Faculty Opinions recommendation of Structural basis of microtubule severing by the hereditary spastic paraplegia protein spastin.

Lipid droplets (LDs) are metabolic organelles that store neutral lipids and dynamically respond to changes in energy availability by accumulating or mobilizing triacylglycerols (TAGs). How the plastic behavior of LDs is regulated is poorly understood. Hereditary spastic paraplegia is a central motor axonopathy predominantly caused by mutations in SPAST, encoding the microtubule-severing protein spastin. The spastin-M1 isoform localizes to nascent LDs in mammalian cells; however, the mechanistic significance of this targeting is not fully explained. Here, we show that tightly controlled levels of spastin-M1 are required to inhibit LD biogenesis and TAG accumulation. Spastin-M1 maintains the morphogenesis of the ER when TAG synthesis is prevented, independent from microtubule binding. Moreover, spastin plays a microtubule-dependent role in mediating the dispersion of LDs from the ER upon glucose starvation. Our results reveal a dual role of spastin to shape ER tubules and to regulate LD movement along microtubules, opening new perspectives for the pathogenesis of hereditary spastic paraplegia.

Download Full-text

Truncating mutations of SPAST associated with hereditary spastic paraplegia indicate greater accumulation and toxicity of the M1 isoform of spastin

Molecular Biology of the Cell ◽

10.1091/mbc.e17-01-0047 ◽

2017 ◽

Vol 28 (13) ◽

pp. 1728-1737 ◽

Cited By ~ 12

Author(s):

Joanna M. Solowska ◽

Anand N. Rao ◽

Peter W. Baas

Keyword(s):

Neurite Outgrowth ◽

Hereditary Spastic Paraplegia ◽

Pathogenic Mutation ◽

Full Length ◽

Start Codon ◽

Spastic Paraplegia ◽

Wild Type ◽

Microtubule Severing ◽

Corticospinal Tracts ◽

Over Time

The SPAST gene, which produces two isoforms (M1 and M87) of the microtubule-severing protein spastin, is the chief gene mutated in hereditary spastic paraplegia. Haploinsufficiency is a popular explanation for the disease, in part because most of the >200 pathogenic mutations of the gene are truncating and expected to produce only vanishingly small amounts of shortened proteins. Here we studied two such mutations, N184X and S245X, and our results suggest another possibility. We found that the truncated M1 proteins can accumulate to notably higher levels than their truncated M87 or wild-type counterparts. Reminiscent of our earlier studies on a pathogenic mutation that generates full-length M1 and M87 proteins, truncated M1 was notably more detrimental to neurite outgrowth than truncated M87, and this was true for both N184X and S245X. The greater toxicity and tendency to accumulate suggest that, over time, truncated M1 could damage the corticospinal tracts of human patients. Curiously, the N184X mutation triggers the reinitiation of translation at a third start codon in SPAST, resulting in synthesis of a novel M187 spastin isoform that is able to sever microtubules. Thus microtubule severing may not be as reduced as previously assumed in the case of that mutation.

Download Full-text

Coexistence of Hereditary Spastic Paraplegia Type 4 and Narcolepsy: A Case Report

Case Reports in Neurology ◽

10.1159/000512404 ◽

2021 ◽

pp. 84-91

Author(s):

Takahiro Nagai ◽

Yoko Sunami ◽

Risa Kato ◽

Megumi Sugai ◽

Makoto Takahara ◽

...

Keyword(s):

Cognitive Decline ◽

Hereditary Spastic Paraplegia ◽

Clinical Symptoms ◽

Splice Site Mutation ◽

Motor Symptom ◽

Spastic Paraplegia ◽

Site Mutation ◽

Neuronal Morphogenesis ◽

Microtubule Severing

Spastic paraplegia type 4 (SPG4) is the most common type of hereditary spastic paraplegia (HSP) caused by the mutations in the <i>SPAST</i> gene, which encodes a microtubule-severing protein named spastin. Spastin regulates the number and mobility of microtubules and is essential for axonal outgrowth and neuronal morphogenesis. Herein, we report a patient with SPG4 harboring a novel donor splice site mutation in the <i>SPAST</i> gene (c.1616+1dupG). Although SPG4 usually manifests itself as a pure form of HSP, this patient exhibited a slow progressive cognitive decline and also developed narcolepsy type 2 (narcolepsy without cataplexy) prior to the onset of SPG4. Recently, cognitive decline has attracted attention as a main non-motor symptom of SPG4. However, this is the first reported case of a patient developing both SPG4 and narcolepsy, although it remains unclear whether the manifestation of the two diseases is a coincidence or an association. In this report, we describe the clinical symptoms and genetic background of the patient.

