Confocal micrograph of a Drosophila melanogaster neuromuscular junction. Signals represent immunoreactivity for different antibodies. Red, anti-horseradish peroxidase labelling neuronal membranes; green, anti-D-spastin; blue, anti-acetylated tubulin, which detects stable and long-lived microtubule filaments. Note that regions where D-spastin is enriched also appear to be regions where stable microtubules are excluded. Image courtesy of K. Broadie, Vanderbilt University, USA.

Hereditary spastic paraplegia is a devastating motor disorder that causes spastic weakness of the lower limbs and eventual axonal degeneration. More than 40% of all cases are associated with mutations in one gene, spastin, but little is known about how such mutations cause the disease. Reporting in Current Biology, Trotta et al. found that spastin mutations might lead to defects in neurotransmission by affecting microtubule functions.

Spastin is an ATPase that contains a microtubule-interacting domain. Therefore, the authors asked whether microtubule structure or function might be affected by abnormal spastin expression. They studied this in the Drosophila melanogaster neuromuscular junction (NMJ) synapse, which has been widely used to assay functions of the microtubule cytoskeleton and to model its role in inherited neurological diseases.

Spastin is expressed at high levels in the nervous system of both mammals and D. melanogaster, but its subcellular localization has not been characterised. Trotta et al. found that the protein is highly enriched in axons and synaptic connections. They showed that spastin co-localizes with a synaptic vesicle protein, synaptotagmin, indicating that spastin is expressed in the synaptic vesicle pool domain of the presynaptic bouton.

The authors showed that knockdown of ubiquitous spastin expression by RNA interference causes lethality. When they knocked down spastin expression specifically in the nervous system, the animals had very poor coordination and locomotor abilities. Interestingly, overexpression of spastin, either ubiquitously or specifically in the nervous system, was also lethal to embryos or early larvae. The authors suggest that a correct dose of spastin expression is crucial for normal development.

Spastin interacts with microtubules and prevents their assembly in vitro. The authors assessed the effects of altered spastin expression in vivo on microtubule assembly and synaptic transmission. They found that, at the NMJ presynaptic terminal, spastin knockdown in the neuron led to an accumulation of acetylated α-tubulin, the post-translationally modified form of tubulin that occurs only in structurally stable microtubules. Conversely, neuron-specific overexpression of spastin caused a reduction in stabilized tubulin and, often, the stabilized tubulin network was no longer detectable.

The authors showed that these effects on microtubule assembly correlated with changes in synaptic transmission. When they stimulated the motor nerve and measured glutamate-gated synaptic currents in the voltage-clamped muscle, they found that loss of spastin expression resulted in an increase in current amplitude, whereas spastin overexpression had the opposite effect. These effects could be reversed by pharmacological agents that affect microtubule stability. Normal functions were restored in spastin knockdown flies by nocodazole, which disassembles microtubules, and in flies overexpressing spastin by taxol, which stabilizes tubulin monomers. In both cases synaptic transmission was indistinguishable from that in normal animals.

The study shows that spastin is enriched at the synapse and controls synaptic transmission by regulating microtubule assembly. Trotta et al. conclude that it is likely that defects in microtubule stability are the primary cause of hereditary spastic paraplegia. This mechanistic insight has significant implications in designing therapeutic strategies to treat the illness.