You can’t keep a good mitochondrion down. Or maybe you can. In today’s Cell, researchers led by Zu-Hang Sheng at the National Institute for Neurological Disorders and Stroke report that the protein syntaphilin docks mitochondria to microtubules, impeding their transport and preventing their delivery to synapses. Genetically ablating syntaphilin boosts the mobility of mitochondria and enhances short-term facilitation, suggesting that syntaphilin may play a role in regulating neural activity, the scientists report. The finding may help researchers understand the relationship between defective mitochondrial transport and neurological and neurodegenerative disorders.
Being energy hogs, neurons are packed with mitochondria, particularly their synapses. Sheng and colleagues previously showed that syntaphilin can suppress presynaptic function (see Lao et al., 2000), which prompted them to go after the function of the protein. Using immunohistochemistry, first author Jian-Sheng Kang and colleagues found that syntaphilin (SNPH) occurs mostly in axons. Furthermore, using cultured hippocampal neurons, they found that the majority of SNPH colocalized with axonal mitochondria, and that deleting an axon-sorting domain redistributed SNPH into mitochondria in various other cellular locales, including the soma and dendrites. The finding suggests a specific relationship between wild-type syntaphilin and axonal mitochondria.
To test if that relationship might affect mitochondrial transport, Kang and colleagues observed the organelles with time-lapse confocal microscopy. They labeled mitochondria with red fluorescent protein (DsRed) and syntaphilin with green fluorescent protein (GFP). The researchers found that while 38 percent of axonal mitochondria are normally on the move, mitochondria bound to GFP-SNPH were stationary. It appears that SNPH binding to mitochondria locks them to microtubules, since SNPH lacking its microtubule binding domain did not impede mitochondrial motion. In fact, by simply tacking syntaphilin’s microtubule binding domain to the mitochondrial outer membrane protein TOM22, the researchers were able to block transport of the organelles. Interestingly, the syntaphilin microtubule binding domain is unlike any other known microtubule binding protein, suggesting that “SNPH likely acts as a docking receptor specific for axonal mitochondria through a unique interaction with the microtubule-based cytoskeleton,” write the authors.
Why does the SNPH-mitochondrial interaction matter? To address this, the authors made SNPH knockout mice. Both homo- and heterozygous strains were viable and fertile. Though their overall appearance was similar to wild-type mice, they did have subtle impairments in motor ability. For example, while the SNPH nulls balanced fine on the rotorod at constant speed, they faltered sooner than wild-type mice when the rod accelerated. As for axonal mitochondria, substantially more (76 percent) of them were on the move in hippocampal neurons cultured from SNPH knockouts than in neurons from wild-type mice (36 percent).
What might happen to the neuron when its mitochondria move more? For one, the density of axonal mitochondria drops (~ 1 per 10 μm in SNPH KOs versus 1.7 in wild-type); for another, the ratio of anterograde to retrograde mitochondrial movement changes from 1.27 to 1.03. These two changes “suggest that the SNPH-mediated docking/retention contributes to the maintenance of proper mitochondrial density in axons,” write the authors. Synapses also appear to be affected. The proportion of presynaptic boutons with proximal mitochondria within 0.5 μm goes down from 57 percent to 33 percent in SNPH knockouts, though the density of boutons stays normal. Given the primary role of mitochondria as energy providers, their failure to dock in and around synapses could deprive some of much needed ATP. The authors suggest this might be particularly significant for neurons with long axonal processes such as motor neurons.
Could the redistribution of mitochondria in SNPH null animals affect synaptic transmission? To test this, the authors measured excitatory post-synaptic currents (EPSC) in cultured hippocampal neurons. They found that while basal activity was normal in SNPH-negative neurons, a train of high-frequency pulses induced enhanced currents that indicate short-term facilitation. “One possible explanation for this observation is a rapid buildup of intracellular [Ca2+] in presynaptic terminals during intensive stimulation,” suggest the authors. In support of this, they demonstrated increased Ca2+ using fluorescent imaging and were able to abolish the enhanced short-term facilitation phenotype using the calcium chelator EGTA-AM. This would be in keeping with a proposed role for mitochondria in calcium homeostasis: mitochondria help balance calcium at some synapses by mopping up Ca2+ released by tetanic stimulation (see Jonas et al., 2006).
Though Sheng and colleagues previously reported that syntaphilin is involved in synaptic endocytosis (see Das et al., 2003), these findings also support a role of syntaphilin in docking mitochondria to microtubules and regulating their motility. “Such a mechanism may enable neurons to maintain proper densities of stationary mitochondria within axons and in the proximity of synapses," write the authors. Given that defective mitochondrial transport has been implicated in Charcot-Marie-Tooth Disease (see Baloh et al., 2007), Huntington disease (see Trushina et al., 2004), and potentially other neurodegenerative disorders (see for review Stokin and Goldstein, 2006), the findings may help researchers more fully understand the role of mitochondrial transport in neurons and neurologic disorders.—Tom Fagan
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No Available Further Reading
- Kang JS, Tian JH, Pan PY, Zald P, Li C, Deng C, Sheng ZH. Docking of axonal mitochondria by syntaphilin controls their mobility and affects short-term facilitation. Cell. 2008 Jan 11;132(1):137-48. PubMed.