Isoprenylation of Monomeric GTPases in Human Trabecular Meshwork Cells

Small monomeric GTPases, including those belonging to the Rho family, regulate a diverse array of intracellular signaling pathways which affect vesicle transport/trafficking, endocytosis, cell cycle progression, cell contractility, and formation of stress fibers or focal adhesions. Functional activation of newly synthesized small monomeric GTPases is facilitated by a multistep post-translational process involving transferase-catalyzed addition of farnesyl or geranylgeranyl isoprenoids to conserved cysteine residues within a unique carboxy terminal CaaX motif. Here, using well-established and widely available contemporary methodologies, detailed protocols by which to semi-quantitatively evaluate the functional consequence of post-translational isoprenylation in human trabecular meshwork cells are described. We introduce the concept that isoprenylation alone is itself a key regulator of mammalian Rho GTPase expression and turnover.