Mouse Mammary Tumor Virus c-rel Transgenic Mice Develop Mammary Tumors

Abstract

Amplification, overexpression, or rearrangement of the c-rel gene, encoding the c-Rel NF-κB subunit, has been reported in solid and hematopoietic malignancies. For example, many primary human breast cancer tissue samples express high levels of nuclear c-Rel. While the Rev-T oncogene v-rel causes tumors in birds, the ability of c-Rel to transform in vivo has not been demonstrated. To directly test the role of c-Rel in breast tumorigenesis, mice were generated in which overexpression of mouse c-rel cDNA was driven by the hormone-responsive mouse mammary tumor virus long terminal repeat (MMTV-LTR) promoter, and four founder lines identified. In the first cycle of pregnancy, the expression of transgenic c-rel mRNA was observed, and levels of c-Rel protein were increased in the mammary gland. Importantly, 31.6% of mice developed one or more mammary tumors at an average age of 19.9 months. Mammary tumors were of diverse histology and expressed increased levels of nuclear NF-κB. Analysis of the composition of NF-κB complexes in the tumors revealed aberrant nuclear expression of multiple subunits, including c-Rel, p50, p52, RelA, RelB, and the Bcl-3 protein, as observed previously in human primary breast cancers. Expression of the cancer-related NF-κB target genes cyclin D1, c-myc, and bcl-xl was significantly increased in grossly normal transgenic mammary glands starting the first cycle of pregnancy and increased further in mammary carcinomas compared to mammary glands from wild-type mice or virgin transgenic mice. In transient transfection analysis in untransformed breast epithelial cells, c-Rel-p52 or -p50 heterodimers either potently or modestly induced cyclin D1 promoter activity, respectively. Lastly, stable overexpression of c-Rel resulted in increased cyclin D1 and NF-κB p52 and p50 subunit protein levels. These results indicate for the first time that dysregulated expression of c-Rel, as observed in breast cancers, is capable of contributing to mammary tumorigenesis.

ACKNOWLEDGMENTS

R.R.-M. and D.W.K. contributed equally to this work.

We thank T. Gilmore, S. Hann, N. Rice, and M. Ernst for generously providing clones and antibodies. The excellent technical assistance of Judy E. Walls, Lee Gazourian, Alan Lau, and Aundrea Oliver is acknowledged. The excellent assistance of Keisha Bratton in preparation of the manuscript is acknowledged.

This work was supported by grants from the Massachusetts Department of Public Health Breast Cancer Program (to E.L.-B. and R.R.-M.), the Department of Army (DAMD 17-01 10158 to R.R.-M.), and the UCD Mutant Mouse Regional Resource Center (NIH/NCRR U42-RR14905 to R.D.C) and by grants NIH RO1 CA82742 and PO5 ES11624 (G.E.S. and D.C.S.).