K-ras nanoclustering is subverted by overexpression of the scaffold protein galectin-3.
The spatial organization of K-Ras proteins into nanoclusters on the plasma membrane is essential for high-fidelity signal transduction. The mechanism underlying K-Ras nanoclustering is unknown. We show here that K-Ras.GTP recruits Galectin-3 (Gal-3) from the cytosol to the plasma membrane where it becomes an integral nanocluster component. Importantly, we show that the cytosolic level of Gal-3 determines the magnitude of K-Ras.GTP nanoclustering and signal output. The beta-sheet layers of the Gal-3 carbohydrate recognition domain contain a hydrophobic pocket that may accommodate the farnesyl group of K-Ras. V125A substitution within this hydrophobic pocket yields a dominant negative Gal-3(V125A) mutant that inhibits K-Ras activity. Gal-3(V125A) interaction with K-Ras.GTP reduces K-Ras.GTP nanocluster formation, which abrogates signal output from the Raf/mitogen-activated protein (MAP)/extracellular signal-regulated kinase (ERK; MEK) pathway. Gal-3(V125A) negatively regulates cell growth and reduces cellular transformation. Thus, regulation of K-Ras nanocluster formation and signal output by Gal-3 critically depends on the integrity of the Gal-3 hydrophobic pocket. These results show that Gal-3 overexpression in breast cancer cells, which increases K-Ras signal output, represents oncogenic subversion of plasma membrane nanostructure.
Shalom-Feuerstein
Ruby
R
Department of Neurobiology, The George S. Wise Faculty of Life Sciences, Tel Aviv University, Tel Aviv, Israel.
Plowman
Sarah J
SJ
Rotblat
Barak
B
Ariotti
Nicholas
N
Tian
Tianhai
T
Hancock
John F
JF
Kloog
Yoel
Y
D000818:Animals; D001943:Breast Neoplasms / Q000378:metabolism* / Q000473:pathology; D002448:Cell Adhesion; D049109:Cell Proliferation; D002478:Cells, Cultured; D006224:Cricetinae; D003593:Cytoplasm / Q000378:metabolism; D048049:Extracellular Signal-Regulated MAP Kinases / Q000378:metabolism; D005434:Flow Cytometry; D037502:Galectin 3 / Q000235:genetics / Q000378:metabolism*; D011905:Genes, ras / Q000502:physiology*; D006160:Guanosine Triphosphate / Q000378:metabolism; D006801:Humans; D015151:Immunoblotting; D047468:Immunoprecipitation; D007668:Kidney / Q000378:metabolism* / Q000473:pathology; D018613:Microscopy, Confocal; D008856:Microscopy, Fluorescence; D020929:Mitogen-Activated Protein Kinase Kinases / Q000378:metabolism; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism; D019908:Proto-Oncogene Proteins c-raf / Q000378:metabolism; D015398:Signal Transduction
D037502:Galectin 3; D006160:Guanosine Triphosphate; D019908:Proto-Oncogene Proteins c-raf; D048049:Extracellular Signal-Regulated MAP Kinases; D020928:Mitogen-Activated Protein Kinases; D020929:Mitogen-Activated Protein Kinase Kinases