Developmental stage determines estrogen receptor alpha expression and non-genomic mechanisms that control IGF-1 signaling and mammary proliferation in mice
Jie Tian, … , Claudio J. Conti, Robin Fuchs-Young
Jie Tian, … , Claudio J. Conti, Robin Fuchs-Young
Published December 19, 2011
Citation Information: J Clin Invest. 2012;122(1):192-204. https://doi.org/10.1172/JCI42204.
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Research Article Oncology

Developmental stage determines estrogen receptor alpha expression and non-genomic mechanisms that control IGF-1 signaling and mammary proliferation in mice

Abstract

Insulin like growth factor–1 (IGF-1) stimulates increased proliferation and survival of mammary epithelial cells and also promotes mammary tumorigenesis. To study the effects of IGF-1 on the mammary gland in vivo, we used BK5.IGF-1 transgenic (Tg) mice. In these mice, IGF-1 overexpression is controlled by the bovine keratin 5 promoter and recapitulates the paracrine exposure of breast epithelium to stromal IGF-1 that is seen in women. Studies have shown that BK5.IGF-1 Tg mice are more susceptible to mammary tumorigenesis than wild-type littermates. Investigation of the mechanisms underlying increased mammary cancer risk, reported here, revealed that IGF-1 preferentially activated the PI3K/Akt pathway in glands from prepubertal Tg mice, resulting in increased cyclin D1 expression and hyperplasia. However, in glands from postpubertal Tg mice, a pathway switch occurred and activation of the Ras/Raf/MAPK pathway predominated, without increased cyclin D1 expression or proliferation. We further showed that in prepubertal Tg glands, signaling was mediated by formation of an ERα/IRS-1 complex, which activated IRS-1 and directed signaling via the PI3K/Akt pathway. Conversely, in postpubertal Tg glands, reduced ERα expression failed to stimulate formation of the ERα/IRS-1 complex, allowing signaling to proceed via the alternate Ras/Raf/MAPK pathway. These in vivo data demonstrate that changes in ERα expression at different stages of development direct IGF-1 signaling and the resulting tissue responses. As ERα levels are elevated during the prepubertal and postmenopausal stages, these may represent windows of susceptibility during which increased IGF-1 exposure maximally enhances breast cancer risk.

Authors

Jie Tian, Thomas R. Berton, Stephanie H. Shirley, Isabel Lambertz, Irma B. Gimenez-Conti, John DiGiovanni, Kenneth S. Korach, Claudio J. Conti, Robin Fuchs-Young

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Figure 5

ERα expression levels determine the IGF-1 signal transduction pathway in the mammary gland.

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ERα expression levels determine the IGF-1 signal transduction pathway in...
(A) Western blots of extracts from prepubertal mammary glands reveal increased formation of the ERα/IRS-1 complex, which can be immunoprecipitated with antibodies to either IRS-1 (left panel) or ERα (right panel). (B) Conversely, in extracts from postpubertal mammary glands, comparatively little ERα is co-precipitated with IRS-1 antibodies, indicating that complex formation is greatly reduced. (C) Side-by side comparison of anti–IRS-1 immunoprecipitates, showing that ERα/IRS-1 complexes are detected in glandular extracts from prepubertal Tg mice. In order to ultimately load equivalent amounts of total IRS-1, 250 μg of glandular extracts from prepubertal mice and 500 μg of extracts from postpubertal mice were incubated with anti–IRS-1. Western blot analysis of total and phosphorylated c-Raf (Ser259) in extracts of (D) prepubertal and (E) postpubertal Tg and WT mammary glands. Each lane represents pooled glands from 4–6 mice of the same age and genotype. Representative Western blots from 2–3 different pools per genotype and age are shown in A, B, D, and E. A representative Western blot of 3 independent experiments is shown in C. Bar graphs in D and E represent mean ± SD of densitometric analysis of blots from 5–6 different pools/lanes of each genotype and age. Statistical analysis performed using Student’s t test; *P < 0.05, p–c-Raf/c-Raf ratio versus WT.