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. 2019 Mar 21;8(3):265. doi: 10.3390/cells8030265

Figure 2.

Figure 2

AQP2 and USP4 interact in vitro and in vivo. For studies in cultured mpkCCD14 cells, cells were grown in the presence of dDAVP for 4 days to induce AQP2 expression. On the day of the experiment, cells were washed with DMEM/F12 and incubated in DMEM/F12 for 2 h at 37 °C. Subsequently, cells were treated with either vehicle or dDAVP for 20 min at 37 °C prior to immunoprecipitation (IP) using an anti-AQP2 antibody. For studies in MDCK-hAQP2 cells, on the day of the experiment, cells were washed with DMEM media and subsequently treated with either vehicle or forskolin for 20 min prior to IP using anti-AQP2 antibody. (A) Representative immunoblots of USP4 and AQP2 interaction in mpkCCD14 cells. (B) Representative immunoblots of USP4 and AQP2 interaction in hAQP2-MDCK cells. (C) Representative immunoblots of USP4 and AQP2 in samples immunoprecipitated from mouse kidney tubules acutely treated with either vehicle or dDAVP (30 min) using anti-AQP2 or anti-USP4 antibodies. Each lane represents an individual protein sample. All samples were run on the same membrane and had the same exposure time. Breaks in the images are to show the immunoprecipitation controls (antibody (AB) alone or lysate alone) next to the samples. NS = non-specific band.