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Cripto-1 inhibits beta-casein expression in mammary epithelial cells through a p21ras-and phosphatidylinositol
M L De Santis1, S Kannan, G H Smith
1Tumor Growth Factor Section, National Cancer Institute, NIH, Bethesda, Maryland 20892, USA.
Abstract:
Cripto-1 (CR-1) is a recently discovered protein of the epidermal growth factor family that does not directly activate any of the known erbB type 1 tyrosine kinase receptors. Also, CR-1 stimulates the growth of HC-11 mouse mammary epithelial cells. We found that prior treatment of HC-11 cells with exogenous CR-1 induced a competency response to the lactogenic hormones dexamethasone, insulin, and prolactin (DIP) with respect to the induction of the milk protein beta-casein. In contrast, simultaneous treatment of mouse HC-11 cells with CR-1 in the presence of DIP inhibited beta-casein expression. The inhibitory effects of CR-1 on beta-casein expression in response to DIP were not unique to this mouse mammary epithelial cell line, because beta-casein and whey acidic protein expression in primary mouse mammary explant cultures established from midpregnant mice were also differentially inhibited by several epidermal growth factor-related peptides including CR-1. The mitogenic and differentiation effects of CR-1 are mediated by the binding of CR-1 to a cell surface receptor that is known to activate the ras/raf/mitogen-activated protein kinase (MAPK)/MAPK kinase pathway. The inhibitory response of CR-1 in HC-11 cells on beta-casein expression after treatment with DIP can be attenuated by B581, a peptidomimetic farnesyltransferase inhibitor that blocks p21ras farnesylation and activation, and by the phosphatidylinositol 3'-kinase (PI3k) inhibitor LY 294002 but not by PD 98059, a MAPK kinase inhibitor that blocks MAPK activation. These data suggest that the ability of CR-1 to block lactogenic hormone-induced expression of beta-casein is mediated through a p21ras-dependent, PI3k-mediated pathway. This is further substantiated by the observation that CR-1 is able to stimulate the tyrosine phosphorylation of the p85 PI3k regulatory subunit and to increase the activity of PI3k in HC-11 cells.
Insights
Cripto-1 (CR-1) protein differentially affects mammary cell growth and milk protein expression. While CR-1 can prime cells for lactogenic hormones, it also inhibits beta-casein expression via a p21ras-dependent pathway.
Area of Science:
- Cell Biology
- Molecular Biology
- Biochemistry
Background:
- Cripto-1 (CR-1) is an epidermal growth factor family protein.
- CR-1 influences mammary epithelial cell growth and differentiation.
- Its precise role in lactogenic hormone signaling is not fully understood.
Purpose of the Study:
- To investigate the dual role of CR-1 in mammary epithelial cells.
- To elucidate the signaling pathways mediating CR-1's effects on beta-casein expression.
- To determine CR-1's interaction with lactogenic hormone-induced differentiation.
Main Methods:
- Cell culture of HC-11 mouse mammary epithelial cells and primary mammary explants.
- Treatment with CR-1 and lactogenic hormones (dexamethasone, insulin, prolactin).
- Analysis of beta-casein and whey acidic protein expression.
- Pharmacological inhibition of signaling pathways (ras, MAPK, PI3k).
Main Results:
- CR-1 induced cell competency for lactogenic hormones but inhibited beta-casein expression when applied simultaneously.
- These inhibitory effects were observed in both cell lines and primary explants.
- Inhibition was mediated by a p21ras-dependent, phosphatidylinositol 3'-kinase (PI3k)-dependent pathway, not MAPK.
- CR-1 stimulated PI3k activity and tyrosine phosphorylation of its regulatory subunit.
Conclusions:
- CR-1 exhibits context-dependent effects on mammary cell differentiation.
- CR-1 inhibits lactogenic hormone-induced beta-casein expression through a novel p21ras/PI3k signaling axis.
- This finding provides new insights into the regulation of mammary gland development and function.