Metabolic Labeling with an Alkyne-modified Isoprenoid Analog Facilitates Imaging and Quantification of the Prenylome in Cells

Protein prenylation is a post-translational modification that is responsible for membrane association and protein-protein interactions. The oncogenic protein Ras, which is prenylated, has been the subject of intense study in the past 20 years as a therapeutic target. Several studies have shown a correlation between neurodegenerative diseases including Alzheimer's disease and Parkinson's disease and protein prenylation. Here, a method for imaging and quantification of the prenylome using microscopy and flow cytometry is described. We show that metabolically incorporating an alkyne isoprenoid into mammalian cells, followed by a Cu(I)-catalyzed alkyne azide cycloaddition reaction to a fluorophore, allows for detection of prenylated proteins in several cell lines and that different cell types vary significantly in their levels of prenylated proteins. The addition of a prenyltransferase inhibitor or the precursors to the native isoprenoid substrates lowers the levels of labeled prenylated proteins. Finally, we demonstrate that there is a significantly higher (22%) level of prenylated proteins in a cellular model of compromised autophagy as compared to normal cells, supporting the hypothesis of a potential involvement of protein prenylation in abrogated autophagy. These results highlight the utility of total prenylome labeling for studies on the role of protein prenylation in various diseases including aging-related disorders.