Download Full-text

Recognition of C-terminal amino acids in tubulin by pore loops in Spastin is important for microtubule severing

The Journal of Cell Biology ◽

10.1083/jcb.200610072 ◽

2007 ◽

Vol 176 (7) ◽

pp. 995-1005 ◽

Cited By ~ 102

Author(s):

Susan Roehl White ◽

Katia J. Evans ◽

Jeffrey Lary ◽

James L. Cole ◽

Brett Lauring

Keyword(s):

Amino Acids ◽

Conformational Changes ◽

Hereditary Spastic Paraplegia ◽

Spastic Paraplegia ◽

Central Cavity ◽

Aaa Atpase ◽

Microtubule Severing ◽

Microtubule Binding Domain ◽

Terminal Amino ◽

Form Ring

Spastin, an AAA ATPase mutated in the neurodegenerative disease hereditary spastic paraplegia, severs microtubules. Many other AAA proteins form ring-shaped hexamers and contain pore loops, which project into the ring's central cavity and act as ratchets that pull on target proteins, leading, in some cases, to conformational changes. We show that Spastin assembles into a hexamer and that loops within the central pore recognize C-terminal amino acids of tubulin. Key pore loop amino acids are required for severing, including one altered by a disease-associated mutation. We also show that Spastin contains a second microtubule binding domain that makes a distinct interaction with microtubules and is required for severing. Given that Spastin engages the MT in two places and that both interactions are required for severing, we propose that severing occurs by forces exerted on the C-terminal tail of tubulin, which results in a conformational change in tubulin, which releases it from the polymer.

Download Full-text

Defects in ER–endosome contacts impact lysosome function in hereditary spastic paraplegia

The Journal of Cell Biology ◽

10.1083/jcb.201609033 ◽

2017 ◽

Vol 216 (5) ◽

pp. 1337-1355 ◽

Cited By ~ 75

Author(s):

Rachel Allison ◽

James R. Edgar ◽

Guy Pearson ◽

Tania Rizo ◽

Timothy Newton ◽

...

Keyword(s):

Endoplasmic Reticulum ◽

Stem Cell ◽

Hereditary Spastic Paraplegia ◽

Axonal Degeneration ◽

Contact Process ◽

Primary Neurons ◽

Human Stem Cell ◽

Spastic Paraplegia ◽

Microtubule Severing ◽

Cellular Models

Contacts between endosomes and the endoplasmic reticulum (ER) promote endosomal tubule fission, but the mechanisms involved and consequences of tubule fission failure are incompletely understood. We found that interaction between the microtubule-severing enzyme spastin and the ESCRT protein IST1 at ER–endosome contacts drives endosomal tubule fission. Failure of fission caused defective sorting of mannose 6-phosphate receptor, with consequently disrupted lysosomal enzyme trafficking and abnormal lysosomal morphology, including in mouse primary neurons and human stem cell–derived neurons. Consistent with a role for ER-mediated endosomal tubule fission in lysosome function, similar lysosomal abnormalities were seen in cellular models lacking the WASH complex component strumpellin or the ER morphogen REEP1. Mutations in spastin, strumpellin, or REEP1 cause hereditary spastic paraplegia (HSP), a disease characterized by axonal degeneration. Our results implicate failure of the ER–endosome contact process in axonopathy and suggest that coupling of ER-mediated endosomal tubule fission to lysosome function links different classes of HSP proteins, previously considered functionally distinct, into a unifying pathway for axonal degeneration.

