Endogenous spartin, mutated in hereditary spastic paraplegia, has a complex subcellular localization suggesting diverse roles in neurons

Robay, Dimitri; Patel, Harshad B.; Simpson, Michael A.; Brown, Nigel A.; Crosby, Andrew H.

doi:10.1016/j.yexcr.2006.05.003

Cited by 29 publications

(48 citation statements)

References 47 publications

(62 reference statements)

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“…Using immunoblotting, we observed protein bands representing endogenous spartin at ϳ85 and ϳ95 kDa in different human cell lines, which concurs with another report using a different antibody (Robay et al, 2006), and we demonstrated that the 95-kDa band represents a mono-ubiquitinated form of spartin. We estimate that ϳ30 -35% of endogenous spartin is mono-ubiquitinated and that this modification does not depend on EGF stimulation, indicating that spartin constitutively undergoes this post-translational modification.…”

Section: Discussionsupporting

confidence: 91%

“…Recent studies have localized endogenous spartin to the trans-Golgi network, neuritic processes, and nucleus (Robay et al, 2006) and overexpressed spartin to mitochondria (Lu et al, 2006). We previously showed that spartin is both a cytosolic and membrane-associated protein that interacts with Eps15 and Eps15R (Bakowska et al, 2005).…”

Section: Introductionmentioning

confidence: 99%

“…Although a region adjacent to the MIT domain is involved in the interaction of spastin with microtubules (Errico et al, 2004), spastin's MIT domain interacts with charged multivesicular body protein 1B (CHMP1B; Reid et al, 2005), a member of the endosomal sorting complex required for transport (ESCRT)-III complex, which is critical for the sorting of membrane cargoes to the multivesicular bodies (Babst et al, 2002). Structural studies of the MIT domain of Vps4-A indicate that it also interacts with the charged multivesicular body protein family of ESCRT-III proteins (Scott et al, 2005).Recent studies have localized endogenous spartin to the trans-Golgi network, neuritic processes, and nucleus (Robay et al, 2006) and overexpressed spartin to mitochondria (Lu et al, 2006). We previously showed that spartin is both a cytosolic and membrane-associated protein that interacts with Eps15 and Eps15R (Bakowska et al, 2005).…”

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confidence: 99%

“…This impairment might be caused by dysfunctions at pre-and/or postsynaptic structures, but at present we can only speculate about the function of spartin in neurons. A recent study found spartin in synaptic nerve terminals (Robay et al, 2006), and the depletion of EHS-1, an ortholog of Eps15 in Caenorhabditis elegans, resulted in a temperature-dependent defect in movement as well as a decreased number of synaptic vesicles (Salcini et al, 2001). Thus, spartin, similar to Eps15, might be an accessory protein involved in clathrinmediated endocytosis of synaptic vesicles in presynaptic nerve terminals.…”

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confidence: 99%

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Troyer Syndrome Protein Spartin Is Mono-Ubiquitinated and Functions in EGF Receptor Trafficking

Bakowska

Jupille

Fatheddin

et al. 2007

MBoC

133

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Troyer syndrome is an autosomal recessive hereditary spastic paraplegia caused by mutation in the spartin (SPG20) gene, which encodes a widely expressed protein of unknown function. This mutation results in premature protein truncation and thus might signify a loss-of-function disease mechanism. In this study, we have found that spartin is monoubiquitinated and functions in degradation of the epidermal growth factor receptor (EGFR). Upon EGF stimulation, spartin translocates from the cytoplasm to the plasma membrane and colocalizes with internalized EGF-Alexa. Knockdown of spartin by small interfering RNA decreases the rate of EGFR degradation and also affects EGFR internalization, recycling, or both. Furthermore, overexpression of spartin results in a prominent decrease in EGFR degradation. Taken together, our data suggest that spartin is involved in the intracellular trafficking of EGFR and that impaired endocytosis may underlie the pathogenesis of Troyer syndrome. INTRODUCTIONThe hereditary spastic paraplegias (HSPs) comprise a cluster of inherited neurological disorders characterized by progressive spasticity and muscle weakness in the lower limbs Reid, 2003;Fink, 2006;Soderblom and Blackstone, 2006). Classically, the HSPs have been divided into two forms: "pure" when lower extremity spasticity and paraparesis are the only features and "complicated" when additional symptoms are present (Harding, 1983). More than 30 genetic loci (SPG1-33) and 14 gene products have been identified, yielding new insights into the molecular pathways involved in the pathogenesis of the HSPs. Troyer syndrome (SPG20), an autosomal recessive, complicated HSP that manifests in early childhood, is characterized by dysarthria, mental retardation, shortness of stature, and distal muscle wasting in addition to spasticity and weakness of the lower limbs (Cross and McKusick, 1967;Proukakis et al., 2004). To date, the only known mutation involved in Troyer syndrome is a single base deletion in the spartin gene, resulting in a 29-amino acid substitution at the C-terminus and premature truncation of the 666-amino acid protein by 268 residues (Patel et al., 2002;Proukakis et al., 2004).Although the cellular functions of spartin are not known, it harbors two conserved domains, an MIT (contained within microtubule-interacting and trafficking molecules) domain at the N-terminus and a plant-related region at the C-terminus. Although the only known function of the latter domain is in senescence (Panavas et al., 1999), the MIT domain is found in several other proteins, including Vps4-A and -B, sortin 15, and spastin (Ciccarelli et al., 2003), which are involved in vesicle trafficking and binding of microtubules. Interestingly, spastin, a microtubule-severing ATPase (Errico et al., 2002;Trotta et al., 2004;Evans et al., 2005), is mutated in the most common form of autosomal dominant HSP, SPG4 Reid, 2003;Fink, 2006;Soderblom and Blackstone, 2006). Although a region adjacent to the MIT domain is involved in the interaction of spastin with microtubules ...

