文档介绍：SPG20 ***@20050820 1 Spartin: Localization to the mitochondria and interaction with microtubule structure Key Words: Hereditary spastic paraplegia HSP, Spartin SPG20, Mitochondria, MIT Abbreviations list HSP Hereditary spastic paraplegia SPG, spastic paraplegia/ paraparesis gene aa amino acid residual(s) ER endoplasmic reticulum MIT Microtubule Interacting and Trafficking TRS Troyer syndrome SPG20 ***@20050820 2 Abstract Hereditary spastic paraplegia (HSP) describes a diverse group of disorders characterized by length dependent axonal degeneration that causes progressive paraparesis primarily affecting the lower limbs. Troyer syndrome (TRS) isa rare autosomal recessive form of HSP where affected patients have characteristic spastic paraparesis due to retrograde axonal degeneration. TRS is caused by mutations in SPG20, one of which 1100 del A leads toa frameshift in exon 4 and premature termination of corresponding protein spartin (fs369-398x399 ). Normal spartin has 666 aa with a microtubule interacting and trafficking (MIT) domain at 36-114aa. However the function of this protein is unknown and there are no reported studies of its subcellular localisation. We cloned full- length spartin cDNA into a eukaryotic expression vector inframe with an enhanced fluorescent protein (ECFP-Spartin). We also constructed plasmids expressing prematurely truncated spartin (ECFP-Spartin 1-398 )orN terminal deleted spartin (ECFP-Spartin 347-666 ) parison. The Spartin 1- 398 has the same amino acid sequences to that reported in TRS patients. Confocal microscopy ofa variety of transfected neuronal and non-neuronal cells revealed cytoplasmic expression that was primarily punctuate with a weak diffuse cytoplasmic distribution. We established that full-length spartin is located in mitochondria with anelle marker or immunocytochemistry staining. Colocalization measurement revealed that the ECFP-Spartin colocalized with mitochondria and tubulin. Analysis of cells expressing either ECFP-Spartin 1-