Novel mitochondrial protein interactors of immunoglobulin light chains causing heart amyloidosis

Francesca Lavatelli, Esther Imperlini, Stefania Orrù, Paola Rognoni, Daniela Sarnataro, Giuseppina Palladini, Giuseppe Malpasso, Maria Eugenia Soriano, Andrea Di Fonzo, Veronica Valentini, Massimiliano Gnecchi, Stefano Perlini, Francesco Salvatore, Giampaolo Merlini

Research output: Contribution to journalArticle

31 Citations (Scopus)

Abstract

In immunoglobulin (Ig) light-chain (LC) (AL) amyloidosis, AL deposition translates into lifethreatening 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 multiplemyeloma 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.

Original languageEnglish
Pages (from-to)4614-4628
Number of pages15
JournalFASEB Journal
Volume29
Issue number11
DOIs
Publication statusPublished - Nov 1 2015

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Immunoglobulin Light Chains
Mitochondrial Proteins
Amyloidosis
Fibroblasts
Cardiomyopathies
Fluorescence Resonance Energy Transfer
Proteins
Optical Imaging
Imaging techniques
Microscopy
Autosomal Dominant Optic Atrophy
Fluorescence
Acyl-CoA Oxidase
Light
Mitochondria
Stem cells
Cardiac Myocytes
Metabolism
Proteomics
Mass spectrometry

Keywords

  • Functional proteomics
  • Human cardiac cells
  • Protein-misfolding diseases
  • Protein-protein interaction
  • Proteotoxicity

ASJC Scopus subject areas

  • Biochemistry
  • Biotechnology
  • Genetics
  • Molecular Biology
  • Medicine(all)

Cite this

Novel mitochondrial protein interactors of immunoglobulin light chains causing heart amyloidosis. / Lavatelli, Francesca; Imperlini, Esther; Orrù, Stefania; Rognoni, Paola; Sarnataro, Daniela; Palladini, Giuseppina; Malpasso, Giuseppe; Soriano, Maria Eugenia; Di Fonzo, Andrea; Valentini, Veronica; Gnecchi, Massimiliano; Perlini, Stefano; Salvatore, Francesco; Merlini, Giampaolo.

In: FASEB Journal, Vol. 29, No. 11, 01.11.2015, p. 4614-4628.

Research output: Contribution to journalArticle

Lavatelli, Francesca ; Imperlini, Esther ; Orrù, Stefania ; Rognoni, Paola ; Sarnataro, Daniela ; Palladini, Giuseppina ; Malpasso, Giuseppe ; Soriano, Maria Eugenia ; Di Fonzo, Andrea ; Valentini, Veronica ; Gnecchi, Massimiliano ; Perlini, Stefano ; Salvatore, Francesco ; Merlini, Giampaolo. / Novel mitochondrial protein interactors of immunoglobulin light chains causing heart amyloidosis. In: FASEB Journal. 2015 ; Vol. 29, No. 11. pp. 4614-4628.
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AU - Lavatelli, Francesca

AU - Imperlini, Esther

AU - Orrù, Stefania

AU - Rognoni, Paola

AU - Sarnataro, Daniela

AU - Palladini, Giuseppina

AU - Malpasso, Giuseppe

AU - Soriano, Maria Eugenia

AU - Di Fonzo, Andrea

AU - Valentini, Veronica

AU - Gnecchi, Massimiliano

AU - Perlini, Stefano

AU - Salvatore, Francesco

AU - Merlini, Giampaolo

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N2 - In immunoglobulin (Ig) light-chain (LC) (AL) amyloidosis, AL deposition translates into lifethreatening 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 multiplemyeloma 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.

AB - In immunoglobulin (Ig) light-chain (LC) (AL) amyloidosis, AL deposition translates into lifethreatening 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 multiplemyeloma 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.

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