Expression of B Cell Activating Factor Pathway Genes in Mouse Mammary Gland

  • Choi, S. (Major in Molecular Biotechnology, Biotechnology Research Institute, Inst. of Ag. Sci. and Tech. Chonnam National University) ;
  • Jung, D.J. (Major in Molecular Biotechnology, Biotechnology Research Institute, Inst. of Ag. Sci. and Tech. Chonnam National University) ;
  • Bong, J.J. (Major in Molecular Biotechnology, Biotechnology Research Institute, Inst. of Ag. Sci. and Tech. Chonnam National University) ;
  • Baik, M. (Major in Molecular Biotechnology, Biotechnology Research Institute, Inst. of Ag. Sci. and Tech. Chonnam National University)
  • Received : 2006.03.22
  • Accepted : 2006.10.25
  • Published : 2007.02.01


In our previous study, overexpression of extracellular proteinase inhibitor (Expi) gene accelerated apoptosis of mammary epithelial cells, and induced expression of B cell activating factor (BAFF) gene. In this study, we found induction of BAFF-receptor (BAFF-R) gene expression in the Expi-transfected cells. A proliferation-inducing ligand (APRIL) gene is another TNF family member and the closest known relative of BAFF. We found induction of APRIL gene expression in the Expi-overexpressed apoptotic cells. NF-${\kappa}$B gene was also induced in the Expi-overexpressed cells. Expression patterns of BAFF and APRIL pathway-related genes were examined in in vivo mouse mammary gland at various reproductive stages. Expression levels of BAFF gene were very low at early pregnancy, increased from mid-pregnancy, and peaked at lactation, and thereafter decreased at involution stages of mammary gland. Expression of BAFF-R gene was highly induced in involution stages compared to lactation stages. Thus, expression patterns of BAFF-R gene were correlated to apoptotic status of mammary gland: active apoptosis of mammary epithelial cells occurs at involution stage of mammary gland. Expression levels of NF-${\kappa}$B gene were higher in involution stages compared to lactation stages. We analyzed mRNA levels of bcl-2 family genes from different stages of mammary development. Bcl-2 gene expression was relatively constant during lactation and involution stages. There was a slight increase in bcl-xL gene expression in involution stages compared to lactation state. Bax gene expression was highly induced in involution stage. Our results suggest that signaling pathways activated by both BAFF and ARRIL in mammary gland point towards NF-${\kappa}$B activation which causes upregulation of bax.


B Cell Activating Factor;APRIL;Apoptosis;Mammary Gland


Supported by : Korea Research Foundation


  1. Ball, R. K., R. R. Friis, C.-A. Schonenberger, W. Doppler and B. Groner. 1994. Prolactin regulation of $\beta$-casein gene expression and of a cytosolic 120-kd protein in a cloned mouse mammary epithelial cell line. EMBO J. 7:2089-2095.
  2. Chapman, R. S., P. C. Lourenco, E. Tonner, D. J. Flint, S. Selbert, K. Takeda, S. Akira, A. R. Clarke and C. J. Watson. 1999. Suppression of epithelial apoptosis and delayed mammary gland involution in mice with a conditional knockout of Stat3. Genes Dev. 13:2604-2616.
  3. Grimm, S., M. K. Bauer, P. A. Baeuerle and K. Schulze-Osthoff. 1996. Bcl-2 down-regulates the activity of transcription factor NF-kappaB induced upon apoptosis. J. Cell Biol. 134:13-23.
  4. Hahne, M., T. Kataoka, M. Schröter, K. Hofmann, M. Irmler, J.- L. Bodmer, P. Schneider, T. Bornand, N. Holler, L. E. French, B. Sordat, D. Rimoldi and J. Tschopp. 1998. APRIL, a new ligand of the tumor necrosis factor family, stimulates tumor cell growth. J. Exp. Med. 188:1185-1190.
