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We used pharmacological perturbations that alter fatty acid chain saturation to manipulate lipid packing and found septin structures were selectively disrupted at flat regions of the plasma membrane. To determine whether lipid packing is sufficient to impact septin assembly, molecular dynamics simulations were used to design lipid mixtures with varied packing to monitor septin adsorption in vitro. Septins strongly favored loosely packed lipid bilayers, but additional geometrical cues act in conjunction with this membrane property. 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