title: Novel mitochondrial protein interactors of immunoglobulin light chains causing heart amyloidosis.
abstract: In immunoglobulin (Ig) light-chain (LC) (AL) amyloidosis, AL deposition translates into life-threatening cardiomyopathy. Clinical and experimental evidence indicates that soluble cardiotoxic LCs are themselves harmful for cells, by which they are internalized. Hypothesizing that interaction of soluble cardiotoxic LCs with cellular proteins contributes to damage, we characterized their interactome in cardiac cells. LCs were purified from patients with AL amyloidosis cardiomyopathy or multiple myeloma without amyloidosis (the nonamyloidogenic/noncardiotoxic LCs served as controls) and employed at concentrations in the range observed in AL patients' sera. A functional proteomic approach, based on direct and inverse coimmunoprecipitation and mass spectrometry, allowed identifying LC-protein complexes. Findings were validated by colocalization, fluorescence lifetime imaging microscopy (FLIM)-fluorescence resonance energy transfer (FRET), and ultrastructural studies, using human primary cardiac fibroblasts (hCFs) and stem cell-derived cardiomyocytes. Amyloidogenic cardiotoxic LCs interact in vitro with specific intracellular proteins involved in viability and metabolism. Imaging confirmed that, especially in hCFs, cardiotoxic LCs (not controls) colocalize with mitochondria and spatially associate with selected interactors: mitochondrial optic atrophy 1-like protein and peroxisomal acyl-coenzyme A oxidase 1 (FLIM-FRET efficiencies 11 and 6%, respectively). Cardiotoxic LC-treated hCFs display mitochondrial ultrastructural changes, supporting mitochondrial involvement. We show that cardiotoxic LCs establish nonphysiologic protein-protein contacts in human cardiac cells, offering new clues on the pathogenesis of AL cardiomyopathy.
authors.affiliation: *Amyloidosis Research and Treatment Center, Department of Molecular Medicine, **Department of Internal Medicine, Department of Cardiothoracic and Vascular Sciences, Laboratory of Experimental Cardiology for Cell and Molecular Therapy, University of Pavia, and Clinical Chemistry Laboratory, Fondazione Istituto di Ricovero e Cura a Carattere Scientifico Policlinico San Matteo, Pavia, Italy; Centro di Ricerca di Ingegneria Genetica (CEINGE)-Biotecnologie Avanzate, Naples, Italy; Department of Movement Sciences, Parthenope University of Naples, Naples, Italy; Department of Molecular Medicine and Medical Biotechnologies, University of Naples Federico II, Naples, Italy; Department of Biology, University of Padua, Padua, Italy; and Department of Medicine, University of Cape Town, Cape Town, South Africa.
authors.forename: Francesca
authors.initials: F
authors.lastname: Lavatelli
authors.affiliation: *Amyloidosis Research and Treatment Center, Department of Molecular Medicine, **Department of Internal Medicine, Department of Cardiothoracic and Vascular Sciences, Laboratory of Experimental Cardiology for Cell and Molecular Therapy, University of Pavia, and Clinical Chemistry Laboratory, Fondazione Istituto di Ricovero e Cura a Carattere Scientifico Policlinico San Matteo, Pavia, Italy; Centro di Ricerca di Ingegneria Genetica (CEINGE)-Biotecnologie Avanzate, Naples, Italy; Department of Movement Sciences, Parthenope University of Naples, Naples, Italy; Department of Molecular Medicine and Medical Biotechnologies, University of Naples Federico II, Naples, Italy; Department of Biology, University of Padua, Padua, Italy; and Department of Medicine, University of Cape Town, Cape Town, South Africa.
