Feedback control of Wnt signaling based on ultrastable histidine cluster co-aggregation between Naked/NKD and Axin

  1. Melissa V Gammons
  2. Miha Renko
  3. Joshua E Flack
  4. Juliusz Mieszczanek
  5. Mariann Bienz  Is a corresponding author
  1. Medical Research Council, United Kingdom

Abstract

Feedback control is a universal feature of cell signaling pathways. Naked/NKD is a widely conserved feedback regulator of Wnt signaling which controls animal development and tissue homeostasis. Naked/NKD destabilizes Dishevelled, which assembles Wnt signalosomes to inhibit the b-catenin destruction complex via recruitment of Axin. Here, we discover that the molecular mechanism underlying Naked/NKD function relies on its assembly into ultrastable decameric core aggregates via its conserved C-terminal histidine cluster (HisC). HisC aggregation is facilitated by Dishevelled and depends on accumulation of Naked/NKD during prolonged Wnt stimulation. Naked/NKD HisC cores co-aggregate with a conserved histidine cluster within Axin, to destabilize it along with Dishevelled, possibly via the autophagy receptor p62 which binds to HisC aggregates. Consistent with this, attenuated Wnt responses are observed in CRISPR-engineered flies and human epithelial cells whose Naked/NKD HisC has been deleted. Thus, HisC aggregation by Naked/NKD provides context-dependent feedback control of prolonged Wnt responses.

Data availability

All data generated and analysed during this study are included in the manuscript and supporting files.

Article and author information

Author details

  1. Melissa V Gammons

    MRC Laboratory of Molecular Biology, Medical Research Council, Cambridge, United Kingdom
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0001-9661-9331
  2. Miha Renko

    MRC Laboratory of Molecular Biology, Medical Research Council, Cambridge, United Kingdom
    Competing interests
    The authors declare that no competing interests exist.
  3. Joshua E Flack

    MRC Laboratory of Molecular Biology, Medical Research Council, Cambridge, United Kingdom
    Competing interests
    The authors declare that no competing interests exist.
  4. Juliusz Mieszczanek

    MRC Laboratory of Molecular Biology, Medical Research Council, Cambridge, United Kingdom
    Competing interests
    The authors declare that no competing interests exist.
  5. Mariann Bienz

    MRC Laboratory of Molecular Biology, Medical Research Council, Cambridge, United Kingdom
    For correspondence
    mb2@mrc-lmb.cam.ac.uk
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-7170-8706

Funding

Cancer Research UK (C7379/A24639)

  • Mariann Bienz

Medical Research Council (U105192713)

  • Mariann Bienz

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

Reviewing Editor

  1. Melanie Königshoff, University of Colorado, United States

Publication history

  1. Received: June 10, 2020
  2. Accepted: October 6, 2020
  3. Accepted Manuscript published: October 7, 2020 (version 1)
  4. Version of Record published: October 22, 2020 (version 2)

Copyright

© 2020, Gammons et al.

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.

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  1. Melissa V Gammons
  2. Miha Renko
  3. Joshua E Flack
  4. Juliusz Mieszczanek
  5. Mariann Bienz
(2020)
Feedback control of Wnt signaling based on ultrastable histidine cluster co-aggregation between Naked/NKD and Axin
eLife 9:e59879.
https://doi.org/10.7554/eLife.59879

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    Background:

    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.

    Methods:

    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.

    Results:

    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.

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    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.

    Funding:

    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.