Neuronal junctophilins recruit specific CaV and RyR isoforms to ER-PM junctions and functionally alter CaV2.1 and CaV2.2

  1. Stefano Perni
  2. Kurt Beam  Is a corresponding author
  1. University of Colorado Anschutz Medical Campus, United States


Despite their recognized physiological importance, the molecular architecture of ER-PM junctions induced by neuronal junctophilins (JPH3 and JPH4) is still poorly understood and challenging to address in neurons. This is due to the small size of the junctions and to the multiple isoforms of candidate junctional proteins in different brain areas. Using colocalization of tagged proteins expressed in tsA201 cells, and electrophysiology, we compared the interactions of JPH3 and JPH4 with different calcium channels. We found that JPH3 and JPH4 caused junctional accumulation of all the tested high-voltage-activated CaV isoforms, but not a low-voltage-activated CaV. Also, JPH3 and JPH4 noticeably modify CaV2.1 and CaV2.2 inactivation rate. RyR3 moderately colocalized at junctions with JPH4, whereas RyR1 and RyR2 did not. By contrast, RyR1 and RyR3 strongly colocalized with JPH3, and RyR2 moderately. Likely contributing to this difference, JPH3 binds to cytoplasmic domain constructs of RyR1 and RyR3, but not of RyR2.

Data availability

Raw data for peak current vs voltage, inactivation vs voltage, and Pearson's coefficients have been provided with the uploaded manuscript files.

Article and author information

Author details

  1. Stefano Perni

    Physiology & Biophysics, University of Colorado Anschutz Medical Campus, Aurora, United States
    Competing interests
    No competing interests declared.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-0591-4376
  2. Kurt Beam

    Physiology & Biophysics, University of Colorado Anschutz Medical Campus, Aurora, United States
    For correspondence
    Competing interests
    Kurt Beam, Reviewing editor, eLife.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0001-6902-085X


NIH Office of the Director (R01 AR070298)

  • Kurt Beam

The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.

Reviewing Editor

  1. Mark T Nelson, University of Vermont, United States

Publication history

  1. Received: October 22, 2020
  2. Accepted: March 19, 2021
  3. Accepted Manuscript published: March 26, 2021 (version 1)
  4. Version of Record published: April 14, 2021 (version 2)


© 2021, Perni & Beam

This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.


  • 1,064
    Page views
  • 207
  • 7

Article citation count generated by polling the highest count across the following sources: Crossref, PubMed Central, Scopus.

Download links

A two-part list of links to download the article, or parts of the article, in various formats.

Downloads (link to download the article as PDF)

Open citations (links to open the citations from this article in various online reference manager services)

Cite this article (links to download the citations from this article in formats compatible with various reference manager tools)

  1. Stefano Perni
  2. Kurt Beam
Neuronal junctophilins recruit specific CaV and RyR isoforms to ER-PM junctions and functionally alter CaV2.1 and CaV2.2
eLife 10:e64249.

Further reading

    1. Cell Biology
    2. Developmental Biology
    Juan Lu et al.
    Research Article Updated

    Phosphatidylinositol 4-phosphate (PI4P) and phosphatidylinositol 4,5-biphosphate (PIP2) are key phosphoinositides that determine the identity of the plasma membrane (PM) and regulate numerous key biological events there. To date, mechanisms regulating the homeostasis and dynamic turnover of PM PI4P and PIP2 in response to various physiological conditions and stresses remain to be fully elucidated. Here, we report that hypoxia in Drosophila induces acute and reversible depletion of PM PI4P and PIP2 that severely disrupts the electrostatic PM targeting of multiple polybasic polarity proteins. Genetically encoded ATP sensors confirmed that hypoxia induces acute and reversible reduction of cellular ATP levels which showed a strong real-time correlation with the levels of PM PI4P and PIP2 in cultured cells. By combining genetic manipulations with quantitative imaging assays we showed that PI4KIIIα, as well as Rbo/EFR3 and TTC7 that are essential for targeting PI4KIIIα to PM, are required for maintaining the homeostasis and dynamic turnover of PM PI4P and PIP2 under normoxia and hypoxia. Our results revealed that in cells challenged by energetic stresses triggered by hypoxia, ATP inhibition and possibly ischemia, dramatic turnover of PM PI4P and PIP2 could have profound impact on many cellular processes including electrostatic PM targeting of numerous polybasic proteins.

    1. Cell Biology
    2. Medicine
    Eric N Jimenez-Vazquez et al.
    Research Article


    Patients with cardiomyopathy of Duchenne Muscular Dystrophy (DMD) are at risk of developing life-threatening arrhythmias, but the mechanisms are unknown. We aimed to determine the role of ion channels controlling cardiac excitability in the mechanisms of arrhythmias in DMD patients.


    To test whether dystrophin mutations lead to defective cardiac NaV1.5–Kir2.1 channelosomes and arrhythmias, we generated iPSC-CMs from two hemizygous DMD males, a heterozygous female, and two unrelated control males. We conducted studies including confocal microscopy, protein expression analysis, patch-clamping, non-viral piggy-bac gene expression, optical mapping and contractility assays.


    Two patients had abnormal ECGs with frequent runs of ventricular tachycardia. iPSC-CMs from all DMD patients showed abnormal action potential profiles, slowed conduction velocities, and reduced sodium (INa) and inward rectifier potassium (IK1) currents. Membrane NaV1.5 and Kir2.1 protein levels were reduced in hemizygous DMD iPSC-CMs but not in heterozygous iPSC-CMs. Remarkably, transfecting just one component of the dystrophin protein complex (α1-syntrophin) in hemizygous iPSC-CMs from one patient restored channelosome function, INa and IK1 densities, and action potential profile in single cells. In addition, α1-syntrophin expression restored impulse conduction and contractility and prevented reentrant arrhythmias in hiPSC-CM monolayers.


    We provide the first demonstration that iPSC-CMs reprogrammed from skin fibroblasts of DMD patients with cardiomyopathy have a dysfunction of the NaV1.5–Kir2.1 channelosome, with consequent reduction of cardiac excitability and conduction. Altogether, iPSC-CMs from patients with DMD cardiomyopathy have a NaV1.5–Kir2.1 channelosome dysfunction, which can be rescued by the scaffolding protein α1-syntrophin to restore excitability and prevent arrhythmias.


    Supported by National Institutes of Health R01 HL122352 grant; ‘la Caixa’ Banking Foundation (HR18-00304); Fundación La Marató TV3: Ayudas a la investigación en enfermedades raras 2020 (LA MARATO-2020); Instituto de Salud Carlos III/FEDER/FSE; Horizon 2020 - Research and Innovation Framework Programme GA-965286 to JJ; the CNIC is supported by the Instituto de Salud Carlos III (ISCIII), the Ministerio de Ciencia e Innovación (MCIN) and the Pro CNIC Foundation), and is a Severo Ochoa Center of Excellence (grant CEX2020-001041-S funded by MICIN/AEI/10.13039/501100011033). American Heart Association postdoctoral fellowship 19POST34380706s to JVEN. Israel Science Foundation to OB and MA [824/19]. Rappaport grant [01012020RI]; and Niedersachsen Foundation [ZN3452] to OB; US-Israel Binational Science Foundation (BSF) to OB and TH [2019039]; Dr. Bernard Lublin Donation to OB; and The Duchenne Parent Project Netherlands (DPPNL 2029771) to OB. National Institutes of Health R01 AR068428 to DM and US-Israel Binational Science Foundation Grant [2013032] to DM and OB.