Farnesyltransferase inhibitors disrupt EGF receptor traffic through modulation of the RhoB GTPase

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Abstract
The Rho family of small GTPases play a pivotal role in the dynamic regulation of the actin cytoskeleton. Recent studies have suggested that these signalling proteins also have wide-ranging functions in membrane trafficking pathways. The Rho family member RhoB was shown to localise to vesicles of the endocytic compartment, suggesting a potential function in regulation of endocytic traffic. In keeping with this, we have previously shown that expression of active RhoB causes a delay in the intracellular trafficking of the epidermal growth factor (EGF) receptor; however, the site of action of RhoB within the endocytic pathway is still unknown. RhoB exists as two prenylated forms in cells: geranylgeranylated RhoB (RhoB-GG) and farnesylated RhoB (RhoB-F). Here we use farnesyltransferase inhibitors (FTIs) to show that prenylation specifies the cellular localisation of RhoB. RhoB-GG localises to multivesicular late endosomes and farnesylated RhoB (RhoB-F) localises to the plasma membrane. The gain of endosomal RhoB-GG elicited by FTI treatment reduces sorting of EGF receptor to the lysosome and increases recycling to the plasma membrane. Ultrastructural analysis shows that activation of RhoB through drug treatment or mutation has no effect the sorting of receptor into late endosomes, but instead inhibits the subsequent transfer of late endosomal receptor to the lysosome.