A mouse model for the role of p130 Crk associated substrate in mammary gland development and cancer.

摘要

p130Cas (Crk associated substrate)/BCAR1 (breast cancer anti-estrogen resistant) is activated in established breast cancer cell lines. Patients with primary breast tumors that express high levels of p130Cas/BCAR1 protein have a poor prognosis. However, the role(s) of p130Cas in breast cancer is still not clear, and p130Cas-deficient mice die at an early embryonic stage because of heart failure, thus precluding an investigation of breast cancer and mammary gland development in this model. We have established a p130Cas dominant-interfering transgenic mouse model by using a constitutively phosphorylated p130Cas substrate domain (SD). Under the control of the mouse mammary tumor virus-long terminal repeat (MMTV-LTR), this construct is preferentially expressed in the epithelium of the mammary gland. In contrast to p130Cas-deficient mice, these mice were viable, breed normally, and passed the transgene in a Mendelian fashion with a slightly smaller litter size. Mice expressed the transgene in the mammary gland and in other tissues where the MMTV-LTR is active, including the salivary gland, harderian gland, spleen, and brain. Transgenic mice have an increased body mass, which is the result of an accumulation of fat and/or an increase in organ weight. Whole mount mammary gland examination showed premature secondary branch development in virgin 6 week old Src*/CasSD transgenic mice and dilated duct formation. To further study the role of p130Cas in breast cancer in vivo, Src*/CasSD mice were mated with transgenic mice carrying an active form of c-Src (c-Src527). The double transgenic mice (Src527/Cas) are viable with normal litter size. Lastly, we have attempted to establish a dominant-active p130Cas molecule by targeting the Src*/CasSD molecule to focal adhesion complexes through the SH3 domain of p130Cas (Src*/CasSD-CSH3, NSH3Src*/CasSD). Expression of these constructs in 3Y1 cells strongly increased the overall tyrosine phosphorylation of cellular proteins. These changes are associated with a change in morphology compared to vector control or Src*/CasSD expressing cells. Src*/CasSD-CSH3 and NSH3-Src*/CasSD expressing cells are spindle shaped and highly refractile, which suggests that these cells are more motile and transformed. In summary, we established a viable mouse model, which can be used to study the role of signaling pathway downstream of the p130Cas SD in mammary gland development and breast cancer.