feuerstein
Ruby
R
Department of Neurobiochemistry, The George S. Wise Faculty of Life Sciences, Tel-Aviv University, 69978 Tel-Aviv, Israel.
Levy
Ran
R
Makovski
Victoria
V
Raz
Avraham
A
Kloog
Yoel
Y
D000818:Animals; D015153:Blotting, Western; D006224:Cricetinae; D004815:Epidermal Growth Factor / Q000494:pharmacology; D037502:Galectin 3 / Q000502:physiology*; D011905:Genes, ras / Q000502:physiology*; D006160:Guanosine Triphosphate / Q000378:metabolism; D006801:Humans; D047468:Immunoprecipitation; D009369:Neoplasms / Q000378:metabolism*; D034741:RNA, Small Interfering / Q000494:pharmacology; D051381:Rats; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D014407:Tumor Cells, Cultured / Q000187:drug effects / Q000378:metabolism; D020663:ras Guanine Nucleotide Exchange Factors / Q000378:metabolism*; D018631:ras Proteins / Q000378:metabolism*
D037502:Galectin 3; C462507:RASGRP4 protein, human; D034741:RNA, Small Interfering; D020663:ras Guanine Nucleotide Exchange Factors; D004815:Epidermal Growth Factor; D006160:Guanosine Triphosphate; D018631:ras Proteins; D013755:Tetradecanoylphorbol Acetate