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PLoS Biol. 2018 Aug 27;16(8):e2004624. doi: 10.1371/journal.pbio.2004624. eCollection 2018 Aug.

A high-throughput screen of real-time ATP levels in individual cells reveals mechanisms of energy failure.

Author information

1
Gladstone Institute of Neurological Disease, San Francisco, California, United States of America.
2
Department of Pediatrics, University of California, San Francisco, California, United States of America.
3
Department of Radiation Oncology, University of California, San Francisco, California, United States of America.
4
Graduate Program in Biomedical Sciences, University of California, San Francisco, California, United States of America.
5
Department of Cellular and Molecular Pharmacology, University of California, San Francisco, California, United States of America.
6
Department of Urology, University of California, San Francisco, California, United States of America.
7
Helen Diller Family Comprehensive Cancer Center, University of California, San Francisco, California, United States of America.
8
Department of Laboratory Medicine, University of California, San Francisco, California, United States of America.
9
Department of Biochemistry and Biophysics and Institute for Neurodegenerative Diseases, University of California, San Francisco, California, United States of America.
10
Chan Zuckerberg Biohub, San Francisco, California, United States of America.
11
Department of Neurology, University of California, San Francisco, California, United States of America.
12
Graduate Program in Neuroscience, University of California, San Francisco, California, United States of America.

Abstract

Insufficient or dysregulated energy metabolism may underlie diverse inherited and degenerative diseases, cancer, and even aging itself. ATP is the central energy carrier in cells, but critical pathways for regulating ATP levels are not systematically understood. We combined a pooled clustered regularly interspaced short palindromic repeats interference (CRISPRi) library enriched for mitochondrial genes, a fluorescent biosensor, and fluorescence-activated cell sorting (FACS) in a high-throughput genetic screen to assay ATP concentrations in live human cells. We identified genes not known to be involved in energy metabolism. Most mitochondrial ribosomal proteins are essential in maintaining ATP levels under respiratory conditions, and impaired respiration predicts poor growth. We also identified genes for which coenzyme Q10 (CoQ10) supplementation rescued ATP deficits caused by knockdown. These included CoQ10 biosynthetic genes associated with human disease and a subset of genes not linked to CoQ10 biosynthesis, indicating that increasing CoQ10 can preserve ATP in specific genetic contexts. This screening paradigm reveals mechanisms of metabolic control and genetic defects responsive to energy-based therapies.

PMID:
30148842
PMCID:
PMC6110572
DOI:
10.1371/journal.pbio.2004624
[Indexed for MEDLINE]
Free PMC Article

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