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

Charuta C. Palsuledesai, Joshua D. Ochocki, Michelle M. Kuhns, Yen Chih Wang, Janel K. Warmka, Dustin S. Chernick, Elizabeth V. Wattenberg, Ling Li, Edgar A. Arriaga, Mark D. Distefano

Research output: Contribution to journalArticlepeer-review

20 Scopus citations

Abstract

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.

Original languageEnglish (US)
Pages (from-to)2820-2828
Number of pages9
JournalACS Chemical Biology
Volume11
Issue number10
DOIs
StatePublished - Oct 21 2016

Bibliographical note

Funding Information:
This work was supported by the National Institutes of Health grants R01 GM058842 and R01 GM084152 (M.D.D.), R01 AG020866 (E.A.A.), R01 AG031846 (L.L.), T32 GM08700 (M.M.K.), and T32 GM008347 (J.D.O.) as well as by the National Science Foundation grant CHE-1308655 (M.D.D.).

Publisher Copyright:
© 2016 American Chemical Society.

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