RESUMO
Controversy has long surrounded the question of whether spontaneous lateral demixing of membranes into coexisting liquid phases can organize proteins and lipids on micron scales within unperturbed, living cells. A clear answer hinges on observation of hallmarks of a reversible phase transition. Here, by directly imaging micron-scale membrane domains of yeast vacuoles both in vivo and cell free, we demonstrate that the domains arise through a phase separation mechanism. The domains are large, have smooth boundaries, and can merge quickly, consistent with fluid phases. Moreover, the domains disappear above a distinct miscibility transition temperature (Tmix) and reappear below Tmix, over multiple heating and cooling cycles. Hence, large-scale membrane organization in living cells under physiologically relevant conditions can be controlled by tuning a single thermodynamic parameter.