Construction and identification of FRT cell line stably expressing UT-A2

AIM: To obtain an FRT cell line that can stably express urea transporter A2 (UT-A2) and provide a cell model for screening UT-A2 inhibitors. METHODS: FRT cells stably expressing aquaporins 1 (AQP1) and YFP were transfected with the recombinant plasmid pUB6/V5-UT-A2 by eukaryotic expression plasmid-lipoplast mediating pathway. The stable UT-A2-FRT cell line was cloned by selection with BSD and confirmed by Western blotting and immunofluorescence staining. The urea permeability across the plasma membrane was detected by a 2 mol/L urea lysis assay. RESULTS: We have obtained a stable UT-A2-FRT cell line. Western blotting analysis showed that UT-A2 protein was expressed stably in this cell line. The immunofluorescence staining detection indicated UT-A2 expression in the plasma membrane. It was found that there was significant urea permeability in this cell line by 2 mol/L urea lysis assay. CONCLUSION: We constructed an FRT cell line that could stably express UT-A2 in plasma membrane in the non-renal epithelia cell. The cell line will be used to screen UT-A2 inhibitors.