Relaxation of Loaded ESCRT-III Spiral Springs Drives Membrane Deformation.
Bottom Line: We reasoned that Snf7 spirals could function as spiral springs.Furthermore, we observed that the elastic expansion of compressed Snf7 spirals generated an area difference between the two sides of the membrane and thus curvature.This spring-like activity underlies the driving force by which ESCRT-III could mediate membrane deformation and fission.
Affiliation: University of Geneva, Department of Biochemistry, quai Ernest Ansermet 30, 1211 Geneva 4, Switzerland.Show MeSH
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Mentions: To characterize the initial events leading to patch formation, we first studied the early steps of Snf7 patch nucleation by TIRFM, which allowed us to quantify the approximate number of Snf7 molecules in diffraction limited spots from their fluorescence intensity (see Experimental Procedures for quantification). Patch nucleation started with the appearance of a fluorescent diffraction limited spot (nucleus, Figure 4A) containing 50 ± 20 monomers (n = 9). At a Snf7 concentration of 300 nM, the intensity of the nuclei remained constant for several minutes until these nuclei started to grow (Figure 4A). Under these conditions, the number of nuclei is very low (Figure 1C). To increase the number of nuclei and to obtain more robust statistics, we nucleated Snf7 assemblies by adding 1 μM ESCRT-II and 1 μM Vps20 to a 75 nM Snf7 solution. Under these conditions, many Snf7 nuclei appeared on the membrane surface (Figure 4B and Figure S4A) and remained stable for several tens of minutes, consistent with our observations with Snf7 alone (see Figure 4A).
Affiliation: University of Geneva, Department of Biochemistry, quai Ernest Ansermet 30, 1211 Geneva 4, Switzerland.