A non-linear system patterns Rab5 GTPase on the membrane

Proteins can self-organize into spatial patterns via non-linear dynamic interactions on cellular membranes. Modelling and simulations have shown that small GTPases can generate patterns by coupling guanine nucleotide exchange factors (GEF) to effectors, generating a positive feedback of GTPase activation and membrane recruitment. Here, we reconstituted the patterning of the small GTPase Rab5 and its GEF/effector complex Rabex5/Rabaptin5 on supported lipid bilayers. We demonstrate a ‘handover’ of Rab5 from Rabex5 to Rabaptin5 upon nucleotide exchange. A minimal system consisting of Rab5, RabGDI and a complex of full length Rabex5/Rabaptin5 was necessary to pattern Rab5 into membrane domains. Rab5 patterning required a lipid membrane composition mimicking that of early endosomes, with PI(3)P enhancing membrane recruitment of Rab5 and acyl chain packing being critical for domain formation. The prevalence of GEF/effector coupling in nature suggests a possible universal system for small GTPase patterning involving both protein and lipid interactions.