Cellular Localization of Green Fluorescent Protein (GFP)-tagged Sushi Domain Containing 3 (SUSD3) in T47D Breast Cancer Cells

Abstract

BACKGROUND: Microarray analyses from breast cancer samples (n=50) of postmenopausal women with favorable and unfavorable response to aromatase inhibitors (AI) was performed. Differential expression of SUSD3 in AI responders and in ERa+/PR+ tumors was observed. Among the patients with ERa+ tumors, those who responded had 2.5x higher expression of SUSD3 suggesting that SUSD3 may be a better molecular predictor of AI responsiveness. We have previously shown that SUSD3 expression is strongly correlated to ERa and can be induced by estradiol (E2) treatment. Induction of SUSD3 by E2 is likely a direct effect of ERa- downstream signaling as demonstrated by cycloheximide treatment. Additionally, we have demonstrated ERa binding to a likely promoter region of SUSD3. Our objective was thus, to localize SUSD3 in attempts to better understand its possible function in breast cancer. MATERIALS AND METHOD(S): GFP-tagged ORF clone of Homo sapiens SUSD3 driven by CMV promoter was transfected into T47D cells using Promega FuGene HD transfection reagent. Cells were fixed with cold methanol and processed. DAPI staining was utilized for nuclear localization. Confocal imaging was performed. RESULT(S): SUSD3 was localized to the cellular membrane and was highly concentrated upon cell-to-cell contact. CONCLUSION(S): The cellular localization of SUSD3 to the cellular membrane, its strong correlation with ERa, and its induction by estradiol offer possible insights to SUSD3's functionality. We hypothesize a potential role of SUSD3 in cell adhesion and cell-to-cell contact.