An amphipathic helix enables septins to sense micron-scale membrane curvature

Abstract

The geometry of cells is well described by membrane curvature. Septins are filament forming, GTP-binding proteins that assemble on positive, micrometer-scale curvatures. Here, we examine the molecular basis of curvature sensing by septins. We show that differences in affinity and the number of binding sites drive curvature-specific adsorption of septins. Moreover, we find septin assembly onto curved membranes is cooperative and show that geometry influences higher-order arrangement of septin filaments. Although septins must form polymers to stay associated with membranes, septin filaments do not have to span micrometers in length to sense curvature, as we find that single septin complexes have curvature-dependent association rates.We trace this ability to an amphipathic helix (AH) located on the C-terminus of Cdc12. The AH domain is necessary and sufficient for curvature sensing both in vitro and in vivo. These data show that curvature sensing by septins operates at much smaller length scales than the micrometer curvatures being detected. We thank the Gladfelter lab and Danny Lew for useful discussions, Matthias Garten for ideas in setting up the rod assay, the UNC EM facility (Victoria Madden and Kristen White) for support with SEM and HHMI Faculty Scholars award to ASG. Author contributionsKSC, BLW conducted experiments, BLW and JMC constructed strains for experiments, BLW and JMC constructed plasmids for experiments, KSC, BLW, and ASG designed experiments.

Septin 9-containging filaments and Golgi assembly depend on two polybasic domains

“…Despite that our in vitro studies were done with the monomeric septin 9, our results give information on how septin 9 membrane binding determinants might influence the localization of septin oligomers to membranes. These results are consistent with a recent report of the presence of AHs structures in oligomeric septin filaments able to sense macroscopic curvatures (Cannon et al, 2018).…”

The long and short of membrane curvature sensing by septins

“…The polybasic motifs turned out to be important for septin polymerization, and it was the AHs that mediated membrane association. Yet another recent study (7) exploited synthetic vesicles and "wavy" membrane surfaces to come to essentially the same conclusions as Cannon et al (1) in terms of yeast septin filament length/orientation/packing and curvature preference.…”

“…Remarkably, a mutant truncating this sequence was isolated decades ago in an unbiased screen and represents a commonly used genetic tool, since in the mutant cells higher-order septin assemblies disappear rapidly upon a shift to high temperature, but the molecular basis was unknown. The Cdc12 AH was found to be both necessary and sufficient for curvature preference in vitro, and when the authors made an equivalent truncation mutation in the Ashbya homologue of Cdc12, septin association with curved membranes in vivo was drastically reduced (1).…”

Transfection of choanoflagellates illuminates their cell biology and the ancestry of animal septins

“…, 2010) for membranes that have distinct geometries (Bridges et al. , 2016; Cannon et al. , 2018) and lipid compositions (Casamayor and Snyder, 2003; Tanaka-Takiguchi et al.…”