Involvement of a membrane-bound form of glutamate dehydrogenase in the association of lysosomes to microtubules.
A 50-kDa membrane protein corresponding to a membrane-bound isoform of glutamate dehydrogenase was proposed as a molecular species that could mediate lysosome-microtubule interactions. This protein, isolated from purified lysosome membranes, is a peripheral membrane protein with an ATP-dependent microtubule binding activity. We have produced antibodies against the purified 50-kDa protein to investigate its role in the association of lysosomes to microtubules using a cell-free reconstitution assay and cell microinjection. Pretreatment of purified lysosomes with the antibodies inhibited the association of these vesicles to microtubules. The blocking effect of antibodies was demonstrated by a differential sedimentation method and negative staining electron microscopy, allowing us to quantify the amount of microtubules interacting with lysosomes and the proportion of lysosomes bound to microtubules, respectively. Affinity-purified antibodies microinjected into intact cells altered the distribution of lysosomes that appeared less clustered in the vicinity of nuclei. The antibody-induced lysosome dispersion was assessed by quantitative videomicroscope analyses. These data show that the 50-kDa membrane protein could act, through its microtubule binding activity, as a molecule of attachment of lysosomes to microtubules. This membrane-bound isoform of glutamate dehydrogenase could be involved in the microtubule-dependent perinuclear localization of lysosomes.
Rajas
F
F
Institut National de la Santé et de la Recherche Médicale, Unité 369, Faculté de Médecine Lyon-RTH Laënnec, 69372 Lyon Cédex 08, France. u369@cimacpcu.univ-lyon1.fr
Gire
V
V
Rousset
B
B
D000818:Animals; D000906:Antibodies / Q000276:immunology; D000918:Antibody Specificity; D002462:Cell Membrane / Q000201:enzymology; D002478:Cells, Cultured; D005969:Glutamate Dehydrogenase / Q000276:immunology / Q000378:metabolism*; D008099:Liver / Q000166:cytology / Q000201:enzymology / Q000648:ultrastructure; D008247:Lysosomes / Q000378:metabolism*; D008854:Microscopy, Electron; D008856:Microscopy, Fluorescence; D008870:Microtubules / Q000378:metabolism*; D013552:Swine
D000906:Antibodies; D005969:Glutamate Dehydrogenase