Fluorescent nucleoside derivatives as a tool for the detection of concentrative nucleoside transporter activity using confocal microscopy and flow cytometry.
The abundance and function of transporter proteins at the plasma membrane are likely to be crucial in drug responsiveness. Functional detection of human concentrative nucleoside transporters (hCNTs) is of interest for predicting drug sensitivity because of their ability to transport most nucleoside-derived drugs. In the present study, two fluorescent nucleoside analogues, uridine-furan and etheno-cytidine, were evaluated as tools to study in vivo nucleoside transporter-related functions. These two molecules showed high affinity interactions with hCNT1 and hCNT3 and were shown to be substrates of both transporters. Both fluorescence microscopy and flow cytometry experiments showed that uridine-furan uptake was better suited for distinguishing cells that express hCNT1 or hCNT3. These data highlight the usefulness of fluorescent nucleoside derivatives, as long as they fulfill the requirements of confocal microscopy and flow cytometry, for in vivo analysis of hCNT-related function.
†Department of Biochemistry and Molecular Biology, University of Barcelona, Institute of Biomedicine (IBUB), Barcelona, Spain.|‡Oncology Programme, National Biomedical Research Institute on Liver and Gastrointestinal Diseases (CIBER ehd), Instituto de Salud Carlos III, Barcelona, Spain.|⊥Department of Organic Chemistry, University of Barcelona, Institute of Biomedicine (IBUB), Barcelona, Spain.
Ana
A
Claudio-Montero
†Department of Biochemistry and Molecular Biology, University of Barcelona, Institute of Biomedicine (IBUB), Barcelona, Spain.|‡Oncology Programme, National Biomedical Research Institute on Liver and Gastrointestinal Diseases (CIBER ehd), Instituto de Salud Carlos III, Barcelona, Spain.
Itziar
I
Pinilla-Macua
†Department of Biochemistry and Molecular Biology, University of Barcelona, Institute of Biomedicine (IBUB), Barcelona, Spain.|‡Oncology Programme, National Biomedical Research Institute on Liver and Gastrointestinal Diseases (CIBER ehd), Instituto de Salud Carlos III, Barcelona, Spain.
Paula
P
Fernández-Calotti
#Department of Nutrition, Food Science and Physiology, University of Navarra, Pamplona, Spain.
Carlos
C
Sancho-Mateo
#Department of Nutrition, Food Science and Physiology, University of Navarra, Pamplona, Spain.
María Pilar
MP
Lostao
§Hematopathology Unit, Hospital Clínic and Institut d'Investigacions Biomèdiques August Pi i Sunyer (IDIBAPS), Barcelona, Spain.
Dolors
D
Colomer
⊥Department of Organic Chemistry, University of Barcelona, Institute of Biomedicine (IBUB), Barcelona, Spain.
Anna
A
Grandas
†Department of Biochemistry and Molecular Biology, University of Barcelona, Institute of Biomedicine (IBUB), Barcelona, Spain.|‡Oncology Programme, National Biomedical Research Institute on Liver and Gastrointestinal Diseases (CIBER ehd), Instituto de Salud Carlos III, Barcelona, Spain.
Marçal
M
Pastor-Anglada
D005434:Flow Cytometry / Q000379:methods*; D006801:Humans; D018613:Microscopy, Confocal / Q000379:methods*; D033703:Nucleoside Transport Proteins / Q000378:metabolism*; D009705:Nucleosides / Q000737:chemistry*
D033703:Nucleoside Transport Proteins; D009705:Nucleosides
confocal microscopy; drug transporters; flow cytometry; fluorescent nucleosides