  5. Lin, B., C. Williams-Skipp, Y. Tao, M. S. Schleicher, L. L. Cano, R. C. Duke and R. Scheinman. 1999. NF-kappaB functions as both a proapoptotic and antiapoptotic regulatory factor within a single cell type. Cell Death Differ. 6:570-582.
  6. Negrini, M., E. Silini, C. Kozak, Y. Tsujimoto and C. M. Croce. 1987. Molecular analysis of mbcl-2: structure and expression of the murine gene homologous to the human gene involved in follicular lymphoma. Cell 49:455-463.
  7. Rayet, B. and C. Gelinas. 1999. Aberrant rel/nfkb genes and activity in human cancer. Oncogene 18:6938-6947.
  8. Schneider, P., F. MacKay, V. Steiner, K. Hofmann, J. L. Bodmer, N. Holler, C. Ambrose, P. Lawton, S. Bixler, H. Acha-Orbea, D. Valmori, P. Romero, C. Werner-Favre, R. H. Zubler, J. L. Browning and J. Tschopp. 1999. BAFF, a novel ligand of the tumor necrosis factor family, stimulates B cell growth, J. Exp. Med. 189:1747-1756.
  9. Seol, M. B., J. J. Bong and M. Baik. 2006. Involvement of cathepsin D in apoptosis of mammary epithelial cells. Asian-Aust. J. Anim. Sci. 19:1100-1105.
  10. Talhouk, R. S., M. J. Bissell and Z. Werb. 1992. Coordinated expression of extracellular matrix-degrading proteinases and their inhibitors regulates mammary epithelial function during involution. J. Cell Biol. 118:1271-1282.
  11. Van Antwerp, D. J., S. J. Martin, T. Kafri, D. R. Green and I. M. Verma. 1996. Suppression of TNF-alpha-induced apoptosis by NF-kappaB. Sci. 274:787-789.
  12. Walker, N. I., R. E. Bennett and J. F. Kerr. 1989. Cell death by apoptosis during involution of the lactating breast in mice and rats. Am. J. Anat. 185:19-32.
  13. Walton, K. D., K. U. Wagner, E. B. Rucker. 3rd, J. M. Shillingford, K. Miyoshi and L. Hennighausen. 2001. Conditional deletion of the bcl-x gene from mouse mammary epithelium results in accelerated apoptosis during involution but does not compromise cell function during lactation. Mech. Dev. 109:281-293.
  14. Zheng, Y., F. Ouaaz, P. Bruzzo, V. Singh, S. Gerondakis and A. A. Beg. 2001. NF-kappa B RelA (p65) is essential for TNFalphainduced fas expression but dispensable for both TCR induced expression and activation-induced cell death. J. Immunol. 166:4949-4957.
  15. Dear, T. N., I. A. Ramshaw and R. F. Kefford. 1988. Differential expression of a novel gene, WDNM1, in nonmetastatic rat mammary adenocarcinoma cells. Cancer Res. 48:5203-5209.
  16. Marti, A., Z. Feng, H. J. Altermatt and R. Jaggi. 1997. Milk accumulation triggers apoptosis of mammary epithelial cells. Eur. J. Cell Biol. 73:158-165.
  17. Marti, A., H. Lazar, P. Ritter and R. Jaggi. 1999. Transcription factor activities and gene expression during mouse mammary gland involution. J. Mammary Gland Biol. Neoplasia 4:145-152.
  18. Ravi, R., G. C. Bedi, L. W. Engstrom, Q. Zeng, B. Mookerjee, C. Gelinas, E. J. Fuchs and A. Bedi. 2001. Regulation of death receptor expression and TRAIL/Apo2L-induced apoptosis by NF-kappaB. Nat. Cell Biol. 3:409-416.
  19. Feng, Z., A. Marti, B. Jehn, H. J. Altermatt, G. Chicaiza and R. Jaggi. 1995. Glucocorticoid and progesterone inhibit involution and programmed cell death in the mouse mammary gland. J. Cell Biol. 131:1095-1103.
