Cytoplasmic dynein/dynactin mediates the assembly of aggresomes.
Aggresomes are pericentrosomal cytoplasmic structures into which aggregated, ubiquitinated, misfolded proteins are sequestered. Misfolded proteins accumulate in aggresomes when the capacity of the intracellular protein degradation machinery is exceeded. Previously, we demonstrated that an intact microtubule cytoskeleton is required for the aggresome formation [Johnston et al., 1998: J. Cell Biol. 143:1883-1898]. In this study, we have investigated the involvement of microtubules (MT) and MT motors in this process. Induction of aggresomes containing misfolded DeltaF508 CFTR is accompanied by a redistribution of the retrograde motor cytoplasmic dynein that colocalizes with aggresomal markers. Coexpression of the p50 (dynamitin) subunit of the dynein/dynactin complex prevents the formation of aggresomes, even in the presence of proteasome inhibitors. Using in vitro microtubule binding assays in conjunction with immunogold electron microscopy, our data demonstrate that misfolded DeltaF508 CFTR associate with microtubules. We conclude that cytoplasmic dynein/dynactin is responsible for the directed transport of misfolded protein into aggresomes. The implications of these findings with respect to the pathogenesis of neurodegenerative disease are discussed.
Johnston
Jennifer A
JA
Department of Biological Sciences, Stanford University, Stanford, California, USA. jjohnston@elanpharma.com
Illing
Michelle E
ME
Kopito
Ron R
RR
D000941:Antigens / Q000378:metabolism / Q000648:ultrastructure; D002478:Cells, Cultured; D019005:Cystic Fibrosis Transmembrane Conductance Regulator / Q000235:genetics / Q000378:metabolism; D003593:Cytoplasm / Q000378:metabolism* / Q000648:ultrastructure; D000072159:Dynactin Complex; D004398:Dyneins / Q000378:metabolism / Q000648:ultrastructure; D005057:Eukaryotic Cells / Q000378:metabolism* / Q000648:ultrastructure; D005455:Fluorescent Antibody Technique; D005786:Gene Expression Regulation / Q000502:physiology; D049452:Green Fluorescent Proteins; D006801:Humans; D007150:Immunohistochemistry; D002479:Inclusion Bodies / Q000378:metabolism* / Q000648:ultrastructure; D008164:Luminescent Proteins; D008854:Microscopy, Electron; D008869:Microtubule-Associated Proteins / Q000378:metabolism / Q000648:ultrastructure; D008870:Microtubules / Q000378:metabolism* / Q000648:ultrastructure; D020409:Molecular Motor Proteins / Q000378:metabolism* / Q000648:ultrastructure; D019636:Neurodegenerative Diseases / Q000378:metabolism / Q000503:physiopathology; D015388:Organelles / Q000378:metabolism* / Q000648:ultrastructure; D017510:Protein Folding; D021381:Protein Transport / Q000502:physiology; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism
D000941:Antigens; C505032:CFTR protein, human; C000606361:DCTN2 protein, human; D000072159:Dynactin Complex; D008164:Luminescent Proteins; D008869:Microtubule-Associated Proteins; D020409:Molecular Motor Proteins; D011993:Recombinant Fusion Proteins; C086178:pericentrin; D019005:Cystic Fibrosis Transmembrane Conductance Regulator; D049452:Green Fluorescent Proteins; D004398:Dyneins