TY - JOUR
T1 - Localization of IFN-γ-Activated Stat1 and IFN Regulatory Factors 1 and 2 in Breast Cancer Cells
AU - Connett, Judith M.
AU - Hunt, Steven R.
AU - Hickerson, Suzanne M.
AU - Wu, Susan J.
AU - Doherty, Gerard M.
PY - 2003/11
Y1 - 2003/11
N2 - The aim of the present work was to evaluate the induction and localization of Stat1, interferon (IFN) regulatory factor-1 (IRF-1), and IRF-2 after IFN-γ exposure of human breast cancer cell lines, SKBR3, MDA468, MCF7, and BT20. Results from growth assays, Western staining, electrophoretic mobility shift assay (EMSA), and immunohistochemical staining were collated to test our hypothesis that immunohistochemical analysis of Stat1, IRF-1, and IRF-2 would provide additional information about the functionality of the IFN-γ signaling pathway in human tumor lines. EMSA results showed that in each of four cell lines, Stat1 expression was increased and demonstrated functional activity after IFN-γ stimulation. Western and EMSA analysis showed upregulation of IRF-1 but not IRF-2 in each cell line. Confocal microscopy of cells stained for Stat1, IRF-1, and IRF-2 confirmed the results and also provided novel information about the intracellular localization of proteins and intercellular variations in responses. The proportion of cells with IRF-1 stimulation and translocation was positively correlated with the IFN-y growth suppression in vitro. In conclusion, using four independent assays, we have demonstrated that heterogeneity in IFN-γ-mediated upregulation of signal transduction proteins can be detected in vitro and that these differences can explain distinct cellular growth effects.
AB - The aim of the present work was to evaluate the induction and localization of Stat1, interferon (IFN) regulatory factor-1 (IRF-1), and IRF-2 after IFN-γ exposure of human breast cancer cell lines, SKBR3, MDA468, MCF7, and BT20. Results from growth assays, Western staining, electrophoretic mobility shift assay (EMSA), and immunohistochemical staining were collated to test our hypothesis that immunohistochemical analysis of Stat1, IRF-1, and IRF-2 would provide additional information about the functionality of the IFN-γ signaling pathway in human tumor lines. EMSA results showed that in each of four cell lines, Stat1 expression was increased and demonstrated functional activity after IFN-γ stimulation. Western and EMSA analysis showed upregulation of IRF-1 but not IRF-2 in each cell line. Confocal microscopy of cells stained for Stat1, IRF-1, and IRF-2 confirmed the results and also provided novel information about the intracellular localization of proteins and intercellular variations in responses. The proportion of cells with IRF-1 stimulation and translocation was positively correlated with the IFN-y growth suppression in vitro. In conclusion, using four independent assays, we have demonstrated that heterogeneity in IFN-γ-mediated upregulation of signal transduction proteins can be detected in vitro and that these differences can explain distinct cellular growth effects.
UR - http://www.scopus.com/inward/record.url?scp=0345305266&partnerID=8YFLogxK
U2 - 10.1089/107999003322558755
DO - 10.1089/107999003322558755
M3 - Article
C2 - 14651776
AN - SCOPUS:0345305266
SN - 1079-9907
VL - 23
SP - 621
EP - 630
JO - Journal of Interferon and Cytokine Research
JF - Journal of Interferon and Cytokine Research
IS - 11
ER -