Overexpression of ROCK in human breast cancer cells: evidence that ROCK activity mediates intracellular membrane traffic of lysosomes.
Small GTPase Rho and its downstream effectors, ROCK family of Rho-associated serine-threonine kinases, are thought to participate in cell morphology, motility, and tumor progression through regulating the rearrangement of actin cytoskeleton. Here we present evidence that transfection of human breast cancer cells with cDNA encoding a dominant active mutant of ROCK causes dispersal of lysosomal vesicles throughout the cytoplasm without perturbing the machinery of the endocytic pathway. The intracellular distribution of lysosomes and endocytosed transferrin, an early endosomal marker, were further assessed by confocal immunofluorescence microscopy. In the active ROCK transfected cells the lysosomal proteins, cathepsin D, LIMPII, and LAMP1, were found throughout the cytoplasm in dispersed small vesicles, which were accessible to the endocytosed Texas Red-labeled transferrin. 3D-image analysis of lysosomal distribution in the active ROCK transfectants revealed abundant punctate signals in the peripheral region of the basal plasma membrane. Cells expressing vector alone did not exhibit these alterations. Wortmannin, a phosphatidylinositol 3-kinase inhibitor, induced LIMPII-positive/ transferrin negative large vacuoles in the perinuclear region, and disappearence of the dispersed small vesicular structures. To our knowledge, this is the first evidence that increasing ROCK expression contributes to selective cellular dispersion of lysosomes in invasive breast cancer cells.
Nishimura
Yukio
Y
Graduate School of Pharmaceutical Sciences, Kyushu University, Fukuoka 812-8582, Japan. nishimur@bioc.phar.kyushu-u.ac.jp
Itoh
Kazuyuki
K
Yoshioka
Kiyoko
K
Tokuda
Kazuo
K
Himeno
Masaru
M
D000730:Androstadienes / Q000494:pharmacology; D015703:Antigens, CD / Q000378:metabolism; D001943:Breast Neoplasms / Q000201:enzymology*; D018955:CD36 Antigens / Q000378:metabolism; D002402:Cathepsin D / Q000378:metabolism; D004705:Endocytosis / Q000502:physiology; D011992:Endosomes / Q000187:drug effects / Q000201:enzymology / Q000648:ultrastructure; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005260:Female; D006801:Humans; D007425:Intracellular Membranes / Q000201:enzymology*; D047908:Intracellular Signaling Peptides and Proteins; D052118:Lysosomal-Associated Membrane Protein 1; D051907:Lysosomal Membrane Proteins; D008247:Lysosomes / Q000201:enzymology* / Q000648:ultrastructure; D008565:Membrane Proteins; D018613:Microscopy, Confocal; D008856:Microscopy, Fluorescence; D000081082:Phosphoinositide-3 Kinase Inhibitors; D017346:Protein Serine-Threonine Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D021381:Protein Transport; D051116:Receptors, Scavenger; D011990:Receptors, Transferrin / Q000276:immunology; D012795:Sialoglycoproteins; D015398:Signal Transduction; D014162:Transfection; D014168:Transferrin / Q000378:metabolism; D014407:Tumor Cells, Cultured; D015854:Up-Regulation; D000077191:Wortmannin; D054460:rho-Associated Kinases
D000730:Androstadienes; D015703:Antigens, CD; D018955:CD36 Antigens; D004791:Enzyme Inhibitors; D047908:Intracellular Signaling Peptides and Proteins; D052118:Lysosomal-Associated Membrane Protein 1; D051907:Lysosomal Membrane Proteins; D008565:Membrane Proteins; D000081082:Phosphoinositide-3 Kinase Inhibitors; D051116:Receptors, Scavenger; D011990:Receptors, Transferrin; C074299:SCARB2 protein, human; D012795:Sialoglycoproteins; D014168:Transferrin; D017346:Protein Serine-Threonine Kinases; D054460:rho-Associated Kinases; D002402:Cathepsin D; D000077191:Wortmannin