  20. Boudreau, N., C. J. Sympson, Z. Werb and M. J. Bissell. 1995. Suppression of ICE and apoptosis in mammary epithelial cells by extracellular matrix. Sci. 267:891-893.
  21. Chomozynski, P. and N. Sacchi. 1987. Single-step method of RNA isolation by acid guanidinium thiocyanate-phenol-chloroform extraction. Anal. Biochem. 162:156-159.
  22. Gonzalez-Garcia, M., R. Perez-Ballestero, L. Ding, L. Duan, L. H. Boise, C. B. Thompson and G. Nunez. 1994. Bcl-xL is the major Bcl-x mRNA form expressed during murine development and its product localizes to mitochondria. Development 120:3033-3042.
  23. Jung, D. J., J. J. Bong and M. Baik. 2004. Extracellular proteinase inhibitor-accelerated apoptosis is associated with B cell activating factor in mammary epithelial cells. Exp. Cell Res. 292:115-122.
  24. Kim, S. B., J. S. Kim, J. H. Lee, W. J. Yoon, D. S. Lee, M. S. Ko, B. S. Kwon, D. H. Choi, H. R. Cho, B. J. Lee, D. K. Chung, H. W. Lee and J. W. Park. 2006. NF-kappaB activation is required for cisplatin-induced apoptosis in head and neck squamous carcinoma cells. FEBS Lett. 580:311-318.
  25. Ghosh, S., A. M. Gifford, L. R. Riviere, P. Tempst, G. P. Nolan and D. Baltimore. 1990. Cloning of the p50 DNA binding subunit of NF-$\kappa$B: homology to rel and dorsal. Cell 62:1019-1029.
  26. Thompson, J. S., S. A. Bixler, F. Qian, K. Vora, M. L. Scott, T. G. Cachero, C. Hession, P. Schneider, I. D. Sizing, C. Mullen, P. Strauch, M. Zafari, C. D. Benjamin, J. Tschopp, J. L. Browning and C. Ambrose. 2001. BAFF-R, a newly identified TNF receptor that specifically interacts with BAFF. Sci. 293:2108-2111.
  27. Lund, L. R., J. Romer, N. Thomasset, H. Solberg, C. Pyke, M. J. Bissell, K. Dano and Z. Werb. 1996. Two distinct phases of apoptosis in mammary gland involution: proteinaseindependent and dependent pathways. Development 122:181-193.
  28. Beg, A. A. and D. Baltimore. 1996. An essential role for NFkappaB in preventing TNF-alpha-induced cell death. Sci. 274:782-784.
  29. Schorr, K., M. Li, S. Krajewski, J. C. Reed and P. A. Furth. 1999. Bcl-2 gene family and related proteins in mammary gland involution and breast cancer. J. Mammary Gland Biol. Neoplasia 4:153-164.
  30. Lee, M. J., O. Han, K. Back, Y. J. Choi and M. G. Baik. 2001. Induction of lysozyme gene expression during involution of mouse mammary gland. Asian-Aust. J. Anim. Sci. 14:462-466.
  31. Laabi, Y., A. Egle and A. Strasser. 2001. TNF cytokine family: more BAFF-ling complexities. Current Biol. 11:R1013-1016.
  32. Ha, S. H., S. R. Lee, T. H. Lee, Y. M. Kim, M. Baik and Y. J. Choi. 2001. The expression of Bok is regulated by serum in HC11 mammary epithelial cells. Mol. Cells 12:368-371.
  33. Metcalfe, A. D., A. Gilmore, T. Klinowska, J. Oliver, A. J. Valentijn, R. Brown, A. Ross, G. MacGregor, J. A. Hickman and C. H. Streuli. 1999. Developmental regulation of Bcl-2 family protein expression in the involuting mammary gland. J. Cell Sci. 112(Pt 11):1771-1783.
  34. Li, M., X. Liu, G. Robinson, U. Bar-Peled, K. U. Wagner, W. S. Young, L. Hennighausen and P. A. Furth. 1997. Mammaryderived signals activated programmed cell death during the first stage of mammary gland involution. Proc. Natl. Acad. Sci. 94:3425-3430.
