Journal
NATURE COMMUNICATIONS
Volume 12, Issue 1, Pages -Publisher
NATURE PORTFOLIO
DOI: 10.1038/s41467-021-25212-3
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Funding
- Office of the Assistant Secretary of Defense for Health Affairs [W81XWH-16-1-0002]
- NSF [DGE1106400]
- Society of Lab Automation and Screening Graduate Education Fellowship
- NIH [1K99AG065200-01A1, R01CA203018, R01AG055891-01]
- NSF CAREER [CBET1056035]
- Howard Hughes Medical Institute
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This research introduces a novel method to simultaneously detect protein complexes in stressed cell populations, revealing potential cellular responses to specific treatments. The study also highlights the impact of non-chemical stress on cellular heterogeneity of F-actin. The assay developed in this work overcomes selectivity limitations to biochemically quantify single-cell protein complexes perturbed with diverse stimuli.
Multimeric cytoskeletal protein complexes orchestrate normal cellular function. However, protein-complex distributions in stressed, heterogeneous cell populations remain unknown. Cell staining and proximity-based methods have limited selectivity and/or sensitivity for endogenous multimeric protein-complex quantification from single cells. We introduce micro-arrayed, differential detergent fractionation to simultaneously detect protein complexes in hundreds of individual cells. Fractionation occurs by 60 s size-exclusion electrophoresis with protein complex-stabilizing buffer that minimizes depolymerization. Proteins are measured with a similar to 5-hour immunoassay. Co-detection of cytoskeletal protein complexes in U2OS cells treated with filamentous actin (F-actin) destabilizing Latrunculin A detects a unique subpopulation (similar to 2%) exhibiting downregulated F-actin, but upregulated microtubules. Thus, some cells may upregulate other cytoskeletal complexes to counteract the stress of Latrunculin A treatment. We also sought to understand the effect of non-chemical stress on cellular heterogeneity of F-actin. We find heat shock may dysregulate filamentous and globular actin correlation. In this work, our assay overcomes selectivity limitations to biochemically quantify single-cell protein complexes perturbed with diverse stimuli.
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