Toxicity and endocytosis of spinocerebellar ataxia type 6 polyglutamine domains: role of myosin IIb |
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Authors: | Marquèze-Pouey Béatrice Martin-Moutot Nicole Sakkou-Norton Marie Lévêque Christian Ji Yong Cornet Véronique Hsiao Wendy L Seagar Michael |
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Affiliation: | INSERM U641, Marseille F-13916, France. marqueze@ciml.univ-mrs.fr |
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Abstract: | Spinocerebellar ataxia type 6 (SCA6) is a dominantly inherited neurodegenerative disease caused by a small expansion of CAG repeats in the sequence coding for the cytoplasmic C-terminal region of the Ca(v)2.1 subunit of P/Q-type calcium channels. We have tested the toxicity of mutated Ca(v)2.1 C-terminal domains expressed in the plasma membrane. In COS-7 cells, CD4-green fluorescent protein fused to Ca(v)2.1 C-terminal domains containing expanded 24 polyglutamine (Q) tracts displayed increased toxicity and stronger expression at the cell surface relative to 'normal' 12 Q tracts, partially because of reduced endocytosis. Glutathione S-transferase pull-down and proteomic analysis indicated that Ca(v)2.1 C-termini interact with the heavy and light chains of cerebellar myosin IIB, a molecular motor protein. This interaction was confirmed by coimmunoprecipitation from rat cerebellum and COS-7 cells and shown to be direct by binding of in vitro-translated (35)S-myosin IIB heavy chain. In COS-7 cells, incremented polyglutamine tract length increased the interaction with myosin IIB. Furthermore, the myosin II inhibitor blebbistatin reversed the effects of polyglutamine expansion on plasma membrane expression. Our findings suggest a key role of myosin IIB in promoting accumulation of mutant Ca(v)2.1Ct at the plasma membrane and suggest that this gain of function might contribute to the pathogenesis of SCA6. |
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Keywords: | CAG repeat calcium channels cell toxicity myosin neurodegenerative disease spinocerebellar ataxia |
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