Non-SUMOylated CRMP2 decreases NaV1.7 currents via the endocytic proteins Numb, Nedd4-2 and Eps15

Kimberly Gomez,Cynthia L Madura,Rajesh Khanna,Dongzhi Ran,Aubin Moutal

doi:10.1186/s13041-020-00714-1

Kimberly Gomez, Cynthia L Madura + Show 3 more

Open Access

https://doi.org/10.1186/s13041-020-00714-1

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Journal: Molecular brain	Publication Date: Jan 21, 2021
Citations: 17	License type: open-access

Affiliation: University of Arizona

Abstract

Voltage-gated sodium channels are key players in neuronal excitability and pain signaling. Functional expression of the voltage-gated sodium channel NaV1.7 is under the control of SUMOylated collapsin response mediator protein 2 (CRMP2). When not SUMOylated, CRMP2 forms a complex with the endocytic proteins Numb, the epidermal growth factor receptor pathway substrate 15 (Eps15), and the E3 ubiquitin ligase Nedd4-2 to promote clathrin-mediated endocytosis of NaV1.7. We recently reported that CRMP2 SUMO-null knock-in (CRMP2K374A/K374A) female mice have reduced NaV1.7 membrane localization and currents in their sensory neurons. Preventing CRMP2 SUMOylation was sufficient to reverse mechanical allodynia in CRMP2K374A/K374A female mice with neuropathic pain. Here we report that inhibiting clathrin assembly in nerve-injured male CRMP2K374A/K374A mice precipitated mechanical allodynia in mice otherwise resistant to developing persistent pain. Furthermore, Numb, Nedd4-2 and Eps15 expression was not modified in basal conditions in the dorsal root ganglia (DRG) of male and female CRMP2K374A/K374A mice. Finally, silencing these proteins in DRG neurons from female CRMP2K374A/K374A mice, restored the loss of sodium currents. Our study shows that the endocytic complex composed of Numb, Nedd4-2 and Eps15, is necessary for non-SUMOylated CRMP2-mediated internalization of sodium channels in vivo.

Highlights

NaV1.7 is a voltage-gated sodium channel highly expressed in nociceptive neurons and in the dorsal horn of the spinal cord [1]
Of relevance to the role of N aV1.7 in pain, we found that C RMP2K374A/K374A mice of both sexes failed to develop mechanical allodynia after a spinal nerve injury (SNI) [37], supporting the conclusion that collapsin response mediator protein 2 (CRMP2) SUMOylation-dependent regulation of NaV1.7 still holds in chronic neuropathic pain in male mice
In male CRMP2K374A/K374A mice, we found higher paw withdrawal thresholds at day 42 in mice subjected to SNI compared to their WT littermates (Fig. 1; raw quantified numerical data for this and all subsequent figures is shown in Additional file 1: Table S1)

Summary

Introduction

NaV1.7 is a voltage-gated sodium channel highly expressed in nociceptive neurons and in the dorsal horn of the spinal cord [1]. Loss of CRMP2 SUMOylation results in: (i) reduced NaV1.7–CRMP2 binding; (ii) increased N aV1.7 internalization via association and recruitment of a tripartite complex of proteins containing the endocytic protein Numb, the E3 ubiquitin ligase, the neuronal precursor cell expressed developmentally downregulated-4 type 2 (Nedd4-2), and epidermal growth factor receptor pathway substrate 15 (Eps15); and (iii) decreased NaV1.7 surface expression and currents [25]. This reduction can be rescued by blocking clathrin mediated endocytosis or by deleting the endocytic proteins Numb, Nedd or Eps15 [25]. Eps binds to the monoubiquitinated channel to induce the initial curvature of the membrane and allow for the clathrin coated pit to form [34, 35]

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