Repeated FRAP of the actin-binding protein CapG in the cell nucleus-a functional assay for EGF signaling in the single live breast cancer cell.
Journal
Scientific reports
ISSN: 2045-2322
Titre abrégé: Sci Rep
Pays: England
ID NLM: 101563288
Informations de publication
Date de publication:
05 Oct 2024
05 Oct 2024
Historique:
received:
19
10
2023
accepted:
23
09
2024
medline:
6
10
2024
pubmed:
6
10
2024
entrez:
5
10
2024
Statut:
epublish
Résumé
Compartmentalization and differential distribution of proteins within a cell maintain cellular function and viability. CapG is a gelsolin-related actin-binding protein that distributes in steady state diffusively throughout cytoplasm and cell nucleus. To detect changes in CapG's nucleocytoplasmic shuttling in response to external stimuli on the single cell level, we established repeated FRAP experiments of one and the same breast cancer cell. With this experimental set up, we found that ATP-depletion reversibly decreased CapG's shuttling into the cell nucleus. The addition of epidermal growth factor (EGF) increased CapG's nuclear shuttling within minutes. Serum-starvation doubled the number of breast cancer cells from 40 to 80% displaying increased CapG shuttling in response to EGF. Testing five different potential CapG phosphorylation sites, we found that serine 70 mediates the increase in CapG's nuclear shuttling triggered by EGF. Thus, repeated FRAP of CapG in the cell nucleus can be used as functional readout of signaling cascades in the same single live breast cancer cell.
Identifiants
pubmed: 39369027
doi: 10.1038/s41598-024-73887-7
pii: 10.1038/s41598-024-73887-7
doi:
Substances chimiques
Epidermal Growth Factor
62229-50-9
Microfilament Proteins
0
CAPG protein, human
148412-71-9
Nuclear Proteins
0
Types de publication
Journal Article
Langues
eng
Sous-ensembles de citation
IM
Pagination
23159Informations de copyright
© 2024. The Author(s).
Références
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