Physics, biology and the right chemistry

Abstract

Joint studies that involve biologists and physicists are becoming more frequent and have contributed to the identification and understanding of physical parameters underlying key biological processes. Here, we illustrate the main findings resulting from a 10-year collaboration between a cell biologist and an experimental physicist, both interested in the mechanisms of intracellular transport and membrane dynamics in eukaryotic cells.

Figure 1.. Formation of membrane nanotubes from giant unilamellar vesicles (GUVs) by molecular motors

Membrane tubes similar to those involved in intracellular transport can be pulled by kinesin motor proteins bound to GUVs that move along immobilized microtubules in the presence of ATP. The kinesins can be bound either via small streptavidin-coated polystyrene beads (top) or via streptavidin molecules associated with the lipid bilayer itself (bottom).

Figure 2.. Confocal image of a network of nanotubes formed from a giant unilamellar vesicle (GUV) containing fluorescent lipids and pulled by kinesin motors

Image courtesy of C. Leduc.

Figure 3.. General scheme of the experimental system used to study forces on a membrane

A membrane nanotube is pulled from a giant unilamellar vesicle (GUV) aspirated in a micropipette (left). A bead (orange sphere) trapped in optical tweezers is attached to the GUV. The tube is formed by pulling the micropipette away from the GUV. At equilibrium, the force required to pull a tube is calculated from the bead displacement and from the tweezers’ stiffness calibration.

Figure 4.. Creation of a gradient of ADP-ribosylation factor 1 (Arf1) along a membrane tube connected to a giant unilamellar vesicle (GUV)

A gradient of Arf1 molecules (green fluorescent label) is created along a membrane tube containing lipids (red fluorescent label) pulled using a bead (large green sphere, right) trapped in an optical tweezers. The gradient is due to the competition between diffusion of Arf1-GTP from the giant unilamellar vesicle (GUV) on the left into the pulled membrane tube (green arrows) and the dissociation of Arf1-GDP induced by ArfGAP1 hydrolysis of Arf1-GTP, which occurs in the tube because of its high curvature. The low curvature of the GUV membrane prevents ArfGAP1 binding and protects Arf1-GTP from hydrolysis.

Figure 5.. Proposed model for the association and dissociation of COPI coatomer in vivo

Coat proteins are recruited to the site of vesicle budding by membrane-bound Arf1 in its GTP form, and begin to deform the donor membrane. Sensing membrane curvature, ArfGAP1 is recruited to the budding site where it hydrolyzes GTP bound to Arf1, which then dissociates. As long as the budding site is attached to the donor membrane, the GTP form of Arf1 is replenished at the budding site. Once dissociated, the new vesicle lacks a fresh supply of Arf1-GTP. After all the Arf1-GTP has been hydrolyzed by ArfGAP1, the COPI coat dissociates from the newly formed transport vesicle or tubule.