authors.forename: Esther
authors.initials: E
authors.lastname: Imperlini
authors.affiliation: *Amyloidosis Research and Treatment Center, Department of Molecular Medicine, **Department of Internal Medicine, Department of Cardiothoracic and Vascular Sciences, Laboratory of Experimental Cardiology for Cell and Molecular Therapy, University of Pavia, and Clinical Chemistry Laboratory, Fondazione Istituto di Ricovero e Cura a Carattere Scientifico Policlinico San Matteo, Pavia, Italy; Centro di Ricerca di Ingegneria Genetica (CEINGE)-Biotecnologie Avanzate, Naples, Italy; Department of Movement Sciences, Parthenope University of Naples, Naples, Italy; Department of Molecular Medicine and Medical Biotechnologies, University of Naples Federico II, Naples, Italy; Department of Biology, University of Padua, Padua, Italy; and Department of Medicine, University of Cape Town, Cape Town, South Africa.
authors.forename: Stefania
authors.initials: S
authors.lastname: Orrù
authors.affiliation: *Amyloidosis Research and Treatment Center, Department of Molecular Medicine, **Department of Internal Medicine, Department of Cardiothoracic and Vascular Sciences, Laboratory of Experimental Cardiology for Cell and Molecular Therapy, University of Pavia, and Clinical Chemistry Laboratory, Fondazione Istituto di Ricovero e Cura a Carattere Scientifico Policlinico San Matteo, Pavia, Italy; Centro di Ricerca di Ingegneria Genetica (CEINGE)-Biotecnologie Avanzate, Naples, Italy; Department of Movement Sciences, Parthenope University of Naples, Naples, Italy; Department of Molecular Medicine and Medical Biotechnologies, University of Naples Federico II, Naples, Italy; Department of Biology, University of Padua, Padua, Italy; and Department of Medicine, University of Cape Town, Cape Town, South Africa.
authors.forename: Paola
authors.initials: P
authors.lastname: Rognoni
authors.affiliation: *Amyloidosis Research and Treatment Center, Department of Molecular Medicine, **Department of Internal Medicine, Department of Cardiothoracic and Vascular Sciences, Laboratory of Experimental Cardiology for Cell and Molecular Therapy, University of Pavia, and Clinical Chemistry Laboratory, Fondazione Istituto di Ricovero e Cura a Carattere Scientifico Policlinico San Matteo, Pavia, Italy; Centro di Ricerca di Ingegneria Genetica (CEINGE)-Biotecnologie Avanzate, Naples, Italy; Department of Movement Sciences, Parthenope University of Naples, Naples, Italy; Department of Molecular Medicine and Medical Biotechnologies, University of Naples Federico II, Naples, Italy; Department of Biology, University of Padua, Padua, Italy; and Department of Medicine, University of Cape Town, Cape Town, South Africa.
authors.forename: Daniela
authors.initials: D
authors.lastname: Sarnataro
authors.affiliation: *Amyloidosis Research and Treatment Center, Department of Molecular Medicine, **Department of Internal Medicine, Department of Cardiothoracic and Vascular Sciences, Laboratory of Experimental Cardiology for Cell and Molecular Therapy, University of Pavia, and Clinical Chemistry Laboratory, Fondazione Istituto di Ricovero e Cura a Carattere Scientifico Policlinico San Matteo, Pavia, Italy; Centro di Ricerca di Ingegneria Genetica (CEINGE)-Biotecnologie Avanzate, Naples, Italy; Department of Movement Sciences, Parthenope University of Naples, Naples, Italy; Department of Molecular Medicine and Medical Biotechnologies, University of Naples Federico II, Naples, Italy; Department of Biology, University of Padua, Padua, Italy; and Department of Medicine, University of Cape Town, Cape Town, South Africa.
authors.forename: Giuseppina
authors.initials: G
authors.lastname: Palladini
authors.affiliation: *Amyloidosis Research and Treatment Center, Department of Molecular Medicine, **Department of Internal Medicine, Department of Cardiothoracic and Vascular Sciences, Laboratory of Experimental Cardiology for Cell and Molecular Therapy, University of Pavia, and Clinical Chemistry Laboratory, Fondazione Istituto di Ricovero e Cura a Carattere Scientifico Policlinico San Matteo, Pavia, Italy; Centro di Ricerca di Ingegneria Genetica (CEINGE)-Biotecnologie Avanzate, Naples, Italy; Department of Movement Sciences, Parthenope University of Naples, Naples, Italy; Department of Molecular Medicine and Medical Biotechnologies, University of Naples Federico II, Naples, Italy; Department of Biology, University of Padua, Padua, Italy; and Department of Medicine, University of Cape Town, Cape Town, South Africa.