  35. Mackay, F. and C. Ambrose. 2003. The TNF family members BAFF and APRIL: the growing complexity. Cytokine and Growth Factor Reviews 14:311-324.
  36. Topper, Y. J. and C. S. Freeman. 1980. Multiple hormone interactions in the developmental biology of the mammary gland. Physiol. Rev. 60:1049-1106.
  37. Clarkson, R. W., J. L. Heeley, R. Chapman, F. Aillet, R. T. Hay, A. Wyllie and C. J. Watson. 2000. NF-kappaB inhibits apoptosis in murine mammary epithelia. J. Biol. Chem. 275:12737-12742.
  38. Yu, G., T. Boone, J. Delaney, N. Hawkins, M. Kelley, M. Ramakrishnan, S. McCabe, J. Qiu, M. Kornuc, X.-Z. Xia, J. Guo, M. Stolina, W. J. Boyle, I. Sarosi, H. Hsu, G. Senaldi and L. E. Theill. 2000. APRIL and TALL-I and receptors BCMA and TACI: system for regulating humoral immunity. Nat. Immunol. 1:252-256.
  39. Visconti, R., J. Cerutti, S. Battista, M. Fedele, F. Trapasso, K. Zeki, M. P. Miano, F. de Nigris, L. Casalino, F. Curcio, M. Santoro and A. Fusco. 1997. Expression of the neoplastic phenotype by human thyroid carcinoma cell lines requires NFkappaB p65 protein expression. Oncogene 15:1987-1994.
  40. Barger, S. W., D. Horster, K. Furukawa, Y. Goddman, J. Krieglestein and M. P. Mattson. 1995. Tumor necrosis factors alpha and beta protect neurons against amyloid beta-peptide toxicity: evidence for involvement of a kappa B-binding factor and attenuation of peroxide and $Ca^{2+}$ accumulation. Proc. Natl. Acad. Sci. USA 92:9328-9332.
  41. Rivera-Walsh, I., M. Waterfield, G. Xiao, A. Fong and S. C. Sun. 2001. NF-kappaB signaling pathway governs TRAIL gene expression and human T-cell leukemia virus-I Tax-induced Tcell death. J. Biol. Chem. 276:40385-40388.
  42. Dear, T. N., D. A. McDonald and R. F. Kefford. 1989. Transcriptional down-regulation of a gene, WDNM2, in metastatic DMBA-8 cells. Cancer Res. 49:5323-5328.
  43. Dear, T. N. and R. F. Kefford. 1991. The WDNM1 gene product is a novel member of the 'four-disulphide core' family of proteins. Biochem. Biophys. Res. Comm. 176:247-254.
  44. Roth, W., B. Wagenknecht, A. Klumpp, U. Naumann, M. Hahne, J. Tschopp and M. Weller. 2001. APRIL, a new member of the tumor necrosis factor family, modulates death ligand-induced apoptosis. Cell Death Differ. 8:403-410.
  45. Strange, R., F. Li, S. Saurer, A. Burkhardt and R. R. Friis. 1992. Apoptotic cell death and tissue remodelling during mouse mammary gland involution. Development 115:49-58.
  46. Kim, T. W., H. B. Moon and S. J. Kim. 2003. Interleukin-10 is upregulated by prolactin and serum-starvation in cultured mammary epithelial cells. Mol. Cells 16:168-172.
  47. Hunot, S., B. Brugg, D. Ricard, P. P. Michel, M. P. Muriel, M. Ruberg, B. A. Faucheux, Y. Agid and E. C. Hirsch. 1997. Nuclear translocation of NF-kappaB is increased in dopaminergic neurons of patients with parkinson disease. Proc. Natl. Acad. Sci. USA 94:7531-7536.
  48. Oltvai, Z. N., C. L. Milliman and S. J. Korsmeyer. 1993. Bcl-2 heterodimerizes in vivo with a conserved homolog, Bax, that accelerates programmed cell death. Cell 74:609-619.