Temperature, lipid charge and septin dependence of lateral lipid diffusion in biomimetic membrane systems

Abstract

Plasma membrane dynamics have become increasingly of interest due to recent discoveries on anomalous lipid diffusion in live cells. A specific characteristic has been found in the exponential relation between lateral lipid diffusion and temperature. The finding was reported to be surprising as a linear dependence was theoretically predicted. In this study, the gap between simple theoretical models and experimental in vivo results on lipid diffusion is investigated by studying model membrane systems experimentally. Specifically, lateral lipid diffusion is further characterized as function of temperature, lipid charge and asymmetry. To induce anomalous diffusion, a peripheral protein known to act as a diffusion barrier, fly-septin, was associated to the lipid bilayer to explore its effect on lipid slow-down.

Lateral lipid diffusion was measured by fluorescence recovery after photobleaching (FRAP) on a bare, or with associated septin, model phospholipid membrane system. The supported lipid bilayer (SLB) model membrane systems were formed by vesicle fusion (VF) or Langmuir-Blodgett/vesicle fusion (LBVF), specifically for asymmetric
SLBs. Temperature variations were induced by using an in-house built sample heater. The quality of the lipid bilayers was defined by investigating influencing variables; lipid batches, substrate surface treatment, SLB formation methods and lipid tracers.

Results show that membrane viscosity exhibits linear scaling to lipid diffusion. Furthermore, lateral lipid diffusion appears to be charge dependent and influenced by lipid composition of both leaflets. Considering the influence of septin, lateral lipid diffusion is reproducibly increased as compared to a bare membrane, in contrast to the reported septin induced lipid slow-down due to a diffusion barrier mechanism. These findings have brought the gap between theoretical models and membrane dynamics one step closer together.