Nongenomic oestrogen signalling in oestrogen receptor negative breast cancer cells: a role for the angiotensin II receptor AT1.
BACKGROUND
Oestrogens can mediate some of their cell survival properties through a nongenomic mechanism that involves the mitogen-activated protein kinase (MAPK) pathway. The mechanism of this rapid signalling and its dependence on a membrane bound oestrogen receptor (ER), however, remains controversial. The role of G-protein-coupled receptor and epidermal growth factor (EGF) receptor in an ER-independent signalling pathway modulated by oestrogen was investigated.


METHODS
ER-positive and ER-negative breast cancer cell lines (MCF-7 and SKBR3) and primary breast cancer cell cultures were used in this study. Cell proliferation was assessed using standard MTT assays. Protein and cAMP levels were detected by Western blotting and ELISA, respectively. Antigen localization was performed by immunocytochemistry, immunohistochemistry and immunofluorescence. Protein knockdown was achieved using small interfering RNA technologies.


RESULTS
EGF and oestrogen, alone and in combination, induced cell proliferation and phosphorylation of MAPK proteins Raf and ERK (extracellular signal regulated kinase)1/2 in both ER-negative SKBR3 and ER-positive MCF-7 human breast cancer cell lines. Increased Raf phosphorylation was also observed in primary human breast cultures derived from ER-positive and ER-negative breast tumours. Oestrogen induced an increase in intracellular cAMP in ER-negative SKBR3 human breast cancer cells. Oestrogen-mediated cell growth and phosphorylation of MAPK was modified by the EGF receptor antagonist AG1478, the G-protein antagonist pertussis toxin, and the angiotensin II receptor antagonist saralasin. Knockdown of angiotensin II type 1 receptor (AT1) protein expression with small interfering RNA attenuated oestrogen-induced Raf phosphorylation in ER-negative cells. AT1 receptor was found to be expressed in the cell membrane of breast tumour epithelial cells.


CONCLUSIONS
These findings provide evidence that, in breast cancer cells, oestrogen can signal through AT1 to activate early cell survival mechanisms in an ER-independent manner.
Lim
Kheng Tian
KT
School of Medicine and Medical Science, St, Vincent's University Hospital, Elm Park, Dublin, Ireland. ktlim@doctors.org.uk
Cosgrave
Niamh
N
Hill
Arnold David
AD
Young
Leonie S
LS
D001943:Breast Neoplasms / Q000503:physiopathology*; D049109:Cell Proliferation; D002470:Cell Survival; D000242:Cyclic AMP / Q000378:metabolism; D005260:Female; D006801:Humans; D020929:Mitogen-Activated Protein Kinase Kinases / Q000378:metabolism; D010766:Phosphorylation; D034741:RNA, Small Interfering; D044140:Receptor, Angiotensin, Type 1 / Q000502:physiology*; D011960:Receptors, Estrogen / Q000502:physiology*; D015398:Signal Transduction; D014407:Tumor Cells, Cultured
D034741:RNA, Small Interfering; D044140:Receptor, Angiotensin, Type 1; D011960:Receptors, Estrogen; D000242:Cyclic AMP; D020929:Mitogen-Activated Protein Kinase Kinases