Mutant huntingtin impairs post-Golgi trafficking to lysosomes by delocalizing optineurin/Rab8 complex from the Golgi apparatus.
Huntingtin regulates post-Golgi trafficking of secreted proteins. Here, we studied the mechanism by which mutant huntingtin impairs this process. Colocalization studies and Western blot analysis of isolated Golgi membranes showed a reduction of huntingtin in the Golgi apparatus of cells expressing mutant huntingtin. These findings correlated with a decrease in the levels of optineurin and Rab8 in the Golgi apparatus that can be reverted by overexpression of full-length wild-type huntingtin. In addition, immunoprecipitation studies showed reduced interaction between mutant huntingtin and optineurin/Rab8. Cells expressing mutant huntingtin produced both an accumulation of clathrin adaptor complex 1 at the Golgi and an increase of clathrin-coated vesicles in the vicinity of Golgi cisternae as revealed by electron microscopy. Furthermore, inverse fluorescence recovery after photobleaching analysis for lysosomal-associated membrane protein-1 and mannose-6-phosphate receptor showed that the optineurin/Rab8-dependent post-Golgi trafficking to lysosomes was impaired in cells expressing mutant huntingtin or reducing huntingtin levels by small interfering RNA. Accordingly, these cells showed a lower content of cathepsin D in lysosomes, which led to an overall reduction of lysosomal activity. Together, our results indicate that mutant huntingtin perturbs post-Golgi trafficking to lysosomal compartments by delocalizing the optineurin/Rab8 complex, which, in turn, affects the lysosomal function.
del Toro
Daniel
D
Departament de Biologia Cellular, Immunologia i Neurociències, Facultat de Medicina, IDIBAPS, Universitat de Barcelona, E-08036 Barcelona, Spain.
Alberch
Jordi
J
Lázaro-Diéguez
Francisco
F
Martín-Ibáñez
Raquel
R
Xifró
Xavier
X
Egea
Gustavo
G
Canals
Josep M
JM
D000818:Animals; D018797:Cell Cycle Proteins; D002462:Cell Membrane / Q000378:metabolism; D002478:Cells, Cultured; D005136:Eye Proteins / Q000032:analysis / Q000378:metabolism*; D006056:Golgi Apparatus / Q000378:metabolism*; D000071058:Huntingtin Protein; D007150:Immunohistochemistry; D008247:Lysosomes / Q000378:metabolism*; D026901:Membrane Transport Proteins; D051379:Mice; D009154:Mutation; D009419:Nerve Tissue Proteins / Q000032:analysis / Q000235:genetics / Q000502:physiology*; D009687:Nuclear Proteins / Q000032:analysis / Q000235:genetics / Q000502:physiology*; D021381:Protein Transport / Q000502:physiology; D018808:Transcription Factor AP-1 / Q000378:metabolism; D020691:rab GTP-Binding Proteins / Q000032:analysis / Q000378:metabolism*
D018797:Cell Cycle Proteins; D005136:Eye Proteins; C506812:Htt protein, mouse; D000071058:Huntingtin Protein; D026901:Membrane Transport Proteins; D009419:Nerve Tissue Proteins; D009687:Nuclear Proteins; C501288:Optn protein, mouse; C521976:Rab8 protein, mouse; D018808:Transcription Factor AP-1; D020691:rab GTP-Binding Proteins