SorCS1-mediated Sorting of Neurexin in Dendrites Maintains Presynaptic Function

Background

Neurons are polarized cells with two distinct compartments: dendrites and axons. These contain the post-and pre-synaptic specialization, respectively, which vary greatly in protein composition. Proper neuronal development, neuronal function and neuronal plasticity depend on the precise targeting of proteins to the right compartment. The bulk of protein synthesis in neurons occurs at the cell body. Thus, a central conundrum in Neuroscience remains: how are axonal and dendritic proteins differentially located to distinct compartments within the same cell?

Neurons have developed different mechanisms to ensure proper axonal protein targeting¹. For instance, when protein subcellular distribution initially happens in a non-discriminatory fashion, protein cargo can be removed exclusively from the dendritic surface through a mechanism called selective endocytosis/retention². As a result, protein cargo becomes enriched at the axonal surface. Alternatively, axonal cargo can be first inserted into the somatodendritic plasma membrane, and only then internalized from the cell surface and rerouted via endosomally derived carriers to the axon through a mechanism called transcytosis².

In this preprint, Ribeiro et al. examined the role of the sorting receptor SorCS1 in the transcytosis-mediated axonal targeting of Neurexin (Nrxn), a presynaptic cell adhesion molecule essential for synaptogenesis and neuronal transmission³. SorCS1 belongs to the family of VPS10P-domain sorting receptors⁴, which are prominently expressed in the brain. They have recently emerged as key regulators of intracellular trafficking of synaptic receptors^5–7 , but the mechanism through which they do this is yet uncharacterized. Here, Ribeiro and colleagues elucidate part of the cellular and molecular mechanisms that SorCS proteins use to regulate the trafficking of synaptic receptors.

 

Key Findings

First, the authors show that the axonal surface of mature cortical primary neurons displays twice as much endogenous Nrxn1a as the dendritic surface, indicating polarization towards the axon (see figure below). However, when the cells are permeabilized, Nrxn1a is mostly detected in dendrites. Permeabilization makes it possible to detect also intracellular proteins, indicating that Nrxn1a is trafficked to both compartments. To understand the mechanism responsible for axonal polarization of Nrxn1α the authors performed live-cell imaging experiments to monitor the transport of Nrxn1a throughout the secretory pathway. They showed that Nrxn1a is first trafficked to dendrites, appearing in the axon only after a long delay. Thus, they hypothesized that newly synthesized Nrxn1a is first trafficked to the somatodendritic compartment, where it is inserted into the plasma membrane. Indeed, when they monitored endocytosis, Nrxn1a was internalized from this membrane and further transcytosed, through the cell, from the dendritic surface to the axonal compartment.

Figure. (a) Surface endogenous Nrxn1a is axonally polarized in mouse cortical neurons. Red arrowheads indicate the axon and blue asterisks mark the cell body. (b) ratio of the axonal/dendritic HA intensity. DIV, days in vitro. Figure adapted from the preprint figure 3 by Ribeiro et al. made available under a CC-BY-NC-ND 4.0 International license.

 

One factor that could be involved in this process is SorCS1, an endosomal sorting receptor that cycles between the plasma membrane and the endosomal pathway⁴. Indeed, as previous work had shown that SorCS1 directly binds Nrxn in cis⁷, the authors tested whether this protein was required for Nrxn1α transcytosis. Loss of SorCS1 resulted in an accumulation of Nrxn1a on the dendritic surface and its depletion from the axonal surface, shifting Nrxn1α surface polarization from axonal to dendritic. This polarization defect was due to an impaired transition of internalized Nrxn1a from early endosomes (EEs) to recycling endosomes (REs) in the somatodendritic compartment. Normally, upon internalization from the dendritic plasma membrane, cargo proteins are incorporated into small carrier vesicles that fuse with existing EEs. From EEs, cargo is sorted to REs. When this transition is affected (like in the case of SorCS1 loss), cargo accumulates in EE, and is either degraded via lysosomes or recycled back to the dendritic membrane.

