PUBLICATION
Genetic, cellular and structural characterization of the membrane potential-dependent cell-penetrating peptide translocation pore
- Authors
- Trofimenko, E., Grasso, G., Heulot, M., Chevalier, N., Deriu, M.A., Dubuis, G., Arribat, Y., Serulla, M., Michel, S., Vantomme, G., Ory, F., Dam, L.C., Puyal, J., Amati, F., Lüthi, A., Danani, A., Widmann, C.
- ID
- ZDB-PUB-211030-9
- Date
- 2021
- Source
- eLIFE 10: (Journal)
- Registered Authors
- Amati, Francesca, Arribat, Yoan
- Keywords
- cell biology, mouse, zebrafish
- MeSH Terms
-
- Animals
- Cell Line
- Cell-Penetrating Peptides/chemistry
- Cell-Penetrating Peptides/genetics*
- Cell-Penetrating Peptides/metabolism
- HeLa Cells
- Humans
- Membrane Potentials
- Mice
- Mice, Inbred C57BL
- Potassium Channels/genetics*
- Potassium Channels/metabolism
- Protein Transport
- Rats
- Rats, Sprague-Dawley
- Zebrafish
- PubMed
- 34713805 Full text @ Elife
Citation
Trofimenko, E., Grasso, G., Heulot, M., Chevalier, N., Deriu, M.A., Dubuis, G., Arribat, Y., Serulla, M., Michel, S., Vantomme, G., Ory, F., Dam, L.C., Puyal, J., Amati, F., Lüthi, A., Danani, A., Widmann, C. (2021) Genetic, cellular and structural characterization of the membrane potential-dependent cell-penetrating peptide translocation pore. eLIFE. 10:.
Abstract
Cell-penetrating peptides (CPPs) allow intracellular delivery of bioactive cargo molecules. The mechanisms allowing CPPs to enter cells are ill-defined. Using a CRISPR/Cas9-based screening, we discovered that KCNQ5, KCNN4, and KCNK5 potassium channels positively modulate cationic CPP direct translocation into cells by decreasing the transmembrane potential (Vm). These findings provide the first unbiased genetic validation of the role of Vm in CPP translocation in cells. In silico modeling and live cell experiments indicate that CPPs, by bringing positive charges on the outer surface of the plasma membrane, decrease the Vm to very low values (-150 mV or less), a situation we have coined megapolarization that then triggers formation of water pores used by CPPs to enter cells. Megapolarization lowers the free energy barrier associated with CPP membrane translocation. Using dyes of varying dimensions in CPP co-entry experiments, the diameter of the water pores in living cells was estimated to be 2(-5) nm, in accordance with the structural characteristics of the pores predicted by in silico modeling. Pharmacological manipulation to lower transmembrane potential boosted CPPs cellular internalization in zebrafish and mouse models. Besides identifying the first proteins that regulate CPP translocation, this work characterized key mechanistic steps used by CPPs to cross cellular membrane. This opens the ground for strategies aimed at improving the ability of cells to capture CPP-linked cargos in vitro and in vivo.
Genes / Markers
Expression
Phenotype
Mutations / Transgenics
Human Disease / Model
Sequence Targeting Reagents
Fish
Orthology
Engineered Foreign Genes
Mapping