Abstract Biological membranes are highly dynamic notably due to the constant exchange of vesicles between the different compartments of the cell. In addition, the dynamic nature of membranes is also caused by their inherently fluid properties, with the diffusion of both proteins and lipids within their leaflets. Lipid diffusion is particularly difficult to study in vivo but recent advances in optical microscopy and lipid visualization now enable the characterization of lipid lateral motion. Here, we review the methods that enable the characterization of lipid lateral diffusion in plants. We will then discuss the parameters that impact lipid diffusion in membranes and explore their consequences on the formation of membrane domains at different scales. Finally, we will address how controlled lipid diffusion impact membrane functions during cell signaling, development and environmental interactions.