title: The hereditary spastic paraplegia protein spastin interacts with the ESCRT-III complex-associated endosomal protein CHMP1B.
abstract: Pure hereditary spastic paraplegia is characterized by length-dependent degeneration of the distal ends of long axons. Mutations in spastin are the most common cause of the condition. We set out to investigate the function of spastin using a yeast two-hybrid approach to identify interacting proteins. Using full-length spastin as bait, we identified CHMP1B, a protein associated with the ESCRT (endosomal sorting complex required for transport)-III complex, as a binding partner. Several different approaches confirmed the physiological relevance of the interaction in mammalian cells. Epitope-tagged CHMP1B and spastin showed clear cytoplasmic co-localization in Cos-7 and PC12 cells. CHMP1B and spastin interacted specifically in vitro and in vivo in beta-lactamase protein fragment complementation assays, and spastin co-immunoprecipitated with CHMP1B. The interaction was mediated by a region of spastin lying between residues 80 and 196 and containing a microtubule interacting and trafficking domain. Expression of epitope-tagged CHMP1B in mammalian cells prevented the development of the abnormal microtubule phenotype associated with expression of ATPase-defective spastin. These data point to a role for spastin in intracellular membrane traffic events and provide further evidence to support the emerging recognition that defects in intracellular membrane traffic are a significant cause of motor neuron pathology.
authors.affiliation: Cambridge Institute for Medical Research, University of Cambridge, Cambridge, UK. ereid@hgmp.mrc.ac.uk
authors.forename: Evan
authors.initials: E
authors.lastname: Reid
authors.forename: James
authors.initials: J
authors.lastname: Connell
authors.forename: Thomas L
authors.initials: TL
authors.lastname: Edwards
authors.forename: Simon
authors.initials: S
authors.lastname: Duley
authors.forename: Stephanie E
authors.initials: SE
authors.lastname: Brown
authors.forename: Christopher M
authors.initials: CM
authors.lastname: Sanderson
mesh_terms: D000251:Adenosine Triphosphatases; D000818:Animals; D019556:COS Cells; D002135:Calcium-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D006224:Cricetinae; D056827:Endosomal Sorting Complexes Required for Transport; D011992:Endosomes / Q000378:metabolism*; D006801:Humans; D046912:Multiprotein Complexes / Q000235:genetics / Q000378:metabolism*; D009687:Nuclear Proteins / Q000235:genetics / Q000378:metabolism*; D016716:PC12 Cells; D011485:Protein Binding / Q000235:genetics / Q000502:physiology; D021381:Protein Transport / Q000235:genetics / Q000502:physiology; D051381:Rats; D015419:Spastic Paraplegia, Hereditary / Q000235:genetics / Q000378:metabolism*; D000074181:Spastin; D020798:Two-Hybrid System Techniques; D033921:Vesicular Transport Proteins
chemical_list: C443709:CHMP1A protein, human; C541483:CHMP1B protein, human; D002135:Calcium-Binding Proteins; D056827:Endosomal Sorting Complexes Required for Transport; D046912:Multiprotein Complexes; D009687:Nuclear Proteins; D033921:Vesicular Transport Proteins; D000251:Adenosine Triphosphatases; D000074181:Spastin; C400283:SPAST protein, human