Monitoring bile acid transport in single living cells using a genetically encoded Förster resonance energy transfer sensor
| Autor: | Stan F.J. van de Graaf, Leo W. J. Klomp, Saskia W.C. van Mil, Maarten Merkx, Ingrid T. G. W. Bijsmans, Misha V. Golynskiy, Lieke M. van der Velden |
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| Přispěvatelé: | Protein Engineering, Macro-Organic Chemistry, Adult Psychiatry, Amsterdam Gastroenterology Endocrinology Metabolism, Gastroenterology and Hepatology |
| Jazyk: | angličtina |
| Rok vydání: | 2013 |
| Předmět: |
Cytoplasm
medicine.drug_class Cell Organic Anion Transporters Sodium-Dependent Receptors Cytoplasmic and Nuclear Biosensing Techniques Biology digestive system Bile Acids and Salts chemistry.chemical_compound Chenodeoxycholic acid Fluorescence Resonance Energy Transfer medicine Animals Humans Zebrafish Fluorescent Dyes Cell Nucleus Membrane Glycoproteins Symporters Hepatology Bile acid Membrane Transport Proteins Metabolism Peroxisome G protein-coupled bile acid receptor medicine.anatomical_structure chemistry Biochemistry Farnesoid X receptor Carrier Proteins CYP8B1 |
| Zdroj: | Hepatology, 75(2), 740-752. Wiley Hepatology (Baltimore, Md.), 57(2), 740-752. John Wiley and Sons Ltd |
| ISSN: | 0270-9139 1527-3350 |
| DOI: | 10.1002/hep.26012 |
| Popis: | Bile acids are pivotal for the absorption of dietary lipids and vitamins and function as important signaling molecules in metabolism. Here, we describe a genetically encoded fluorescent bile acid sensor (BAS) that allows for spatiotemporal monitoring of bile acid transport in single living cells. Changes in concentration of multiple physiological and pathophysiological bile acid species were detected as robust changes in Forster resonance energy transfer (FRET) in a range of cell types. Specific subcellular targeting of the sensor demonstrated rapid influx of bile acids into the cytoplasm and nucleus, but no FRET changes were observed in the peroxisomes. Furthermore, expression of the liver fatty acid binding protein reduced the availability of bile acids in the nucleus. The sensor allows for single cell visualization of uptake and accumulation of conjugated bile acids, mediated by the Na+-taurocholate cotransporting protein (NTCP). In addition, cyprinol sulphate uptake, mediated by the putative zebrafish homologue of the apical sodium bile acid transporter, was visualized using a sensor based on the zebrafish farnesoid X receptor. The reversible nature of the sensor also enabled measurements of bile acid efflux in living cells, and expression of the organic solute transporter αβ (OSTαβ) resulted in influx and efflux of conjugated chenodeoxycholic acid. Finally, combined visualization of bile acid uptake and fluorescent labeling of several NTCP variants indicated that the sensor can also be used to study the functional effect of patient mutations in genes affecting bile acid homeostasis. Conclusion: A genetically encoded fluorescent BAS was developed that allows intracellular imaging of bile acid homeostasis in single living cells in real time. (HEPATOLOGY 2013) |
| Databáze: | OpenAIRE |
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