{ "abstract": "Sarcomeres, the basic repeating unit of striated muscle, are joined together by crosslinked actin filaments found at the boundaries of muscle sarcomeres, termed Z-discs. Z-discs play a key role in cardiac signalling and disease, however, the arrangement and function of many of the proteins present in the Z-disc remain to be understood. Here, we determined the organisation of 3 key proteins, ZASP, ɑ-actinin-2 and the Z1Z2 epitope of titin, located within the Z-disc. We fluorescently labelled these proteins in cardiac myofibrils using Adhirons specific to each protein and used interferometric photoactivated localization microscopy (iPALM) to obtain the 3D position of these proteins to a high precision (<10nm in x,y,z). We then used PERPL (Pattern Extraction from Relative Positions of Localisations: available at https://github.com/oubino/z_disk/releases/tag/v0.0.3) to analyse patterns in the relative positions of the proteins and reveal their underlying organisation. This analysis revealed that ZASP and ɑ-actinin-2 have a similar repeating organisation, but that the organisation of Z1Z2 is different. Author summary: Z-discs connect myosin- and actin-containing filaments from adjacent sarcomeres in striated (cardiac and skeletal) muscle. They contain over 50 different interacting proteins that have structural and/or signalling roles. Understanding their organisation in Z-discs is important for understanding their role. However, it is challenging to image Z-discs as they are narrower (~100 nm wide) than the resolution limit of conventional light microscopes (~250 nm). To overcome this, we used super-resolution imaging (iPALM: interferomic photoactivation and localisation microscopy), which has a resolution better than 10 nm in all three dimensions (x,y and z) to image specific proteins the z-discs of myofibrils from pig cardiac myocytes. We used small (< 4 nm in size) fluorescently-labelled binding proteins (Adhirons) to label ɑ‑actinin-2, a protein that cross-links actin-containing filaments from adjacent sarcomeres; the N-terminal region (Z1Z2) of titin, a large protein that regulates the length of myosin‑containing filaments; or ZASP (Z-band Alternative Spliced PDZ motif), which might help to regulate Z-disc width. We performed a tailored downstream analysis of the iPALM single molecule localisation data to uncover the detailed organisation of each protein. This revealed that ZASP and ɑ-actinin-2 have a similar repeating organisation, while titin Z1Z2 has a different, yet regular organisation throughout the Z-disc.", "abstract_shown": true, "affiliations": [ "School of Molecular and Cellular Biology, Faculty of Biological Sciences, University of Leeds, Leeds, United Kingdom", "Engineering and Physical Sciences Faculty Services, Faculty of Engineering and Physical Sciences, University of Leeds, Leeds, United Kingdom", "Astbury Centre for Structural and Molecular Biology University of Leeds, Leeds, United Kingdom", "Advanced Imaging Center, Janelia Research Campus, Howard Hughes Medical Institute, Ashburn, United States of America" ], "article_type": "Research Article", "arxiv": null, "author_total": 11, "authors": [ { "affs": [ 0 ], "name": "Oliver Umney" }, { "affs": [ 1 ], "name": "Alistair P. Curd" }, { "affs": [ 2 ], "name": "Heather L. Martin" }, { "affs": [ 3 ], "name": "Tarek Lewis" }, { "name": "Anna Ah-San Tang" }, { "name": "Thembaninkosi Gaule" }, { "name": "Iain W. Manfield" }, { "name": "Harikrushnan Balasubramanian" }, { "name": "Satya Khuon" }, { "name": "Jesse Aaron" }, { "name": "Michelle Peckham" } ], "bibtex": "https://research.draconitic.com/paper/61dk49hccddxv.bib", "cited_by": 0, "doi": "10.1371/journal.pcbi.1014309", "fulltext_url": "https://journals.plos.org/ploscompbiol/article?id=10.1371/journal.pcbi.1014309", "id": "61dk49hccddxv", "issns": [ "1553-7358" ], "landing_url": "https://journals.plos.org/ploscompbiol/article?id=10.1371/journal.pcbi.1014309", "language": null, "licence": "CC-BY-4.0", "licence_label": "CC BY 4.0", "licence_source": "journal", "licence_url": "https://creativecommons.org/licenses/by/4.0/", "metadata_licence": null, "native_subjects": [ [ "openalex", "1304|Advanced Fluorescence Microscopy Techniques" ], [ "plos", "/Biology and life sciences/Anatomy/Biological tissue/Muscle tissue/Muscle cells/Myofibrils" ], [ "plos", "/Biology and life sciences/Anatomy/Biological tissue/Muscle tissue/Muscle cells/Myofibrils/Sarcomeres" ], [ "plos", "/Biology and life sciences/Anatomy/Musculoskeletal system/Muscles/Cardiac muscles" ], [ "plos", "/Biology and life sciences/Biochemistry/Proteins/Motor proteins" ], [ "plos", "/Biology and life sciences/Biochemistry/Proteins/Muscle proteins" ], [ "plos", "/Biology and life sciences/Biochemistry/Proteins/Protein domains" ], [ "plos", "/Biology and life sciences/Biochemistry/Proteins/Structural proteins" ], [ "plos", "/Biology and life sciences/Cell biology/Cell motility/Actin filaments" ], [ "plos", "/Biology and life sciences/Cell biology/Cellular types/Animal cells/Muscle cells/Myofibrils" ], [ "plos", "/Biology and life sciences/Cell biology/Cellular types/Animal cells/Muscle cells/Myofibrils/Sarcomeres" ], [ "plos", "/Biology and life sciences/Cell biology/Molecular motors/Motor proteins" ], [ "plos", "/Medicine and health sciences/Anatomy/Biological tissue/Muscle tissue/Muscle cells/Myofibrils" ], [ "plos", "/Medicine and health sciences/Anatomy/Biological tissue/Muscle tissue/Muscle cells/Myofibrils/Sarcomeres" ], [ "plos", "/Medicine and health sciences/Anatomy/Musculoskeletal system/Muscles/Cardiac muscles" ] ], "openalex": "W7214641931", "pdf_url": "https://journals.plos.org/ploscompbiol/article/file?id=10.1371/journal.pcbi.1014309&type=printable", "pmcid": null, "pmid": null, "published": "2026-09-28", "publisher": "Public Library of Science", "ris": "https://research.draconitic.com/paper/61dk49hccddxv.ris", "sources": [ { "first_seen": "2026-10-30T12:17:37Z", "native_id": "10.1371/journal.pcbi.1014309", "source": "plos", "url": "https://journals.plos.org/ploscompbiol/article?id=10.1371/journal.pcbi.1014309" } ], "subjects": [ { "field": "biology", "origin": "plos:cell biology", "subfield": "cell-biology" }, { "field": "medicine", "origin": "plos:anatomy", "subfield": "physiology" } ], "title": "Using iPALM to determine protein organisation in cardiac muscle Z-discs", "title_text": "Using iPALM to determine protein organisation in cardiac muscle Z-discs", "type": "peer-reviewed", "updated_at": "2026-10-30T12:17:37Z", "url": "https://research.draconitic.com/paper/61dk49hccddxv", "venue": "PLOS Computational Biology", "versions": [] }