Tandem detergent-extraction and immunoprecipitation of proteinopathy: Scalable enrichment of ALS-associated TDP-43 aggregates

Baggio A. Evangelista; Shannon R. Cahalan; Joey V. Ragusa; Angie Mordant; Julie C. Necarsulmer; Robert J. Perna; Tejazaditya Ajit; Kristen White; Natalie K. Barker; Xu Tian; Sarah Cohen; Rick Meeker; Laura E. Herring; Todd J. Cohen

iScience (May 2023)

Tandem detergent-extraction and immunoprecipitation of proteinopathy: Scalable enrichment of ALS-associated TDP-43 aggregates

Baggio A. Evangelista,
Shannon R. Cahalan,
Joey V. Ragusa,
Angie Mordant,
Julie C. Necarsulmer,
Robert J. Perna,
Tejazaditya Ajit,
Kristen White,
Natalie K. Barker,
Xu Tian,
Sarah Cohen,
Rick Meeker,
Laura E. Herring,
Todd J. Cohen

Affiliations

Baggio A. Evangelista: Department of Cell Biology and Physiology, University of North Carolina at Chapel Hill, Chapel Hill, NC, USA
Shannon R. Cahalan: Medical Student Training in Aging Research, Center for Aging and Health, University of North Carolina at Chapel Hill, Chapel Hill, NC, USA
Joey V. Ragusa: Department of Cell Biology and Physiology, University of North Carolina at Chapel Hill, Chapel Hill, NC, USA
Angie Mordant: Proteomics Core Facility, Department of Pharmacology, University of North Carolina at Chapel Hill, Chapel Hill, NC, USA
Julie C. Necarsulmer: Department of Cell Biology and Physiology, University of North Carolina at Chapel Hill, Chapel Hill, NC, USA
Robert J. Perna: Department of Neurology, University of North Carolina at Chapel Hill, Chapel Hill, NC, USA
Tejazaditya Ajit: Department of Neurology, University of North Carolina at Chapel Hill, Chapel Hill, NC, USA
Kristen White: Microscopy Services Laboratory, Department of Pathology and Laboratory Medicine, University of North Carolina at Chapel Hill, Chapel Hill, NC, USA
Natalie K. Barker: Proteomics Core Facility, Department of Pharmacology, University of North Carolina at Chapel Hill, Chapel Hill, NC, USA
Xu Tian: Department of Neurology, University of North Carolina at Chapel Hill, Chapel Hill, NC, USA
Sarah Cohen: Department of Cell Biology and Physiology, University of North Carolina at Chapel Hill, Chapel Hill, NC, USA
Rick Meeker: Department of Neurology, University of North Carolina at Chapel Hill, Chapel Hill, NC, USA
Laura E. Herring: Proteomics Core Facility, Department of Pharmacology, University of North Carolina at Chapel Hill, Chapel Hill, NC, USA
Todd J. Cohen: Department of Cell Biology and Physiology, University of North Carolina at Chapel Hill, Chapel Hill, NC, USA; Department of Neurology, University of North Carolina at Chapel Hill, Chapel Hill, NC, USA; Department of Biochemistry and Biophysics, University of North Carolina at Chapel Hill, Chapel Hill, NC, USA; UNC Neuroscience Center, University of North Carolina at Chapel Hill, Chapel Hill, NC, USA; Corresponding author

Journal volume & issue: Vol. 26, no. 5
p. 106645

Abstract

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Summary: Transactive response DNA-binding protein of 43 kDa (TDP-43) is a highly conserved, ubiquitously expressed nucleic acid-binding protein that regulates DNA/RNA metabolism. Genetics and neuropathology studies have linked TDP-43 to several neuromuscular and neurological disorders including amyotrophic lateral sclerosis (ALS) and frontotemporal lobar degeneration (FTLD). Under pathological conditions, TDP-43 mislocalizes to the cytoplasm where it forms insoluble, hyper-phosphorylated aggregates during disease progression. Here, we optimized a scalable in vitro immuno-purification strategy referred to as tandem detergent-extraction and immunoprecipitation of proteinopathy (TDiP) to isolate TDP-43 aggregates that recapitulate those identified in postmortem ALS tissue. Moreover, we demonstrate that these purified aggregates can be utilized in biochemical, proteomics, and live-cell assays. This platform offers a rapid, accessible, and streamlined approach to study ALS disease mechanisms, while overcoming many limitations that have hampered TDP-43 disease modeling and therapeutic drug discovery efforts.

Published in iScience

ISSN: 2589-0042 (Online)
Publisher: Elsevier
Country of publisher: United States
LCC subjects: Science
Website: http://www.cell.com/iscience/home

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