Stable or acute PI3K activation decreases tRAH-induced glycosylated NIS protein expression and NIS-mediated radioactive iodide uptake in MCF-7 cells. (A) 48 hours of tRAH treatment induces fully glycosylated NIS protein in MCF-7 cells transiently transfected with empty vector. Underglycosylated 50 kDa NIS is dominant in MCF-7/PI3K p110αCAAX cells, which minimally respond to tRAH treatment. Transient expression of PI3K p110αCAAX also induces underglycosylated NIS protein expression with a modest decrease in the tRAH-induced fully glycosylated NIS form. (B) tRAH treatment cannot induce NIS function in MCF-7/PI3K p110αCAAX cells. A small increase in basal NIS-mediated radioactive iodide uptake is noted in MCF-7/PI3K p110αCAAX stable clones versus parental MCF-7 cells. Cells were treated with tRAH for 48 hours followed by 125I uptake assay. Perchlorate (ClO4
-) is a specific inhibitor of NIS function. ***p < .0001. (C) Acute expression of activated PI3K p110α decreases tRAH-induced NIS function in MCF-7 cells. A modest increase in basal NIS-mediated radioactive iodide uptake is present in MCF-7 cells transiently transfected with PI3K p110αCAAX versus vector-only controls. Cells were transiently transfected with pcDNA3/PI3K p110αCAAX or empty vector for 6 hours, and then treated with tRAH for 42 hours prior to 125I uptake assay. *p < .05.