Pituitary tumor transforming gene binding factor (PBF): a novel modulator of iodide uptake and target for Src phosphorylation in breast cancer

Although the sodium iodide symporter (NIS) is expressed in 70–80% of breast cancers, only 20–30% is located at the plasma membrane (PM) and therefore functional. Previous work in thyroid cells leads demonstrated that PBF redistributes NIS from the PM into intracellular vesicles, potently reducing radioiodide uptake. We therefore examined whether increased membranous NIS could facilitate radioiodide therapy for breast cancer. Immunofluorescent microscopy revealed co-localisation between NIS, phosphorlyated (pY174) and total PBF in MCF-7 cells, with PBF transfection leading to increased intracellular NIS staining. Treatment with the Src family kinase (SFK) inhibitor dasatinib inhibited PBF phosphorylation and led to increased NIS PM staining and decreased co-localisation with PBF. PBF significantly repressed radioiodide uptake in MCF-7 and MDA-MB-231 cells expressing exogenous NIS (25 and 30% reduction respectively, both n=3 and P<0.05), which was restored by dasatanib treatment (1.75- and 2-fold increase, n=3, P<0.05). Two PBF mutants were next utilised: i) EEN170-172AAA, lacking the predicted Src consensus sequence, and ii) Y174A, disrupting the endocytosis motif. EEN170-172AAA was unphosphorylated, incapable of reducing radioiodide uptake and showed WT subcellular localisation, whereas Y174A did not reduce iodide uptake and was retained in the PM. These data suggest that phosphorylation of PBF, not localisation, is critical for the interaction between NIS and PBF. To investigate Src phosphorylation of PBF, a mutant form of Src (T341I), resistant to dasatinib, was utilised. In the presence of T341I Src, dasatinib no longer rescued PBF’s repression of radioiodide uptake in MCF-7 cells, confirming that Src is the kinase responsible for the phosphorylation of PBF. Taken together, these data suggest that PBF alters the subcellular location of NIS and that Src’s phosphorylation of PBF modulates the ability of breast carcinoma cell-lines to uptake radioiodide. This has critical implications for adapting NIS as a potential therapy in breast cancer.