PUBLICATION

Targeting the Microtubule EB1-CLASP2 Complex Modulates NaV1.5 at Intercalated Discs

Authors
Marchal, G.A., Jouni, M., Chiang, D.Y., Pérez-Hernández Duran, M., Podliesna, S., Yu, N., Casini, S., Potet, F., Veerman, C.C., Klerk, M., Lodder, E.M., Mengarelli, I., Guan, K., Vanoye, C.G., Rothenberg, E., Charpentier, F., Redon, R., George, A., Verkerk, A.O., Bezzina, C.R., MacRae, C.A., Burridge, P., Delmar, M., Galjart, N.J., Portero, V., Remme, C.A.
ID
ZDB-PUB-210608-12
Date
2021
Source
Circulation research   129(3): 349-365 (Journal)
Registered Authors
Keywords
NaV1.5, intercalated discs, microtubule, trafficking
MeSH Terms
  • Action Potentials
  • Animals
  • Arrhythmias, Cardiac/genetics*
  • Arrhythmias, Cardiac/metabolism
  • Cells, Cultured
  • Glycogen Synthase Kinase 3 beta/metabolism
  • HEK293 Cells
  • Humans
  • Loss of Function Mutation
  • Male
  • Mice
  • Mice, Inbred C57BL
  • Microtubule-Associated Proteins/genetics
  • Microtubule-Associated Proteins/metabolism*
  • Myocytes, Cardiac/metabolism*
  • Myocytes, Cardiac/physiology
  • NAV1.5 Voltage-Gated Sodium Channel/genetics
  • NAV1.5 Voltage-Gated Sodium Channel/metabolism*
  • Protein Transport
  • Zebrafish
(all 20)
PubMed
34092082 Full text @ Circ. Res.
Abstract
Rationale: Loss-of-function of the cardiac sodium channel NaV1.5 causes conduction slowing and arrhythmias. NaV1.5 is differentially distributed within subcellular domains of cardiomyocytes, with sodium current (INa) being enriched at the intercalated discs (ID). Various pathophysiological conditions associated with lethal arrhythmias display ID-specific INa reduction, but the mechanisms underlying microdomain-specific targeting of NaV1.5 remain largely unknown. Objective: To investigate the role of the microtubule (MT) plus-end tracking proteins end binding protein 1 (EB1) and CLIP-associated protein 2 (CLASP2) in mediating NaV1.5 trafficking and subcellular distribution in cardiomyocytes.Methods and Results: EB1 overexpression in human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) resulted in enhanced whole-cell INa, increased action potential (AP) upstroke velocity (Vmax), and enhanced NaV1.5 localization at the plasma membrane as detected by multi-color stochastic optical reconstruction microscopy (STORM). Fluorescence recovery after photobleaching (FRAP) experiments in HEK293A cells demonstrated that EB1 overexpression promoted NaV1.5 forward trafficking. Knockout of MAPRE1 in hiPSC-CMs led to reduced whole-cell INa, decreased Vmax and AP duration (APD) prolongation. Similarly, acute knockout of the MAPRE1 homolog in zebrafish (mapre1b) resulted in decreased ventricular conduction velocity and Vmax as well as increased APD. STORM imaging and macropatch INa measurements showed that subacute treatment (2-3 hours) with SB216763 (SB2), a GSK3β inhibitor known to modulate CLASP2-EB1 interaction, reduced GSK3β localization and increased NaV1.5 and INa preferentially at the ID region of wild type murine ventricular cardiomyocytes. By contrast, SB2 did not affect whole cell INa or NaV1.5 localization in cardiomyocytes from Clasp2-deficient mice, uncovering the crucial role of CLASP2 in SB2-mediated modulation of NaV1.5 at the ID. Conclusions: Our findings demonstrate the modulatory effect of the MT plus-end tracking protein EB1 on NaV1.5 trafficking and function, and identify the EB1-CLASP2 complex as a target for preferential modulation of INa within the ID region of cardiomyocytes.
Genes / Markers
Marker Marker Type Name
cacna1cGENEcalcium channel, voltage-dependent, L type, alpha 1C subunit
gja1bGENEgap junction protein alpha 1b
kcnh6aGENEpotassium voltage-gated channel, subfamily H (eag-related), member 6a
kcnq1.1GENEpotassium voltage-gated channel, KQT-like subfamily, member 1.1
mapre1aGENEmicrotubule-associated protein, RP/EB family, member 1a
mapre1bGENEmicrotubule-associated protein, RP/EB family, member 1b
mapre2GENEmicrotubule-associated protein, RP/EB family, member 2
mapre3aGENEmicrotubule-associated protein, RP/EB family, member 3a
mapre3bGENEmicrotubule-associated protein, RP/EB family, member 3b
scn12aaGENEsodium channel, voltage gated, type XII, alpha a
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Figures
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Expression
No data available
Phenotype
Fish Conditions Stage Phenotype Figure
AB/TU + CRISPR1-mapre1b + CRISPR2-mapre1b + CRISPR3-mapre1bstandard conditionsProtruding-mouth
AB/TU + CRISPR1-mapre1b + CRISPR2-mapre1b + CRISPR3-mapre1bstandard conditionsProtruding-mouth
1 - 2 of 2
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Mutations / Transgenics
No data available
Human Disease / Model
No data available
Sequence Targeting Reagents
Target Reagent Reagent Type
mapre1bCRISPR1-mapre1bCRISPR
mapre1bCRISPR2-mapre1bCRISPR
mapre1bCRISPR3-mapre1bCRISPR
1 - 3 of 3
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Fish
Fish
AB/TU + CRISPR1-mapre1b + CRISPR2-mapre1b + CRISPR3-mapre1b
1 - 1 of 1
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Antibodies
Orthology
Engineered Foreign Genes
No data available
Mapping
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Citation
Marchal, G.A., Jouni, M., Chiang, D.Y., Pérez-Hernández Duran, M., Podliesna, S., Yu, N., Casini, S., Potet, F., Veerman, C.C., Klerk, M., Lodder, E.M., Mengarelli, I., Guan, K., Vanoye, C.G., Rothenberg, E., Charpentier, F., Redon, R., George, A., Verkerk, A.O., Bezzina, C.R., MacRae, C.A., Burridge, P., Delmar, M., Galjart, N.J., Portero, V., Remme, C.A. (2021) Targeting the Microtubule EB1-CLASP2 Complex Modulates NaV1.5 at Intercalated Discs. Circulation research. 129(3):349-365.