authors.forename: Giuseppe
authors.initials: G
authors.lastname: Malpasso
authors.affiliation: *Amyloidosis Research and Treatment Center, Department of Molecular Medicine, **Department of Internal Medicine, Department of Cardiothoracic and Vascular Sciences, Laboratory of Experimental Cardiology for Cell and Molecular Therapy, University of Pavia, and Clinical Chemistry Laboratory, Fondazione Istituto di Ricovero e Cura a Carattere Scientifico Policlinico San Matteo, Pavia, Italy; Centro di Ricerca di Ingegneria Genetica (CEINGE)-Biotecnologie Avanzate, Naples, Italy; Department of Movement Sciences, Parthenope University of Naples, Naples, Italy; Department of Molecular Medicine and Medical Biotechnologies, University of Naples Federico II, Naples, Italy; Department of Biology, University of Padua, Padua, Italy; and Department of Medicine, University of Cape Town, Cape Town, South Africa.
authors.forename: Maria Eugenia
authors.initials: ME
authors.lastname: Soriano
authors.affiliation: *Amyloidosis Research and Treatment Center, Department of Molecular Medicine, **Department of Internal Medicine, Department of Cardiothoracic and Vascular Sciences, Laboratory of Experimental Cardiology for Cell and Molecular Therapy, University of Pavia, and Clinical Chemistry Laboratory, Fondazione Istituto di Ricovero e Cura a Carattere Scientifico Policlinico San Matteo, Pavia, Italy; Centro di Ricerca di Ingegneria Genetica (CEINGE)-Biotecnologie Avanzate, Naples, Italy; Department of Movement Sciences, Parthenope University of Naples, Naples, Italy; Department of Molecular Medicine and Medical Biotechnologies, University of Naples Federico II, Naples, Italy; Department of Biology, University of Padua, Padua, Italy; and Department of Medicine, University of Cape Town, Cape Town, South Africa.
authors.forename: Andrea
authors.initials: A
authors.lastname: Di Fonzo
authors.affiliation: *Amyloidosis Research and Treatment Center, Department of Molecular Medicine, **Department of Internal Medicine, Department of Cardiothoracic and Vascular Sciences, Laboratory of Experimental Cardiology for Cell and Molecular Therapy, University of Pavia, and Clinical Chemistry Laboratory, Fondazione Istituto di Ricovero e Cura a Carattere Scientifico Policlinico San Matteo, Pavia, Italy; Centro di Ricerca di Ingegneria Genetica (CEINGE)-Biotecnologie Avanzate, Naples, Italy; Department of Movement Sciences, Parthenope University of Naples, Naples, Italy; Department of Molecular Medicine and Medical Biotechnologies, University of Naples Federico II, Naples, Italy; Department of Biology, University of Padua, Padua, Italy; and Department of Medicine, University of Cape Town, Cape Town, South Africa.
authors.forename: Veronica
authors.initials: V
authors.lastname: Valentini
authors.affiliation: *Amyloidosis Research and Treatment Center, Department of Molecular Medicine, **Department of Internal Medicine, Department of Cardiothoracic and Vascular Sciences, Laboratory of Experimental Cardiology for Cell and Molecular Therapy, University of Pavia, and Clinical Chemistry Laboratory, Fondazione Istituto di Ricovero e Cura a Carattere Scientifico Policlinico San Matteo, Pavia, Italy; Centro di Ricerca di Ingegneria Genetica (CEINGE)-Biotecnologie Avanzate, Naples, Italy; Department of Movement Sciences, Parthenope University of Naples, Naples, Italy; Department of Molecular Medicine and Medical Biotechnologies, University of Naples Federico II, Naples, Italy; Department of Biology, University of Padua, Padua, Italy; and Department of Medicine, University of Cape Town, Cape Town, South Africa.
