Playing with lipid packing and membrane curvature: regulation of protein-membrane interaction for the full-length lipid-packing sensor motif of ArfGAP1

ERNESTO AMBROGGIO; BENOÎT SORRE; GUILLAUME DRIN; JOELLE BIGAY; PATRICIA BASSEREAU; JEAN-BAPTISTE MANNEVILLE; BRUNO ANTONNY; BRUNO GOUD

Pavia-Italy

Congreso; The 2008 Golgi meeting: Membrane trafficking in global cellular responses; 2008

  The recruitment of coatomer (COPI) by the small G-protein Arf1 (GTP-bound form) at the Golgi generates highly-curved COPI-coated membrane buds, which give rise to transport vesicles. The GTPase activating protein for Arf1, ArfGAP1, has been shown to preferentially interact with highly curved membranes where it promotes fast GTP hydrolysis on Arf1. This adsorption to  highly-curved membranes is due to the presence of two adjacent lipid-packing sensor motifs, called ALPS 1 and ALPS 2, in the middle of the ArfGAP1 sequence. Here we have studied the interaction of a fluorescent-labeled version of the ALPS 1-ALPS 2 tandem with giant unilamellar vesicles (GUVs) by using fluorescence confocal microscopy. Different local conditions of lipid packing and curvature were imposed to the membrane either by pulling membrane tubes with molecular motors or with a set-up that couples micropipette aspiration and optical tweezers. Our results demonstrate that when regions of high and low curvature coexist within a membrane, ALPS 1-2-Alexa⁴⁸⁸ and ArfGAP1 bind exclusively to the highly curved region. A quantitative analysis indicates that 35 nm is the critical radius for ALPS binding to DOPC tubes. ArfGAP1 activity was studied using a fluorescent version of Arf1. Our data reflects that ArfGAP1 acts preferentially at the highly-curved region of the membrane and in accordance to the reaction-diffusion law. These results led us to propose a novel idea on the activity of ArfGAP1 at the COPI-mediated transport.