Tracking fructose 1,6-bisphosphate dynamics in liver cancer cells using a fluorescent biosensor

  • iScience. 2024 Nov 6;27(12):111336. doi: 10.1016/j.isci.2024.111336.
Israel Pérez-Chávez  1  2  3  4 ,  John N Koberstein  5 ,  Julia Malo Pueyo  1  2  3 ,  Eduardo H Gilglioni  4 ,  Didier Vertommen  6 ,  Nicolas Baeyens  7 ,  Daria Ezeriņa  1  2  3 ,  Esteban N Gurzov  4  8 ,  Joris Messens  1  2  3
Affiliations
  • 1. VIB-VUB Center for Structural Biology, Vlaams Instituut Voor Biotechnologie, B-1050 Brussels, Belgium.
  • 2. Brussels Center for Redox Biology, Vrije Universiteit Brussel, B-1050 Brussels, Belgium.
  • 3. Structural Biology Brussels, Vrije Universiteit Brussel, B-1050 Brussels, Belgium.
  • 4. Signal Transduction and Metabolism Laboratory, Université Libre de Bruxelles (ULB), Brussels, Belgium.
  • 5. HHMI Janelia Research Campus, Ashburn, VA, USA.
  • 6. de Duve Institute, MASSPROT Platform, UCLouvain, 1200 Brussels, Belgium.
  • 7. Laboratoire de Physiologie et de Pharmacologie (LAPP), Université Libre de Bruxelles (ULB), Brussels, Belgium.
  • 8. WELBIO Department, WEL Research Institute, Avenue Pasteur 6, Wavre B-1300, Belgium.
PMID: 39640569 DOI: 10.1016/j.isci.2024.111336
Abstract

HYlight is a genetically encoded fluorescent biosensor that ratiometrically monitors fructose 1,6-bisphosphate (FBP), a key glycolytic metabolite. Given the role of glucose in liver Cancer metabolism, we expressed HYlight in human liver Cancer cells and primary mouse hepatocytes. Through in vitro, in silico, and in cellulo experiments, we showed HYlight's ability to monitor FBP changes linked to glycolysis, not gluconeogenesis. HYlight's affinity for FBP was ∼1 μM and stable within physiological pH range. HYlight demonstrated weak binding to dihydroxyacetone phosphate, and its ratiometric response was influenced by both ionic strength and phosphate. Therefore, simulating cytosolic conditions in vitro was necessary to establish a reliable correlation between HYlight's cellular responses and FBP concentrations. FBP concentrations were found to be in the lower micromolar range, far lower than previous millimolar estimates. Altogether, this biosensor approach offers real-time monitoring of FBP concentrations at single-cell resolution, making it an invaluable tool for the understanding of Cancer metabolism.

Keywords
biochemistry methods; cancer; metabolomics.
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