Actin-cytoskeleton rearrangement modulates proton-induced uptake

Area of Science:

• Cell biology
• Biophysics

Background:

• Elevating proton concentration at the cell surface stimulates membrane invaginations and vesicle formation, enhancing macromolecule uptake.
• The precise mechanisms of vesicle formation and scission in proton-induced uptake (PIU) remain unclear.
• Actin's role in vesicle formation during endocytosis suggests its involvement in PIU.

Purpose of the Study:

• To investigate the involvement of cytoskeletal actin in proton-induced uptake (PIU).
• To understand how actin influences membrane vesiculation and macromolecule uptake under altered proton concentrations.

Main Methods:

• Utilized dextran-FITC uptake as a quantitative measure of PIU, reflecting vesicle scission from the plasma membrane.
• Compared PIU rates in suspended versus adherent cells to assess the impact of cell attachment and cytoskeletal organization.
• Employed pharmacological agents (wortmannin, calyculin-A, cytochalasin-D, rapamycin, latrunculin-A) to modulate actin dynamics and evaluate their effects on PIU.

Main Results:

• PIU rate was constant in suspended cells but gradually increased over time in adherent cells, eventually reaching levels seen in suspended cells.
• In adherent cells, PIU increased by 25% with wortmannin and calyculin-A, suggesting a role for specific signaling pathways.
• Cytochalasin-D, rapamycin, and latrunculin-A elevated PIU in both cell types, indicating broader actin involvement, while high latrunculin-A concentrations inhibited PIU via extensive actin depolymerization.

Conclusions:

• Proton-induced membrane vesiculation is constrained by the structural resistance of actin to plasma membrane bending.
• A degree of cortical actin restructuring is necessary for the successful completion of the scission process in PIU.
• Actin dynamics play a crucial, albeit complex, role in regulating proton-induced macromolecule uptake via membrane vesiculation.