SIRT6 is a DNA double-strand break sensor

  1. Lior Onn
  2. Miguel Portillo
  3. Stefan Ilic
  4. Gal Cleitman
  5. Daniel Stein
  6. Shai Kaluski
  7. Ido Shirat
  8. Zeev Slobodnik
  9. Monica Einav
  10. Fabian Erdel
  11. Barak Akabayov
  12. Debra Toiber  Is a corresponding author
  1. Ben-Gurion University of the Negev, Israel
  2. Heidelberg University, Germany

Abstract

DNA double strand breaks are the most deleterious type of DNA damage. In this work, we show that SIRT6 directly recognizes DNA damage through a tunnel-like structure, with high affinity for double strand breaks. SIRT6 relocates to sites of damage independently of signalling and known sensors. It activates downstream signalling for double strand break repair by triggering ATM recruitment, H2AX phosphorylation and the recruitment of proteins of Homologous Recombination and Non-Homologous End Joining pathways. Our findings indicate that SIRT6 plays a previously uncharacterized role as a DNA damage sensor, a critical factor in initiating the DNA damage response (DDR). Moreover, other Sirtuins share some DSB binding capacity and DDR activation. SIRT6 activates the DDR before the repair pathway is chosen, and prevents genomic instability. Our findings place SIRT6 as a sensor of DSB, and pave the road to dissecting the contributions of distinct double strand break sensors in downstream signalling.

Data availability

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

Article and author information

Author details

  1. Lior Onn

    Department of Life Sciences, Ben-Gurion University of the Negev, Beer Sheva, Israel
    Competing interests
    The authors declare that no competing interests exist.
  2. Miguel Portillo

    Department of Life Sciences, Ben-Gurion University of the Negev, Beer Sheva, Israel
    Competing interests
    The authors declare that no competing interests exist.
  3. Stefan Ilic

    Department of Chemistry, Ben-Gurion University of the Negev, Beer Sheva, Israel
    Competing interests
    The authors declare that no competing interests exist.
  4. Gal Cleitman

    Department of Life Sciences, Ben-Gurion University of the Negev, Beer Sheva, Israel
    Competing interests
    The authors declare that no competing interests exist.
  5. Daniel Stein

    Department of Life Sciences, Ben-Gurion University of the Negev, Beer Sheva, Israel
    Competing interests
    The authors declare that no competing interests exist.
  6. Shai Kaluski

    Department of Life Sciences, Ben-Gurion University of the Negev, Beer Sheva, Israel
    Competing interests
    The authors declare that no competing interests exist.
  7. Ido Shirat

    Department of Life Sciences, Ben-Gurion University of the Negev, Beer Sheva, Israel
    Competing interests
    The authors declare that no competing interests exist.
  8. Zeev Slobodnik

    Department of Life Sciences, Ben-Gurion University of the Negev, Beer Sheva, Israel
    Competing interests
    The authors declare that no competing interests exist.
  9. Monica Einav

    Department of Life Sciences, Ben-Gurion University of the Negev, Beer Sheva, Israel
    Competing interests
    The authors declare that no competing interests exist.
  10. Fabian Erdel

    BioQuant, Heidelberg University, Heidelberg, Germany
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0003-2888-7777
  11. Barak Akabayov

    Department of Chemistry, Ben-Gurion University of the Negev, Beer Sheva, Israel
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-3882-2742
  12. Debra Toiber

    Department of Life Sciences, Ben-Gurion University of the Negev, Beer Sheva, Israel
    For correspondence
    toiber@bgu.ac.il
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-1465-0130

Funding

Israel Science Foundation (188/17)

  • Debra Toiber

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

Reviewing Editor

  1. Katrin Chua, Stanford University, United States

Publication history

  1. Received: September 4, 2019
  2. Accepted: January 23, 2020
  3. Accepted Manuscript published: January 29, 2020 (version 1)
  4. Version of Record published: March 2, 2020 (version 2)
  5. Version of Record updated: March 6, 2020 (version 3)

Copyright

© 2020, Onn 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. Lior Onn
  2. Miguel Portillo
  3. Stefan Ilic
  4. Gal Cleitman
  5. Daniel Stein
  6. Shai Kaluski
  7. Ido Shirat
  8. Zeev Slobodnik
  9. Monica Einav
  10. Fabian Erdel
  11. Barak Akabayov
  12. Debra Toiber
(2020)
SIRT6 is a DNA double-strand break sensor
eLife 9:e51636.
https://doi.org/10.7554/eLife.51636

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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.