PreprintCited by 16Cellular Mechanics and InteractionsAdvanced Proteomics Techniques and ApplicationsMicrotubule and mitosis dynamics

Measuring expression heterogeneity of single-cell cytoskeletal protein complexes

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Vlassakis, Julea, Hansen, Louise L., Higuchi-Sanabria, Ryo, Zhou, Yun, Tsui, C. Kimberly, Dillin, Andrew, Huang, Haiyan, Herr, Amy E.·2020·Nature Communications

Preprint — published as 10.1038/s41467-021-25212-3 · Paper page

Abstract

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 100s of individual cells. Fractionation occurs by 60 s size-exclusion electrophoresis with protein complex-stabilizing buffer that minimizes depolymerization. Co-detection of cytoskeletal protein complexes in U2OS cells treated with filamentous actin (F-actin) destabilizing LatA detects a subpopulation (~11%) exhibiting downregulated F-actin, but upregulated microtubule and intermediate filament protein complexes. Thus, some cells upregulate other cytoskeletal complexes to counteract the stress of LatA treatment. We also sought to understand the effect of non-chemical stress on cellular heterogeneity of F-actin. We find heat shock dysregulates F- and G-actin correlation. The assay overcomes selectivity limitations to biochemically quantify single-cell protein complexes perturbed with diverse stimuli.

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Authors
Vlassakis, J., Hansen, L. L., Higuchi-Sanabria, R., Zhou, Y., Tsui, C. K., Dillin, A., Huang, H., & Herr, A. E.
Affiliations
  • University of California, Berkeley (US)
  • Howard Hughes Medical Institute (US)
Corresponding author
Herr, Amy E.
Venue
Nature Communications
Publisher
openRxiv
Published
2020-09-13
Volume
12
Issue
1
Pages
4969-4969
ISSN
2041-1723 (ISSN-L)
OpenAlex
W3194320991
Cited by
16
References
113
Citations by year
2026: 2 · 2025: 5 · 2024: 2 · 2023: 5 · 2022: 1 · 2021: 1
FWCI
1.17
Type
journal article
Language
en
Open access
Locations
  • Nature Communications (publishedVersion, cc-by)
  • bioRxiv (Cold Spring Harbor Laboratory) (acceptedVersion)
  • PubMed Central (publishedVersion, cc-by)
  • PubMed (publishedVersion)
  • RePEc: Research Papers in Economics (submittedVersion)
  • DOAJ (DOAJ: Directory of Open Access Journals) (submittedVersion, cc-by-sa)
  • eScholarship (California Digital Library) (submittedVersion)
  • eScholarship (California Digital Library) (submittedVersion, cc-by)
Funders
  • National Science Foundation (1056035, 1106400, CAREER)
  • U.S. Department of Defense (W81XWH-16, W81XWH, W81XWH1610002)
  • Howard Hughes Medical Institute
  • U.S. Department of Health and Human Services (1K99AG065200-01A1, R01CA203018, R01AG055891-01)
  • National Institutes of Health (MDA-MB-231, W81XWH, NIH K99)
  • National Institute on Aging (K99AG065200, R01AG055891)
  • National Cancer Institute (R01CA203018)
  • Division of Graduate Education (DGE‐1106400)
  • Division of Chemical, Bioengineering, Environmental, and Transport Systems (CBET-1056035)
Topics
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Cell Biology › Cellular Mechanics and Interactions
  • Physical Sciences › Chemistry › Spectroscopy › Advanced Proteomics Techniques and Applications
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Cell Biology › Microtubule and mitosis dynamics
Keywords
single-cell heterogeneity, F-actin, microtubules, latrunculin A, heat shock, U2OS cells
MeSH terms
Actins, Animals, Cell Differentiation, Cell Line, Cytoskeletal Proteins, Cytoskeleton, Humans, Microtubules, Models, Biological, Genetic Heterogeneity, Heat-Shock Response, Bridged Bicyclo Compounds, Heterocyclic, Thiazolidines, Single-Cell Analysis

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