PIP2 reshapes membranes through asymmetric desorption

Sankalp Shukla, Rui Jin, Jaclyn Robustelli, Zachary E. Zimmerman, Tobias Baumgart

Phosphatidylinositol-4,5-bisphosphate [PIP2] is an important signaling lipid in eukaryotic cell plasma membranes, playing an essential role in diverse cellular processes. The headgroup of PIP2 is highly negatively charged and this lipid displays a high critical micellar concentration compared to housekeeping phospholipid analogs. Given the crucial role of PIP2, it is imperative to study its localization, interaction with proteins, and membrane shaping properties. Biomimetic membranes have served extensively to elucidate aspects including protein-lipid and lipid-lipid interactions, as well as membrane mechanics. Incorporation of PIP2 into biomimetic membranes, however, has at times resulted in discrepant findings described in the literature. With the goal to elucidate the mechanical consequences of PIP2 incorporation, we studied the desorption of PIP2 from biomimetic giant unilamellar vesicles [GUVs] by means of a fluorescent marker. A decrease in fluorescence intensity with the age of the vesicles suggested that PIP2 lipids were being desorbed from the outer leaflet of the membrane. To evaluate whether this desorption was asymmetric, the vesicles were systematically diluted. This resulted in an increase in the number of internally tubulated vesicles within minutes after dilution, suggesting that the desorption was asymmetric and also generated membrane curvature. By means of a saturated chain homolog of PIP2, we showed that the fast desorption of PIP2 is facilitated by presence of an arachidonic lipid tail and possibly due to its oxidation. Through measurements of the pulling force of membrane tethers, we quantified the effect of this asymmetric desorption on the spontaneous membrane curvature. Furthermore, we found that the spontaneous curvature could be modulated by externally increasing the concentration of PIP2 micelles. Given that the local concentration of PIP2 in biological membranes is variable, spontaneous curvature generated by PIP2 may affect the formation of highly curved structures which can serve as initiators for signaling events.

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