Labeling of Phosphatidylinositol Lipid Products in Cells through Metabolic Engineering by Using a Clickable myo‐Inositol Probe

• Published in: Chembiochem : a European journal of chemical biology Vol. 20; no. 2; pp. 172 - 180
• Main Authors: Ricks, Tanei J., Cassilly, Chelsi D., Carr, Adam J., Alves, Daiane S., Alam, Shahrina, Tscherch, Kathrin, Yokley, Timothy W., Workman, Cameron E., Morrell‐Falvey, Jennifer L., Barrera, Francisco N., Reynolds, Todd B., Best, Michael D.
• Format: Journal Article
• Language: English
• Published: WEINHEIM Wiley 18.01.2019; Wiley Subscription Services, Inc
• Subjects: Biochemistry & Molecular Biology; Biological activity; Biosynthesis; Bladder; Bladder cancer; Cancer; Candida albicans - cytology; Candida albicans - growth & development; Candida albicans - metabolism; Cell growth; Cells, Cultured; Chemistry, Medicinal; Click Chemistry; Disease control; Fluorescence; Fluorescence microscopy; Fluorescent Dyes - chemical synthesis; Fluorescent Dyes - chemistry; fluorescent probes; Humans; Inositol; Inositol - chemical synthesis; Inositol - chemistry; Labeling; Life Sciences & Biomedicine; Lipids; Localization; Mass spectrometry; Mass spectroscopy; Metabolic Engineering; metabolic labeling; Optical Imaging; Pharmacology & Pharmacy; Phosphatidylinositol; Phosphatidylinositols - chemistry; phospholipids; Science & Technology; Tracking; lipids; ANALOGS; CHOLINE; PHOSPHATASE; click chemistry; phospholipids; RELEASE; fluorescent probes; INOSITOL; YEAST; SYNTHASE; CHEMISTRY; metabolic labeling; SELECTIVITY
• ISSN: 1439-4227; 1439-7633
• DOI: 10.1002/cbic.201800248

Phosphatidylinositol (PI) lipids control critical biological processes, so aberrant biosynthesis often leads to disease. As a result, the capability to track the production and localization of these compounds in cells is vital for elucidating their complex roles. Herein, we report the design, synthesis, and application of clickable myo‐inositol probe 1 a for bioorthogonal labeling of PI products. To validate this platform, we initially conducted PI synthase assays to show that 1 a inhibits PI production in vitro. Fluorescence microscopy experiments next showed probe‐dependent imaging in T‐24 human bladder cancer and Candida albicans cells. Growth studies in the latter showed that replacement of myo‐inositol with probe 1 a led to an enhancement in cell growth. Finally, fluorescence‐based TLC analysis and mass spectrometry experiments support the labeling of PI lipids. This approach provides a promising means for tracking the complex biosynthesis and trafficking of these lipids in cells. Tracking the trafficking: A clickable myo‐inositol probe for labeling phosphatidylinositol (PI) lipids in live cells has been developed and validated by PI synthase inhibition assays, cellular fluorescence microscopy, TLC imaging, and mass spectrometry. This platform provides an exciting avenue for tracking the biosynthesis and trafficking of PI lipids, which are commonly dysregulated in disease.