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Section: Discussionsupporting

confidence: 91%

Section: Introductionmentioning

confidence: 99%

mentioning

confidence: 99%

mentioning

confidence: 99%

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Troyer Syndrome Protein Spartin Is Mono-Ubiquitinated and Functions in EGF Receptor Trafficking

Bakowska

Jupille

Fatheddin

et al. 2007

MBoC

133

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“…Most studies to date have relied on overexpression systems that have given conflicting results. A single study reporting the localization of the endogenous protein used an antibody whose specificity was not verifed by knockdown experiments and gave discrepant results to all of the overexpression studies [23]. To resolve this controversy, we used a novel spartin antibody in immunofluorescence experiments.…”

Section: Discussionmentioning

confidence: 99%

Endogenous spartin (SPG20) is recruited to endosomes and lipid droplets and interacts with the ubiquitin E3 ligases AIP4 and AIP5

Edwards¹,

Clowes²,

Tsang³

et al. 2009

Biochemical Journal

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The HSPs (hereditary spastic paraplegias) are genetic conditions in which there is distal degeneration of the longest axons of the corticospinal tract, resulting in spastic paralysis of the legs. The gene encoding spartin is mutated in Troyer syndrome, an HSP in which paralysis is accompanied by additional clinical features. There has been controversy over the subcellular distribution of spartin. We show here that, at steady state, endogenous spartin exists in a cytosolic pool that can be recruited to endosomes and to lipid droplets. Cytosolic endogenous spartin is monoubiquitinated and we demonstrate that it interacts via a PPXY motif with the ubiquitin E3 ligases AIP4 [atrophin-interacting protein 4; WWP2 (WW domain-containing E3 ubiquitin protein ligase 2] and AIP5 (WWP1). Surprisingly, the PPXY motif, AIP4 and AIP5 are not required for spartin's ubiquitination, and so we propose that spartin acts as an adaptor for these proteins. Our results suggest that spartin is involved in diverse cellular functions, which may be of relevance to the complex phenotype seen in Troyer syndrome.

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Lack of Spartin Protein in Troyer Syndrome

Bakowska¹,

Wang²,

Xin³

et al. 2008

Arch Neurol

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Background: Hereditary spastic paraplegias (SPG1-SPG33) are characterized by progressive spastic weakness of the lower limbs. A nucleotide deletion (1110delA) in the (SPG20; OMIM 275900) spartin gene is the origin of autosomal recessive Troyer syndrome. This mutation is predicted to cause premature termination of the spartin protein. However, it remains unknown whether this truncated spartin protein is absent or is present and partially functional in patients. Objective: To determine whether the truncated spartin protein is present or absent in cells derived from patients with Troyer syndrome.

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Endogenous spartin, mutated in hereditary spastic paraplegia, has a complex subcellular localization suggesting diverse roles in neurons

Cited by 29 publications

References 47 publications

Troyer Syndrome Protein Spartin Is Mono-Ubiquitinated and Functions in EGF Receptor Trafficking

Troyer Syndrome Protein Spartin Is Mono-Ubiquitinated and Functions in EGF Receptor Trafficking

Endogenous spartin (SPG20) is recruited to endosomes and lipid droplets and interacts with the ubiquitin E3 ligases AIP4 and AIP5

Lack of Spartin Protein in Troyer Syndrome

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