{
  "abstract": "Introduction During heart disease, the cardiac extracellular matrix (ECM) mechanically transforms, resulting in the stiffening of the ECM. Cardiomyocytes sense this ECM change, leading to phenotypic remodelling. A key part of the cardiomyocyte mechanosensing machinery is organised at the costamere; integrins and associated proteins, bridging the ECM and the sarcomere, facilitating force transmission. An integral component of the costamere is the mechanosensitive protein talin, with rod domains which unfold and refold in the presence and absence of physiological forces, exposing cryptic binding sites leading to force-dependent interactions with talin binding partners. While talin has been established as a mechanical sensor, the specifics of its mechanosensing pathway remain elusive.Methods Co-immunoprecipitation, fluorescence recovery after photobleaching, and fluorescence resonance energy transfer experiments were performed on neonate rat cardiomyocytes seeded on PDMS of stiffnesses corresponding to that of embryonic (1 kPa), healthy (6 kPa) and diseased (20–130 kPa) cardiac tissue. In situ proximity ligation was conducted on wildtype and MLP knockout mouse heart sections. The LOVTRAP optogenetics system was performed on the C2C12 mouse myoblast cell line, and fluorescence polarisation in vitro binding experiments used talin R7R8 domain, DLC1 and RIAM synthetic proteins expressed and purified from E. coli BL21. Adenoviral transduction and siRNA were used to overexpress and knockdown proteins, respectively.Results We identified three talin-interacting proteins, DLC1, RIAM and paxillin, which each preferentially bound to talin at specific ECM stiffnesses – paxillin preferred 1 kPa, DLC1 6 kPa and RIAM 20–130 kPa – and subsequently confirmed this in situ in healthy and diseased mouse hearts. The interactions between talin and DLC1 or RIAM were shown to be regulated through direct competition for binding at the cellular and molecular level, yielding Kd values of 4.9 µM and 9.2 µM for DLC1 and RIAM, respectively. These interactions were present even when talin was in a tension-free state during the immunoprecipitation, suggesting the competitive interactions might be further imprinted, i.e. stabilised through other mechanisms such as phosphorylation. Focal adhesion kinase activity was shown to regulate the interaction between talin and DLC1 or RIAM: inhibition of Src family kinases and FAK altered the talin interactions, promoting the talin-RIAM interaction over the talin-DLC1 at 6 kPa. Finally, we identified DLC1, as a major RhoGAP, regulated cardiomyocyte RhoA activity in a stiffness-dependent manner, with DLC1 knockdown resulting in cytoskeletal disruptions.Conclusions This work demonstrated a mechanism of imprinting mechanical information into the talin interactome to finetune RhoA activity, contributing to our knowledge of cardiomyocyte mechanosignalling which can help improve our understanding of cardiac health and disease.",
  "authors": [
    {
      "affiliations": [
        "School of Engineering and Materials Science, Queen Mary University of London, UK",
        "Univ. Bordeaux, CNRS, Bordeaux INP, CBMN, UMR 5248, F-33600 Pessac, France",
        "#equal contribution"
      ],
      "name": "Emilie Marhuenda"
    },
    {
      "affiliations": [
        "School of Engineering and Materials Science, Queen Mary University of London, UK",
        "#equal contribution"
      ],
      "name": "Ioannis Xanthis"
    },
    {
      "affiliations": [
        "School of Engineering and Materials Science, Queen Mary University of London, UK",
        "*Corresponding author"
      ],
      "name": "Poppy O Smith"
    },
    {
      "affiliations": [
        "School of Engineering and Materials Science, Queen Mary University of London, UK"
      ],
      "name": "Aishwarya Prakash"
    },
    {
      "affiliations": [
        "Department of Biochemistry, Cell and Systems Biology, Institute of Systems, Molecular and Integrative Biology, University of Liverpool, Crown Street, Liverpool L69 7ZB, UK"
      ],
      "name": "Till Kallem"
    },
    {
      "affiliations": [
        "School of Engineering and Materials Science, Queen Mary University of London, UK"
      ],
      "name": "Pragati Pandey"
    },
    {
      "affiliations": [
        "School of Engineering and Materials Science, Queen Mary University of London, UK"
      ],
      "name": "Darren Graham Samuel Wilson"
    },
    {
      "affiliations": [
        "Department of Biochemistry, Cell and Systems Biology, Institute of Systems, Molecular and Integrative Biology, University of Liverpool, Crown Street, Liverpool L69 7ZB, UK"
      ],
      "name": "Amar Azad"
    },
    {
      "affiliations": [
        "Department of Biochemistry, Cell and Systems Biology, Institute of Systems, Molecular and Integrative Biology, University of Liverpool, Crown Street, Liverpool L69 7ZB, UK"
      ],
      "name": "Megan Richter"
    },
    {
      "affiliations": [
        "Department of Biochemistry, Cell and Systems Biology, Institute of Systems, Molecular and Integrative Biology, University of Liverpool, Crown Street, Liverpool L69 7ZB, UK"
      ],
      "name": "Davor Pavolvic"
    },
    {
      "affiliations": [
        "Department of Biochemistry, Cell and Systems Biology, Institute of Systems, Molecular and Integrative Biology, University of Liverpool, Crown Street, Liverpool L69 7ZB, UK",
        "Institute of Cardiovascular Sciences, University of Birmingham, Birmingham B15 2TT, UK"
      ],
      "name": "Katja Gehmlich"
    },
    {
      "affiliations": [
        "Division of Cardiovascular Medicine, Radcliffe Department of Medicine and British Heart Foundation Centre of Research Excellence Oxford, University of Oxford, Oxford, OX3 9DU, UK"
      ],
      "name": "Giuseppe Faggian"
    },
    {
      "affiliations": [
        "School of Engineering and Materials Science, Queen Mary University of London, UK",
        "Divisione Ospedaliero Universitaria Cardiochirurgia Verona, Verona, Italy"
      ],
      "name": "Pamela Swiatlowska"
    },
    {
      "affiliations": [
        "National Heart and Lung Institute, Imperial College London, UK"
      ],
      "name": "Elisabeth Ehler"
    },
    {
      "affiliations": [
        "School of Cardiovascular and Metabolic Medicine and Sciences, King’s College London, UK"
      ],
      "name": "James Levitt"
    },
    {
      "affiliations": [
        "School of Cardiovascular and Metabolic Medicine and Sciences, King’s College London, UK"
      ],
      "name": "Simon P Poland"
    },
    {
      "affiliations": [
        "School of Cardiovascular and Metabolic Medicine and Sciences, King’s College London, UK"
      ],
      "name": "Simon Ameer-Beg"
    },
    {
      "affiliations": [
        "Department of Biochemistry, Cell and Systems Biology, Institute of Systems, Molecular and Integrative Biology, University of Liverpool, Crown Street, Liverpool L69 7ZB, UK"
      ],
      "name": "Benjamin T Goult"
    },
    {
      "affiliations": [
        "School of Engineering and Materials Science, Queen Mary University of London, UK"
      ],
      "name": "Thomas Iskratsch"
    }
  ],
  "title": "BS42 Mechanical imprinting of the talin interaction with DLC1 regulates RHOA activity and cardiomyocyte remodelling",
  "uid": "e57457ab-124d-5b23-b64f-f6046351f91a"
}