TY - JOUR
T1 - CRMP2 protein SUMOylation modulates NaV1.7 channel trafficking
AU - Dustrude, Erik T.
AU - Wilson, Sarah M.
AU - Ju, Weina
AU - Xiao, Yucheng
AU - Khanna, Rajesh
PY - 2013/8/23
Y1 - 2013/8/23
N2 - Background: Post-translational modifications of CRMP2 protein direct its regulation of effector proteins. Results: Destruction of a CRMP2 SUMOylation site reduces surface expression and current density of sodium channel NaV1.7. Conclusion: CRMP2 SUMOylation choreographs NaV1.7, but not NaV1.1 or NaV1.3, trafficking. Significance: Learning how neuronal NaV1.7 trafficking is modulated by CRMP2 is important for understanding the mechanism of action of NaV-targeted anti-epileptic and anti-nociceptive drugs. Voltage-gated sodium channel (NaV) trafficking is incompletely understood. Post-translational modifications of NaVs and/or auxiliary subunits and protein-protein interactions have been posited as NaV-trafficking mechanisms. Here, we tested if modification of the axonal collapsin response mediator protein 2 (CRMP2) by a small ubiquitin-like modifier (SUMO) could affect NaV trafficking; CRMP2 alters the extent of NaV slow inactivation conferred by the anti-epileptic (R)-lacosamide, implying NaV-CRMP2 functional coupling. Expression of a CRMP2 SUMOylation-incompetent mutant (CRMP2-K374A) in neuronal model catecholamine A differentiated (CAD) cells did not alter lacosamide-induced NaV slow inactivation compared with CAD cells expressing wild type CRMP2. Like wild type CRMP2, CRMP2-K374A expressed robustly in CAD cells. Neurite outgrowth, a canonical CRMP2 function, was moderately reduced by the mutation but was still significantly higher than enhanced GFP-transfected cortical neurons. Notably, huwentoxin-IV-sensitive NaV1.7 currents, which predominate inCADcells, were significantly reduced inCADcells expressing CRMP2-K374A. Increasing deSUMOylation with sentrin/SUMO-specific protease SENP1 or SENP2 in wild type CRMP2-expressing CAD cells decreased NaV1.7 currents. Consistent with a reduction in current density, biotinylation revealed a significant reduction in surface NaV1.7 levels in CAD cells expressing CRMP2-K374A; surface NaV1.7 expression was also decreased by SENP1 + SENP2 overexpression. Currents in HEK293 cells stably expressing NaV1.7 were reduced by CRMP2-K374A in a manner dependent on the E2-conjugating enzyme Ubc9. No decrement in current density was observed in HEK293 cells co-expressing CRMP2-K374A and NaV1.1 or NaV1.3. Diminution of sodium currents, largely NaV1.7, was recapitulated in sensory neurons expressing CRMP2-K374A. Our study elucidates a novel regulatory mechanism that utilizes CRMP2 SUMOylation to choreograph NaV1.7 trafficking.
AB - Background: Post-translational modifications of CRMP2 protein direct its regulation of effector proteins. Results: Destruction of a CRMP2 SUMOylation site reduces surface expression and current density of sodium channel NaV1.7. Conclusion: CRMP2 SUMOylation choreographs NaV1.7, but not NaV1.1 or NaV1.3, trafficking. Significance: Learning how neuronal NaV1.7 trafficking is modulated by CRMP2 is important for understanding the mechanism of action of NaV-targeted anti-epileptic and anti-nociceptive drugs. Voltage-gated sodium channel (NaV) trafficking is incompletely understood. Post-translational modifications of NaVs and/or auxiliary subunits and protein-protein interactions have been posited as NaV-trafficking mechanisms. Here, we tested if modification of the axonal collapsin response mediator protein 2 (CRMP2) by a small ubiquitin-like modifier (SUMO) could affect NaV trafficking; CRMP2 alters the extent of NaV slow inactivation conferred by the anti-epileptic (R)-lacosamide, implying NaV-CRMP2 functional coupling. Expression of a CRMP2 SUMOylation-incompetent mutant (CRMP2-K374A) in neuronal model catecholamine A differentiated (CAD) cells did not alter lacosamide-induced NaV slow inactivation compared with CAD cells expressing wild type CRMP2. Like wild type CRMP2, CRMP2-K374A expressed robustly in CAD cells. Neurite outgrowth, a canonical CRMP2 function, was moderately reduced by the mutation but was still significantly higher than enhanced GFP-transfected cortical neurons. Notably, huwentoxin-IV-sensitive NaV1.7 currents, which predominate inCADcells, were significantly reduced inCADcells expressing CRMP2-K374A. Increasing deSUMOylation with sentrin/SUMO-specific protease SENP1 or SENP2 in wild type CRMP2-expressing CAD cells decreased NaV1.7 currents. Consistent with a reduction in current density, biotinylation revealed a significant reduction in surface NaV1.7 levels in CAD cells expressing CRMP2-K374A; surface NaV1.7 expression was also decreased by SENP1 + SENP2 overexpression. Currents in HEK293 cells stably expressing NaV1.7 were reduced by CRMP2-K374A in a manner dependent on the E2-conjugating enzyme Ubc9. No decrement in current density was observed in HEK293 cells co-expressing CRMP2-K374A and NaV1.1 or NaV1.3. Diminution of sodium currents, largely NaV1.7, was recapitulated in sensory neurons expressing CRMP2-K374A. Our study elucidates a novel regulatory mechanism that utilizes CRMP2 SUMOylation to choreograph NaV1.7 trafficking.
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U2 - 10.1074/jbc.M113.474924
DO - 10.1074/jbc.M113.474924
M3 - Article
C2 - 23836888
AN - SCOPUS:84883176930
SN - 0021-9258
VL - 288
SP - 24316
EP - 24331
JO - Journal of Biological Chemistry
JF - Journal of Biological Chemistry
IS - 34
ER -