High-throughput identification of calcium-regulated proteins across diverse proteomes
Timothy M. Locke,
Rose Fields,
Hayden Gizinski,
George M. Otto,
Melissa J.S. MacEwen,
Domnita-Valeria Rusnac,
Peixian He,
David M. Shechner,
Chris D. McGann,
Matthew D. Berg,
Judit Villen,
Yasemin Sancak,
Devin K. Schweppe
Affiliations
Timothy M. Locke
Department of Pharmacology, University of Washington, Seattle, WA 98195, USA
Rose Fields
Department of Genome Sciences, University of Washington, Seattle, WA 98195, USA; Brotman Baty Institute for Precision Medicine, Seattle, WA, USA; Institute of Stem Cell and Regenerative Medicine, University of Washington, Seattle, WA, USA
Hayden Gizinski
Department of Pharmacology, University of Washington, Seattle, WA 98195, USA
George M. Otto
Department of Pharmacology, University of Washington, Seattle, WA 98195, USA
Melissa J.S. MacEwen
Department of Pharmacology, University of Washington, Seattle, WA 98195, USA
Domnita-Valeria Rusnac
Department of Pharmacology, University of Washington, Seattle, WA 98195, USA; Howard Hughes Medical Institute, Department of Pharmacology, University of Washington, Seattle, WA 98195, USA
Peixian He
Department of Pharmacology, University of Washington, Seattle, WA 98195, USA
David M. Shechner
Department of Pharmacology, University of Washington, Seattle, WA 98195, USA
Chris D. McGann
Department of Genome Sciences, University of Washington, Seattle, WA 98195, USA; Brotman Baty Institute for Precision Medicine, Seattle, WA, USA; Institute of Stem Cell and Regenerative Medicine, University of Washington, Seattle, WA, USA
Matthew D. Berg
Department of Genome Sciences, University of Washington, Seattle, WA 98195, USA
Judit Villen
Department of Genome Sciences, University of Washington, Seattle, WA 98195, USA
Yasemin Sancak
Department of Pharmacology, University of Washington, Seattle, WA 98195, USA; Corresponding author
Devin K. Schweppe
Department of Genome Sciences, University of Washington, Seattle, WA 98195, USA; Brotman Baty Institute for Precision Medicine, Seattle, WA, USA; Institute of Stem Cell and Regenerative Medicine, University of Washington, Seattle, WA, USA; Corresponding author
Summary: Calcium ions play important roles in nearly every biological process, yet whole-proteome analysis of calcium effectors has been hindered by a lack of high-throughput, unbiased, and quantitative methods to identify protein-calcium engagement. To address this, we adapted protein thermostability assays in budding yeast, human cells, and mouse mitochondria. Based on calcium-dependent thermostability, we identified 2,884 putative calcium-regulated proteins across human, mouse, and yeast proteomes. These data revealed calcium engagement of signaling hubs and cellular processes, including metabolic enzymes and the spliceosome. Cross-species comparison of calcium-protein engagement and mutagenesis experiments identified residue-specific cation engagement, even within well-known EF-hand domains. Additionally, we found that the dienoyl-coenzyme A (CoA) reductase DECR1 binds calcium at physiologically relevant concentrations with substrate-specific affinity, suggesting direct calcium regulation of mitochondrial fatty acid oxidation. These discovery-based proteomic analyses of calcium effectors establish a key resource to dissect cation engagement and its mechanistic effects across multiple species and diverse biological processes.
