DOI QR코드

DOI QR Code

Cloning and Molecular Characterization of Porcine β-casein Gene (CNS2)

  • Lee, Sang-Mi (Department of Animal Science, Chonnam National University) ;
  • Kim, Hye-Min (Department of Animal Science, Chonnam National University) ;
  • Moon, Seung-Ju (Department of Animal Science, Chonnam National University) ;
  • Kang, Man-Jong (Department of Animal Science, Chonnam National University)
  • Received : 2011.07.23
  • Accepted : 2011.11.03
  • Published : 2012.03.01

Abstract

The production of therapeutic proteins from transgenic animals is one of the most important successes of animal biotechnology. Milk is presently the most mature system for production of therapeutic proteins from a transgenic animal. Specifically, ${\beta}$-casein is a major component of cow, goat and sheep milk, and its promoter has been used to regulate the expression of transgenic genes in the mammary gland of transgenic animals. Here, we cloned the porcine ${\beta}$-casein gene and analyzed the transcriptional activity of the promoter and intron 1 region of the porcine ${\beta}$-casein gene. Sequence inspection of the 5'-flanking region revealed potential DNA elements including SRY, CdxA, AML-a, GATA-3, GATA-1 and C/EBP ${\beta}$. In addition, the first intron of the porcine ${\beta}$-casein gene contained the transcriptional enhancers Oct-1, SRY, YY1, C/EBP ${\beta}$, and AP-1, as well as the retroviral TATA box. We estimated the transcriptional activity for the 5'-proximal region with or without intron 1 of the porcine ${\beta}$-casein gene in HC11 cells stimulated with lactogenic hormones. High transcriptional activity was obtained for the 5'-proximal region with intron 1 of the porcine ${\beta}$-casein gene. The ${\beta}$-casein gene containing the mutant TATA box (CATAAAA) was also cloned from another individual pig. Promoter activity of the luciferase vector containing the mutant TATA box was weaker than the same vector containing the normal TATA box. Taken together, these findings suggest that the transcription of porcine ${\beta}$-casein gene is regulated by lactogenic hormone via intron 1 and promoter containing a mutant TATA box (CATAAAA) has poor porcine ${\beta}$-casein gene activity.

Keywords

Lactogenic Hormones;HC11 Cells;TATA Box;Milk Protein;Porcine Casein Gene

References

  1. Alexander, L. J. and C. W. Beattie. 1992. The sequence of porcine beta-casein cDNA. Anim. Genet. 23:369-371. https://doi.org/10.1111/j.1365-2052.1992.tb00160.x
  2. Baev, A. A., I. K. Smirnov and S. I. Gorodetskii. 1987. Primary structure of cDNA for bovine beta-casein. Mol. Biol. 21:255-265.
  3. Boudreau, N., C. Myers and M. J. Bissell. 1995. From laminin to lamin: Regulation of tissue-specific gene expression by the ECM. Trends Cell Biol. 5:1-4. https://doi.org/10.1016/S0962-8924(00)88924-2
  4. Chang, M., J. W. Lee, D. B. Koo, S. T. Shin and Y. M. Han. 2010. Knocking-in of the human thrombopoietin gene on beta-casein locus in bovine fibroblasts. Asian-Aust. J. Anim. Sci. 23:806-813. https://doi.org/10.5713/ajas.2010.90501
  5. Clark, A. J. 1998. The mammary gland as a bioreactor: expression, processing, and production of recombinant proteins. J. Mammary Gland Biol. Neoplasia 3:337-350. https://doi.org/10.1023/A:1018723712996
  6. Doppler, W., B. Groner and R. K. Ball. 1989. Prolactin and glucocorticoid hormones synergistically induce expression of transfected rat beta-casein gene promoter constructs in a mammary epithelial cell line. Proc. Natl. Acad. Sci. USA. 86:104-108. https://doi.org/10.1073/pnas.86.1.104
  7. Gorodetsky, S. I., T. M. Tkach and T. V. Kapelinskaya. 1988. Isolation and characterization of the Bos taurus beta-casein gene. Gene 66:87-96. https://doi.org/10.1016/0378-1119(88)90227-2
  8. Ho, M. Y. and D. Murphy. 2002. The vasopressin gene non-canonical Hogness box: effect on protein binding and promoter function. Mol. Cell. Endocrinol. 186:17-25. https://doi.org/10.1016/S0303-7207(01)00677-3
  9. Houdebine, L. M. 2009. Production of pharmaceutical proteins by transgenic animals. Comp. Immunol. Microbiol. Infect. Dis. 32:107-121. https://doi.org/10.1016/j.cimid.2007.11.005
  10. Kang, Y. K., C. S. Lee, A. S. Chung and K. K. Lee. 1998. Prolactin-inducible enhancer activity of the first intron of the bovine beta-casein gene. Mol. Cells 30:259-265.
  11. Kolb, A. 2003. The first intron of the murine $\beta$-casein gene contains a functional promoter. Biochem. Biophys. Res. Commun. 306:1099-1105. https://doi.org/10.1016/S0006-291X(03)01104-5
  12. Lee, P., H. K. Chung, H. G. Lee, H. C. Lee, J. S. Woo, S. Lee, S. J. Jo, W. K. Chang, H. T. Lee, M. Kwon and J. K. Park. 2008. Cloning and characterization of 5'-untranslated region of porcine beta casein gene (CSN2). Domest. Anim. Endocrinol. 35:245-253. https://doi.org/10.1016/j.domaniend.2008.04.001
  13. Mercier, J. C. and J. L. Viloite. 1993. Structure and function of milk protein genes. J. Dairy Sci. 76:3079-3098. https://doi.org/10.3168/jds.S0022-0302(93)77647-X
  14. Rijnkels, M. 2002. Multispecies comparison of the casein gene loci and evolution of casein gene family. J. Mammary Gland Biol. Neoplasia 7:327-345. https://doi.org/10.1023/A:1022808918013
  15. Rosen, J. M., S. L. Wyszomierski and D. Hadsell. 1999. Regulation of milk protein gene expression. Annu. Rev. Nutr. 19:407-436. https://doi.org/10.1146/annurev.nutr.19.1.407
  16. Schmitt-Ney, M., W. Doppler, R. K. Ball and B. Groner. 1991. Beta-casein gene promoter activity is regulated by the hormone-mediated relief of transcriptional repression and a mammary-gland-specific nuclear factor. Mol. Cell Biol. 11:3745-3755.
  17. Wheeler, M. B. and E. M. Walters. 2001. Transgenic technology and application in swine. Theriogenology 56:1345-1369. https://doi.org/10.1016/S0093-691X(01)00635-5
  18. Xu, L. C., M. Thali and W. Schaffner. 1991. Upstream box/TATA box order is the major determinant of the direction of transcription. Nucleic Acids Res. 19:6699-6704. https://doi.org/10.1093/nar/19.24.6699