VAP spatially stabilizes dendritic mitochondria to locally fuel synaptic plasticity

Background:

Local information processing in neuronal synapses is energy-intensive and requires a continuous supply of ATP for incurring long-term changes at the synapses¹. Making up only 2% of our total body weight, a human brain utilizes about 20% of the body’s total oxygen for its optimal functioning^1,2. As shown previously, inhibiting mitochondrial activity blocks protein synthesis-dependent synaptic plasticity³. This observation means that a stable mitochondrial compartment is required at the dendrites to fuel local changes at the synapses in response to synaptic activity.

While the players involved in docking the mitochondria in axonal terminals have been identified and well characterized, the mechanistic basis of mitochondrial tethering and stabilization within neuronal dendrites remains unknown. To address this knowledge gap, the authors of this preprint screened for proteins that exist close to mitochondria and interact with actin and the microtubule cytoskeleton.

Using a knockdown and imaging approach in hippocampal neurons, the authors were able to hone in on the role of the VAP protein – already known to be involved in several neurological diseases⁴. The authors discovered that the VAPB paralog is particularly localized only to the dendritic compartments. Depletion of VAP from neurons affected the structural plasticity in LTP-induced spines. These findings point to VAP being a major mitochondrial stabilizer near synapses.

Key findings:

To identify the major players in tethering dendritic mitochondria, the authors employed a screening strategy using the APEX tag, which can biotinylate proteins in its vicinity. The APEX tag was targeted to the mitochondrial outer membrane (OMM) and expressed in hippocampal neurons. The captured proteins were identified by mass spectrometry, and the resulting list was filtered for proteins with known interactions with actin and the microtubule cytoskeleton. Eventually, the authors narrowed down to a list of 8 proteins that could be potential players in regulating mitochondrial stability in the dendritic compartments and hence fuel local synaptic plasticity events.

To understand the role of the 8 identified proteins in ensuring dendritic mitochondrial stability and synaptic plasticity, the authors knocked down each of these proteins in hippocampal neurons and measured the actin-mitochondrial interaction in dendrites using Fis1-mCherry and Fis1-Lifeact-GFP. In all knockdown neurons, actin-mitochondrial interaction was significantly reduced in dendrites while unaffected in axons, which suggested a role of these proteins in mitochondrial tethering exclusively to dendritic actin.

To further understand the mechanism by which mitochondrial compartments are stabilized in the dendrites, the authors narrowed their list to three proteins: SNCA, SRGAP2, and VAP. They monitored mitochondrial stability when these proteins were knocked down in hippocampal neurons using photoactivated fluorescence of mitochondria. Knockdown of all three proteins reduced the mitochondrial length at the dendrites. However, of all three knockdowns, only dendrites with VAP knockdown showed a significant decrease in photoactivated mitochondrial compartment fluorescence corresponding to destabilized mitochondrial compartments.

Figure 1 (original fig 6A-B) measuring spine-head width and GCaMP fluorescence before and after synaptic plasticity induction in control and VAP KO hippocampal neurons.

Figure 2 (original fig 6F) Schematic showing VAP as a mitochondrial stabilizer within dendrites supporting synaptic plasticity.

Since VAP is associated with various neurological diseases, the authors then aimed to identify the localization of both paralogs of VAP, VAPA and VAPB, with respect to mitochondria within dendrites and axons. Interestingly, VAPB was specifically enriched near dendritic mitochondria, while VAPA was enriched near dendritic and axonal mitochondria. The specific enrichment of VAPB near dendritic mitochondria made the authors wonder whether VAP protein’s paralog B could be a major stabilizer of dendritic mitochondria and locally support synaptic plasticity. To test this, the authors induced single-spine plasticity using two-photon glutamate uncaging. Spine size was measured in hippocampal neurons with and without VAP knockdown. Interestingly, VAP depletion did not affect the early stages of synaptic plasticity (t=12 mins). However, at later time points (t=22 mins to t=62 mins), spines lacking VAP were unable to retain structural plasticity (increased spine size) (figure 1). Additionally, the ability to exhibit clustered plasticity was also lost in VAP-depleted spines. The authors finally demonstrate a model where VAP tethers and stabilizes dendritic mitochondria, and this spatial stability enables continuous ATP supply via oxidative phosphorylation during synaptic plasticity (see figure 2 above).

What I liked about the preprint:

How a neuron’s intrinsic machinery meets the extensive energy requirements in different compartments is intriguing to me. It is also interesting how different mechanisms exist for mitochondrial stabilization in different neuronal compartments, as loss of each could have diverse but detrimental effects on brain functioning and disease progression. This preprint reflects a significant advancement in our understanding by identifying a critical player facilitating the crucial ATP supply required for long-term synaptic plasticity.

The authors’ approach to identifying new, relevant proteins is impressive to me as this strategy allowed them to screen in a more physiologically relevant model.

Questions to authors:

1. It is my understanding that VAP brings mitochondria and dendritic actin together. Has domain specificity of VAP for actin and the ER been identified? If yes, is the remaining portion of the VAP protein sufficient for mitochondrial tethering?
2. Could VAP-associated mitochondria have enhanced stability via the preclusion of the fission-fusion machinery?
3. As a mitochondrial stabilizer, could VAP be more enriched in active dendrites in comparison to inactive dendrites?

Future Directions:

Studies in the future should focus on validating the role of VAP in stabilizing dendritic mitochondria. Since VAP is enriched in dendrites, it will be interesting to see if only VAP-positive dendritic segments carry mitochondria or if mitochondrial tethering only happens after synaptic activity. Other relevant proteins identified in this preprint should also be explored further for their contribution to dendritic mitochondrial tethering and stabilization.

References:

1. Mink, J. W., Blumenschine, R. J. & Adams, D. B. Ratio of central nervous system to body metabolism in vertebrates: its constancy and functional basis. American Journal of Physiology-Regulatory, Integrative and Comparative Physiology 241, R203–R212 (1981).
2. Harris, J. J., Jolivet, R. & Attwell, D. Synaptic Energy Use and Supply. Neuron 75, 762–777 (2012).
3. Rangaraju, V., Lauterbach, M. & Schuman, E. M. Spatially Stable Mitochondrial Compartments Fuel Local Translation during Plasticity. Cell 176, 73-84.e15 (2019).
4. Nishimura, A. L. et al. A Mutation in the Vesicle-Trafficking Protein VAPB Causes Late-Onset Spinal Muscular Atrophy and Amyotrophic Lateral Sclerosis. The American Journal of Human Genetics 75, 822–831 (2004).

 

doi: https://doi.org/10.1242/prelights.33633

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