Download Full-text

Therapeutic Strategies for Mutant SPAST-Based Hereditary Spastic Paraplegia

Brain Sciences ◽

10.3390/brainsci11081081 ◽

2021 ◽

Vol 11 (8) ◽

pp. 1081

Author(s):

Neha Mohan ◽

Liang Qiang ◽

Gerardo Morfini ◽

Peter W. Baas

Keyword(s):

Present Article ◽

Hereditary Spastic Paraplegia ◽

Therapeutic Strategies ◽

Spastic Paraplegia ◽

Gain Of Function ◽

Disease Etiology ◽

Microtubule Severing ◽

Pathogenic Mutations ◽

Two Factors

Mutations of the SPAST gene that encodes the microtubule-severing enzyme called spastin are the chief cause of Hereditary Spastic Paraplegia. Growing evidence indicates that pathogenic mutations functionally compromise the spastin protein and endow it with toxic gain-of-function properties. With each of these two factors potentially relevant to disease etiology, the present article discusses possible therapeutic strategies that may ameliorate symptoms in patients suffering from SPAST-based Hereditary Spastic Paraplegia, which is usually termed SPG4-HSP.

Download Full-text

Spastin recovery in hereditary spastic paraplegia by preventing neddylation-dependent degradation

Life Science Alliance ◽

10.26508/lsa.202000799 ◽

2020 ◽

Vol 3 (12) ◽

pp. e202000799

Author(s):

Francesca Sardina ◽

Alessandra Pisciottani ◽

Manuela Ferrara ◽

Davide Valente ◽

Marialuisa Casella ◽

...

Keyword(s):

Hereditary Spastic Paraplegia ◽

Cell Model ◽

Neuronal Cell ◽

Spastic Paraplegia ◽

Therapeutic Approaches ◽

Proliferating Cells ◽

Gene Encoding ◽

Microtubule Severing ◽

Protein Levels

Hereditary Spastic Paraplegia (HSP) is a neurodegenerative disease most commonly caused by autosomal dominant mutations in the SPG4 gene encoding the microtubule-severing protein spastin. We hypothesise that SPG4-HSP is attributable to reduced spastin function because of haploinsufficiency; thus, therapeutic approaches which elevate levels of the wild-type spastin allele may be an effective therapy. However, until now, how spastin levels are regulated is largely unknown. Here, we show that the kinase HIPK2 regulates spastin protein levels in proliferating cells, in differentiated neurons and in vivo. Our work reveals that HIPK2-mediated phosphorylation of spastin at S268 inhibits spastin K48-poly-ubiquitination at K554 and prevents its neddylation-dependent proteasomal degradation. In a spastin RNAi neuronal cell model, overexpression of HIPK2, or inhibition of neddylation, restores spastin levels and rescues neurite defects. Notably, we demonstrate that spastin levels can be restored pharmacologically by inhibiting its neddylation-mediated degradation in neurons derived from a spastin mouse model of HSP and in patient-derived cells, thus revealing novel therapeutic targets for the treatment of SPG4-HSP.

Download Full-text

Different fusion tags affect the activity of ubiquitin overexpression on spastin protein stability

European Journal of Histochemistry ◽

10.4081/ejh.2021.3352 ◽

2021 ◽

Vol 65 (4) ◽

Author(s):

Jianyu Zou ◽

Zhenbin Cai ◽

Zhi Liang ◽

Yaozhong Liang ◽

Guowei Zhang ◽

...

Keyword(s):

Protein Stability ◽

Neurite Outgrowth ◽

Hereditary Spastic Paraplegia ◽

Hippocampal Neurons ◽

Fluorescent Protein ◽

The Novel ◽

Spastic Paraplegia ◽

Promoting Effect ◽

Microtubule Severing ◽

Green Fluorescent

Spastin is one of the proteins which lead to hereditary spastic paraplegia (HSP), whose dysfunction towards microtubule severing and membrane transporting is critically important. The present study is to elucidate the mechanisms of the protein stability regulation of spastin. The ubiquitin encoding plasmids are transfected into COS-7 cells with different fusion tags including Green Fluorescent Protein (GFP), mCherry and Flag. The expression level of spastin was detected, microtubule severing activity and neurite outgrowth were quantified. The data showed that ubiquitin overexpression significantly induced the decreased expression of spastin, suppressed the activity of microtubule severing in COS-7 cells and inhibited the promoting effect on neurite outgrowth in cultured hippocampal neurons. Furthermore, when modulating the overexpression experiments of ubiquitin, it was found that relatively small tag like Flag, but not large tags such as GFP or mCherry fused with ubiquitin, retained the activity on spastin stability. The present study investigated the effects of small/large tags addition to ubiquitin and the novel mechanisms of post-transcriptional modifications of spastin on regulating neurite outgrowth, in the attempt to experimentally elucidate the mechanisms that control the level or stability of spastin in hereditary spastic paraplegia.

Download Full-text