There is thus a need for the development of reconstituted in vitro systems, in which the different components and physico-chemical parameters could be finely tuned, and ideally precisely ordered in space and time. To this aim, giant unilamellar vesicles (GUVs), which are spherical entities with a phospholipid bilayer membrane and a diameter (1-100 µm) comparable to that of actual cells, appear as particularly suitable minimal models to mimic living cells. Various methods have been developed to control their size, inner content and composition of their membrane (in lipids and proteins), leading to various relevant examples of synthetic cells, each of them mimicking, reproducing, or allowing to study one or several specific biological features. Hence, the lipid domain formation and organization has been studied using GUVs as models for lipid phase segregation. Indeed, mixtures of unsaturated and saturated lipids with cholesterol in proper proportions exhibit liquid-ordered (Lo) / liquid-disordered (Ld) phase separation. Controlling domain formation in GUVs, either by lipid stoichiometry or externally by temperature or light stimulus, is a well-established process, but the control over the domain spatial distribution remains a highly desired challenge, which has been partially achieved in only a few notable reports. For instance, microstructured substrates have been shown to direct the position of lipid domains at specific locations but this involved the use of supported bilayers and not GUVs. To our knowledge, spatial control over lipid domain position in GUVs has been only realized either on a single vesicle under pneumatic stress or on GUVs squeezed in pH-sensitive gels. The first study showed that, in a GUV compressed into a pancake shape between two walls, the lipid phase in minority tended to localize away from the flat parts in contact with the walls. Such an approach was however limited to a single compressed GUV and did not allow much flexibility in terms of GUV domain localization. In contrast, the second study allowed to impose a broad variety of user-defined shapes to a large number of constrained GUVs, but it required a specific pH-sensitive gel microenvironment and the control over domain distribution was not studied in detail.