Cooperative extraction of membrane nanotubes by molecular motors

doi:10.1073/pnas.0406598101

Cooperative extraction of membrane nanotubes by molecular motors

^*Institut Curie, Unité Mixte de Recherche 168, 26 Rue d'Ulm, F-75248 Paris Cedex 05, France; ^‡Department d'Estructuria i Constituents de la Matèria, Universitat de Barcelona, Avinguda Diagonal, 647, E-08028 Barcelona, Spain; ^§Institut Curie, Unité Mixte de Recherche 144, 26 Rue d'Ulm, F-75248 Paris Cedex 05, France; ^¶Institut de Biologie de Lille, Department 5, Unité Mixte de Recherche 8525, Centre National de la Recherche Scientifique–Université de Lille 2, Campus Calmette, 1, Rue du Pr. Calmette, F-59021 Lille Cedex, France; and ^**Ecole Supérieure de Physique et de Chimie Industrielles, 10 Rue Vauquelin, F-75231 Paris Cedex 05, France

Edited by Tom C. Lubensky, University of Pennsylvania, Philadelphia, PA, and approved October 14, 2004 (received for review September 7, 2004)

Abstract

In eukaryotic cells, nanotubes represent a substantial fraction of transport intermediates between organelles. They are extracted from membranes by molecular motors walking along microtubules. We previously showed that kinesins fixed on giant unilamellar vesicles in contact with microtubules are sufficient to form nanotubes in vitro. Motors were attached to the membrane through beads, thus facilitating cooperative effects. Koster et al. [Koster, G., VanDuijn, M., Hofs, B. & Dogterom, M. (2003) Proc. Natl. Acad. Sci. USA 100, 15583–15588] proposed that motors could dynamically cluster at the tip of tubes when they are individually attached to the membrane. We demonstrate, in a recently designed experimental system, the existence of an accumulation of motors allowing tube extraction. We determine the motor density along a tube by using fluorescence intensity measurements. We also perform a theoretical analysis describing the dynamics of motors and tube growth. The only adjustable parameter is the motor binding rate onto microtubules, which we measure to be 4.7 ± 2.4 s^–1. In addition, we quantitatively determine, for a given membrane tension, the existence of a threshold in motor density on the vesicle above which nanotubes can be formed. We find that the number of motors pulling a tube can range from four at threshold to a few tens away from it. The threshold in motor density (or in membrane tension at constant motor density) could be important for the understanding of membrane traffic regulation in cells.

Footnotes

↵ ∥ To whom correspondence may be addressed. E-mail: jean-francois.joanny{at}curie.fr or patricia.bassereau{at}curie.fr.
↵ † C.L. and O.C. contributed equally to this work.
Author contributions: C.L., O.C., A.R., B.G., J.-F.J., P.B., and J.P. designed research; C.L., O.C., J.-F.J., P.B., and J.P. performed research; C.L. analyzed data; K.Z., P.J., and L.B.-B. contributed new reagents or analytic tools; C.L., O.C., J.-F.J., P.B., and J.P. wrote the paper; and O.C., J.-F.J., and J.P. provided theoretical descriptions.
This paper was submitted directly (Track II) to the PNAS office.
Abbreviations: MT, microtubule; BODIPY PC, β-BODIPY 530/550 C₅-hexadecanoyl phosphatidylcholine; DHPE-Biot-Rhod, rhodamin–biotin dihexadecanoyl-phosphatidylethanolamine; GUV, giant unilamellar vesicle.