{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,7,16]],"date-time":"2026-07-16T12:40:14Z","timestamp":1784205614134,"version":"3.55.0"},"reference-count":40,"publisher":"Wiley","issue":"4","license":[{"start":{"date-parts":[[2025,7,10]],"date-time":"2025-07-10T00:00:00Z","timestamp":1752105600000},"content-version":"vor","delay-in-days":0,"URL":"http:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"content-domain":{"domain":["onlinelibrary.wiley.com"],"crossmark-restriction":true},"short-container-title":["J cachexia sarcopenia muscle"],"published-print":{"date-parts":[[2025,8]]},"abstract":"ABSTRACT<\/jats:title>\n \n Background<\/jats:title>\n \n Plectin, a multifunctional cytolinker and intermediate filament stabilizing protein, is essential for muscle fibre integrity and function. Mutations in the human plectin gene (\n PLEC<\/jats:italic>\n ) cause autosomal recessive epidermolysis bullosa simplex with muscular dystrophy (EBS\u2010MD). The disorganization and aggregation of desmin filaments in conjunction with degenerative changes of the myofibrillar apparatus are key features in the skeletal muscle pathology of EBS\u2010MD. We performed a comprehensive analysis addressing protein homeostasis in this rare protein aggregation disease by using human EBS\u2010MD tissue, plectin knock\u2010out mice and plectin\u2010deficient cells.\n <\/jats:p>\n <\/jats:sec>\n \n Methods<\/jats:title>\n \n Protein degradation pathways were analysed in muscles from EBS\u2010MD patients, muscle\u2010specific conditional plectin knockout (MCK\u2010Cre\/cKO) mice, as well as in plectin\u2010deficient (\n Plec<\/jats:italic>\n \n \u2212\/\u2212<\/jats:italic>\n <\/jats:sup>\n ) myoblasts by electron and immunofluorescence microscopy. To obtain a comprehensive picture of autophagic processes, we evaluated the transcriptional regulation and expression levels of autophagic markers in plectin\u2010deficient muscles and myoblasts (RNA\u2010Seq, qRT\u2010PCR, immunoblotting). Autophagic turnover was dynamically assessed by measuring baseline autophagy as well as specific inhibition and activation in mCherry\u2010EGFP\u2010LC3B\u2010expressing\n Plec<\/jats:italic>\n \n +\/+<\/jats:italic>\n <\/jats:sup>\n and\n Plec<\/jats:italic>\n \n \u2212\/\u2212<\/jats:italic>\n <\/jats:sup>\n myoblasts, and by monitoring primary\n Plec<\/jats:italic>\n \n +\/+<\/jats:italic>\n <\/jats:sup>\n and\n Plec<\/jats:italic>\n \n \u2212\/\u2212<\/jats:italic>\n <\/jats:sup>\n myoblasts using organelle\u2010specific dyes. Wild\u2010type and MCK\u2010Cre\/cKO mice were treated with chloroquine or metformin to assess the effects of autophagy inhibition and activation in\u00a0vivo.\n <\/jats:p>\n <\/jats:sec>\n \n Results<\/jats:title>\n \n Our study identified the accumulation of degradative vacuoles as well as LC3\u2010 and SQSTM1\u2010positive patches in EBS\u2010MD patients, MCK\u2010Cre\/cKO mouse muscles and\n Plec<\/jats:italic>\n \n \u2212\/\u2212<\/jats:italic>\n <\/jats:sup>\n myoblasts. The transcriptional regulation of ~30% of autophagy\u2010related genes was altered, and protein levels of downstream targets of the autophagosomal degradation machinery were elevated in MCK\u2010Cre\/cKO muscle lysates (e.g., LAMP2, BAG3 and SQSTM1 to ~160, ~150 and ~140% of controls, respectively;\n p<\/jats:italic>\n \u2009<\u20090.05). Autophagosome turnover was compromised in mCherry\u2010EGFP\u2010LC3B\u2010expressing\n Plec<\/jats:italic>\n \n \u2212\/\u2212<\/jats:italic>\n <\/jats:sup>\n myoblasts (~40% reduction in median red:green ratio, reduced puncta number, smaller puncta;\n p<\/jats:italic>\n \u2009<\u20090.01). By labelling autophagic compartments with CYTO\u2010ID dye or lysosomes with LYSO\u2010ID, we found reduced signal intensities in primary\n Plec<\/jats:italic>\n \n \u2212\/\u2212<\/jats:italic>\n <\/jats:sup>\n cells (\n p<\/jats:italic>\n \u2009<\u20090.001). Treatment with chloroquine led to drastic swelling of autophagic vacuoles in primary\n Plec<\/jats:italic>\n \n +\/+<\/jats:italic>\n <\/jats:sup>\n myoblasts, while the swelling in\n Plec<\/jats:italic>\n \n \u2212\/\u2212<\/jats:italic>\n <\/jats:sup>\n cells was moderate, establishing a defect in their autophagic clearance. Chloroquine treatment of MCK\u2010Cre\/cKO mice corroborated that loss of plectin coincides with impaired autophagic clearance, while metformin amelioratively induced autophagic flux.\n <\/jats:p>\n <\/jats:sec>\n \n Conclusions<\/jats:title>\n Our work demonstrates that the characteristic protein aggregation pathology in EBS\u2010MD is linked to an impaired autophagic flux. The obtained results open a new perspective on the understanding of the protein aggregation pathology in plectin\u2010related disorders and provide a basis for further pharmacological intervention.<\/jats:p>\n <\/jats:sec>","DOI":"10.1002\/jcsm.70001","type":"journal-article","created":{"date-parts":[[2025,7,11]],"date-time":"2025-07-11T05:59:42Z","timestamp":1752213582000},"update-policy":"https:\/\/doi.org\/10.1002\/crossmark_policy","source":"Crossref","is-referenced-by-count":3,"title":["Impaired Autophagic Flux in Skeletal Muscle of Plectin\u2010Related Epidermolysis Bullosa Simplex With Muscular Dystrophy"],"prefix":"10.1002","volume":"16","author":[{"given":"Michaela\u00a0M.","family":"Zrelski","sequence":"first","affiliation":[{"name":"Division of Cell and Developmental Biology, Center for Anatomy and Cell Biology Medical University of Vienna Vienna 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Anatomy and Cell Biology Medical University of Vienna Vienna Austria"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Leonid","family":"Mill","sequence":"additional","affiliation":[{"name":"MIRA Vision Microscopy GmbH Wangen Germany"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Monika","family":"Kustermann","sequence":"additional","affiliation":[{"name":"Neuromuscular Research Group, Division of Cell and Developmental Biology, Center for Anatomy and Cell Biology Medical University of Vienna Vienna Austria"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Wolfgang\u00a0M.","family":"Schmidt","sequence":"additional","affiliation":[{"name":"Neuromuscular Research Group, Division of Cell and Developmental Biology, Center for Anatomy and Cell Biology Medical University of Vienna Vienna Austria"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Fiona","family":"Norwood","sequence":"additional","affiliation":[{"name":"Department 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