To gain more mechanistic insight into how SorCS1 functions, the authors set out to find factors that interact with this protein. They identified the endosomal associated protein, Rip11, as a new SorCS1 binding partner. Rip11 has been previously involved in sorting of cargo from early to recycling endosomes in non-neuronal cells, and here they showed that it is also required to maintain axon-dendritic polarity of Nrxn1a.

With these findings in mind, Ribeiro et al. proposed that SorCS1 and Rip11 form a protein complex that localizes to dendritic endosomes and sorts internalized Nrxn1α from early to recycling endosomes. This could explain the increase in surface dendritic levels of Nrxn1α and the concomitant decrease in surface axonal levels observed in Sorcs1 KO cells.

Finally, the authors tested the impact of genetic ablation of SorCS1, which decreases the axonal surface targeting of Nrxn1a, on Nrxn-dependent neuronal transmission and synaptogenesis. Indeed, depletion of SorCS1 compromised Nrxn-dependent synapse assembly and neuronal transmission. In summary, these findings expand the list of neuronal cargoes that undergo transcytosis, which so far is very limited², and support the notion that SorCS1-mediated sorting in dendrites is required to maintain axon-dendritic polarity and normal neuronal function.

 

What do I think about this preprint / Open Questions?

I like this preprint a lot! It is a detailed study, and carefully performed. One experiment I particularly like, and that I think should become more common in cell biology research, is the endogenous tagging of proteins using CRISPR/Cas9. For instance, with this technique, the authors showed that Nrxn1α does not only localize to axons, but could also be found (albeit potentially temporarily) in the dendrites. As such, the authors could overcome the lack of good antibodies, and used these observations as the basis for their study. In addition, the live-cell imaging experiments in this work are very impressive. I thought the videos of cargo trafficking from one cellular compartment to the other in real life were awesome!

However, these observations also raise some questions that, in the future, may need further clarification. For instance, what is the biological function of the dendritic pool of Nrxn? Is dendritic Nrxn merely a reservoir of readily-synthesized protein to be delivered to the axonal membrane, or does dendritically localized Nrxn have a function in the dendritic compartment? As dendritic/postsynaptically expressed Nrxn has been shown to inhibit synapse formation⁸, it would be interesting to further study other possible functions of dendritic Nrxn, particularly of intracellular Nrxn, which is abundantly present in dendrites.

One important finding is that the axonal-dendritic balance of Nrxn is regulated by neuronal activity. Does this mean that SorCS1-mediated sorting can dynamically regulate Nrxn polarity? How do neuronal activity and SorCS1-mediated sorting of Nrxn crosstalk in order to ensure proper protein distribution and neuronal functioning? And further, how much can these Nrxn-specific observations be generalized to explain neuronal activity-dependent compartment diversification?

In summary, I think this study provides important insights into the mechanisms controlling the intracellular trafficking of a compartmentally polarized protein, which helps to better understand subcellular protein distribution in polarized cells. With the toolbox the authors developed for this study, I am convinced many more questions can be answered, and thus I look forward to read the follow up stories, too!

 

References

1. Bentley, M. & Banker, G. Nat. Rev. Neurosci. 17, 611–622 (2016).
2. Winckler, B. & Mellman, I. Cold Spring Harb. Perspect. Biol. 2, a001826–a001826 (2010).
3. Südhof, T.C. Cell 171, 745–769 (2017).
4. Willnow, T.E., Petersen, C.M. & Nykjaer, A. Nat. Rev. Neurosci. 9, 899–909 (2008).
5. Ma, Q. et al. JCI Insight 2, (2017).
6. Glerup, S. et al. Mol. Psychiatry 21, 1740–1751 (2016).
7. Savas, J.N. et al. Neuron 87, 764–780 (2015).
8. Taniguchi, H. et al. J. Neurosci. 27, 2815–2824 (2007).

 

 

Tags: axonal targeting, cell biology, cell compartment specialization, intracellular trafficking, neurexins, neuroscience, sorcs1, transcytosis

doi: https://doi.org/10.1242/prelights.8991

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