We recently reported that the isoprenylation and palmitoylation motif present at the C-terminus of human RhoB protein promotes intraluminal vesicle delivery of proteins in cells from organisms as phylogenetically apart as fungi and humans. Here we build on these observations by showing that chimeras of fluorescent proteins bearing this sequence, namely, CINCCKVL, which become isoprenylated and palmitoylated in cells, may be used to mark endolysosomes while preserving their morphology. Indeed, these chimeric proteins are devoid of the effects derived from overexpression of fluorescent constructs of full-length, active proteins widely used as endolysosomal markers, such as Lamp1 or Rab7, which cause lysosomal enlargement, or RhoB, which induces actin stress fibers. Moreover, the fact that lipidation-dependent endolysosomal localization of CINCCKVL chimeras can be ascertained in a wide variety of cells indicates that they follow a path toward endolysosomes that is conserved in diverse species. Therefore, CINCCKVL chimeras serve as robust tools to mark these late endocytic compartments.
Keywords: Rho proteins; fluorescent markers/proteins; isoprenylation; lysosomes/endolysosomes; multivesicular bodies; organelle probe; palmitoylation; subcellular localization.