DISTINCT STRUCTURAL ELEMENTS OF RAB5 DEFINE ITS FUNCTIONAL SPECIFICITY

Members of the rab family of small GTPases are localized to distinct c ellular compartments and function as specific regulators of vesicle tr ansport between organelles. Overexpression of rab5, which is associate d with early endosomes and the plasma membrane, increases the rate of endocytosis [Bucci et al. (1992) Cell, 70, 715 - 728]. From sequence a lignments and molecular modelling we identified structural elements th at might contribute to the definition of the functional specificity of rab5. To test the role of these elements experimentally, we transplan ted them onto rab6, which is associated with the Golgi complex. The ch imeric proteins were assayed for intracellular localization and stimul ation of endocytosis. First, we found that the C-terminus of rab5 coul d target rab6 to the plasma membrane and early endosomes but it did no t confer rab5-like stimulation of endocytosis. Further replacement of other regions revealed that the N-terminus, helix alpha2/loop 5 and he lix alpha3/loop 7 were all required to functionally convert rab6 into rab5. Reciprocal hybrids of rab5 containing these regions replaced wit h those of rab6 were inactive, demonstrating that each region is essen tial for rab5 function. These results indicate that distinct structura l elements specify the localization, membrane association and regulato ry function of rab5.