authors.forename: Massimiliano
authors.initials: M
authors.lastname: Gnecchi
authors.affiliation: *Amyloidosis Research and Treatment Center, Department of Molecular Medicine, **Department of Internal Medicine, Department of Cardiothoracic and Vascular Sciences, Laboratory of Experimental Cardiology for Cell and Molecular Therapy, University of Pavia, and Clinical Chemistry Laboratory, Fondazione Istituto di Ricovero e Cura a Carattere Scientifico Policlinico San Matteo, Pavia, Italy; Centro di Ricerca di Ingegneria Genetica (CEINGE)-Biotecnologie Avanzate, Naples, Italy; Department of Movement Sciences, Parthenope University of Naples, Naples, Italy; Department of Molecular Medicine and Medical Biotechnologies, University of Naples Federico II, Naples, Italy; Department of Biology, University of Padua, Padua, Italy; and Department of Medicine, University of Cape Town, Cape Town, South Africa.
authors.forename: Stefano
authors.initials: S
authors.lastname: Perlini
authors.affiliation: *Amyloidosis Research and Treatment Center, Department of Molecular Medicine, **Department of Internal Medicine, Department of Cardiothoracic and Vascular Sciences, Laboratory of Experimental Cardiology for Cell and Molecular Therapy, University of Pavia, and Clinical Chemistry Laboratory, Fondazione Istituto di Ricovero e Cura a Carattere Scientifico Policlinico San Matteo, Pavia, Italy; Centro di Ricerca di Ingegneria Genetica (CEINGE)-Biotecnologie Avanzate, Naples, Italy; Department of Movement Sciences, Parthenope University of Naples, Naples, Italy; Department of Molecular Medicine and Medical Biotechnologies, University of Naples Federico II, Naples, Italy; Department of Biology, University of Padua, Padua, Italy; and Department of Medicine, University of Cape Town, Cape Town, South Africa salvator@unina.it gmerlini@unipv.it.
authors.forename: Francesco
authors.initials: F
authors.lastname: Salvatore
authors.affiliation: *Amyloidosis Research and Treatment Center, Department of Molecular Medicine, **Department of Internal Medicine, Department of Cardiothoracic and Vascular Sciences, Laboratory of Experimental Cardiology for Cell and Molecular Therapy, University of Pavia, and Clinical Chemistry Laboratory, Fondazione Istituto di Ricovero e Cura a Carattere Scientifico Policlinico San Matteo, Pavia, Italy; Centro di Ricerca di Ingegneria Genetica (CEINGE)-Biotecnologie Avanzate, Naples, Italy; Department of Movement Sciences, Parthenope University of Naples, Naples, Italy; Department of Molecular Medicine and Medical Biotechnologies, University of Naples Federico II, Naples, Italy; Department of Biology, University of Padua, Padua, Italy; and Department of Medicine, University of Cape Town, Cape Town, South Africa salvator@unina.it gmerlini@unipv.it.
authors.forename: Giampaolo
authors.initials: G
authors.lastname: Merlini
mesh_terms: D000328:Adult; D000686:Amyloidosis / Q000378:metabolism* / Q000473:pathology; D000818:Animals; D005260:Female; D005347:Fibroblasts / Q000378:metabolism* / Q000473:pathology; D006331:Heart Diseases / Q000378:metabolism* / Q000473:pathology; D006801:Humans; D007147:Immunoglobulin Light Chains / Q000378:metabolism*; D008297:Male; D024101:Mitochondrial Proteins / Q000378:metabolism*; D032383:Myocytes, Cardiac / Q000378:metabolism* / Q000473:pathology; D051381:Rats; D017207:Rats, Sprague-Dawley
chemical_list: D007147:Immunoglobulin Light Chains; D024101:Mitochondrial Proteins
keywords: functional proteomics; human cardiac cells; protein-misfolding diseases; protein–protein interaction; proteotoxicity