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. 2019 Jun 5;10(1):186–194. doi: 10.1016/j.apsb.2019.05.005

Figure 5.

Figure 5

PKA activator Bt2-cAMP up-regulates OAT3 transport function and phosphorylation. (A) OAT3-expressing cells were treated with Bt2-cAMP (5 nmol/L) for 2 h. 4-min uptake of [3H]-estrone sulfate (ES, 300 nmol/L) was then measured. Transport activity was expressed as percentage of the uptake in control cells. The data corresponded to the uptake of OAT3-expressing cells minus uptake of untreated parental cells. Values are mean ± SD (n = 3); *P < 0.05. (B) Phosphorylation of OAT3. Top panel: OAT3-expressing cells were treated with Bt2-cAMP (5 nmol/L) for 2 h. After treatment, cells were lysed, pre-cleared and immunoprecipitated with anti-myc agarose affinity gel, followed by immunoblotting (IB) with anti-phospho-Ser/Thr antibody. Bottom panel: The identical blot of the top panel was re-probed with anti-myc antibody to determine the amount of OAT3 immunoprecipitated. (C) Densitometry analysis of blot results from Fig. 5B top panel as well as from other experiments. The values are mean ± SD (n = 3); *P < 0.05. Statistical analysis was performed using Student's paired t-tests.