During the first half of the project, we made significant progress on the mechanism underlying the formation of lipid gradients. We found that membrane contact sites are critical sites for the establishment of lipid gradients. This is true at the level of the cell but also at the tissue scale, which we did not anticipate initially. We also found that anionic lipid feedback on the formation of membrane contact sites, thereby regulating the establishment of their own patterns. Using a microfluidic based system and high-resolution time-lapse imaging, we uncovered that not only auxin, but also regulatory peptides, can rapidly impact anionic lipids subcellular accumulation. Finally, we analyzed the function of anionic lipids in the formation of signaling nanodomains at the plasma membrane and their impact on signal specificity. Here, we found that different anionic lipids are critical for the formation of Rho GTPase-containing nanodomains. However, we also uncovered the existence of anionic lipid-independent mechanisms that we will investigate during the second reporting period.