Mammary Gland-Specific Expression of Biologically Active Human Osteoprotegerin in Transgenic Mice

We have recently shown that the regulatory sequence of the uromodulin gene, containing the 3.7kb promoter, exon 1 and a part of exon 2, provided for kidney-specific expression of the reporter lacZ gene in transgenic mice [Zbikowska, Soukhareva, Behnam, Chang, Drews, Lubon, Hammond and Soukharev (2002) Transgenic Res., in the press]. In the present study, we generated transgenic mice harbouring the regulatory sequence of the uromodulin gene to direct the expression of human α1-antitrypsin (α1AT) into urine. Of the 13 founder mice that tested positive by PCR, seven showed the presence of the human protein in their urine. The concentration of the recombinant human (rh) α1AT in the urine, estimated by using ELISA, ranged from 0.5 to 14μg/ml in the F0-generation mice, and reached up to 65μg/ml in the F1 generation. The transgenically produced rh α1AT was found to be N-glycosylated and biologically active. The N-terminal sequence analysis confirmed the identity of the human protein and revealed that the recombinant α1AT was correctly processed with the signal peptide cleaved off. Our results demonstrate for the first time that the uromodulin regulatory sequence provides a very attractive option for the potential large-scale production of functional therapeutic proteins in livestock.

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Differential regulation of rat beta-casein-chloramphenicol acetyltransferase fusion gene expression in transgenic mice

Molecular and Cellular Biology ◽

10.1128/mcb.9.2.560-565.1989 ◽

1989 ◽

Vol 9 (2) ◽

pp. 560-565

Author(s):

K F Lee ◽

S H Atiee ◽

J M Rosen

Keyword(s):

Gene Expression ◽

Mammary Gland ◽

Transgenic Mice ◽

Fusion Gene ◽

Chloramphenicol Acetyltransferase ◽

Fusion Genes ◽

Specific Expression ◽

Casein Gene ◽

Noncoding Exon ◽

Beta Casein

Previous studies in our laboratory have demonstrated the mammary-specific expression of the entire rat beta-casein gene with 3.5 kilobases (kb) of 5' and 3.0 kb of 3' DNA in transgenic mice (Lee et al., Nucleic Acids Res. 16:1027-1041, 1988). In an attempt to localize sequences that dictate this specificity, lines of transgenic mice carrying two different rat beta-casein promoter-bacterial chloramphenicol acetyltransferase (cat) fusion genes have been established. Twenty and eight lines of transgenic mice carrying two fusion genes containing either 2.3 or 0.5 kb, respectively, of 5'-flanking DNA of the rat beta-casein gene along with noncoding exon I and 0.5 kb of intron A were identified, most of which transmitted the transgenes to their offspring in a Mendelian pattern. CAT activity was detected predominantly in the lactating mammary gland of female transgenic mice but not in the male mammary fat pad. A several-hundred-fold variation in the level of cat expression was observed in the mammary gland of different lines of mice, presumably due to the site of integration of the transgenes. CAT activity was increased in the mammary gland during development from virgin to midpregnancy and lactation. Unexpectedly, the casein-cat transgenes were also expressed in the thymus of different lines of both male and female mice, in some cases at levels equivalent to those observed in the mammary gland, and in contrast to the mammary gland, CAT activity was decreased during pregnancy and lactation in the thymus. Thus, 0.5 kb of 5'-flanking DNA of the rat beta-casein gene along with noncoding exon I and 0.5 kb of intron A are sufficient to target bacterial cat gene expression to the mammary gland of lactating mice.

Download Full-text

Differential regulation of rat beta-casein-chloramphenicol acetyltransferase fusion gene expression in transgenic mice.

Molecular and Cellular Biology ◽

10.1128/mcb.9.2.560 ◽

1989 ◽

Vol 9 (2) ◽

pp. 560-565 ◽

Cited By ~ 32

Author(s):

K F Lee ◽

S H Atiee ◽

J M Rosen

Keyword(s):

Gene Expression ◽

Mammary Gland ◽

Transgenic Mice ◽

Fusion Gene ◽

Chloramphenicol Acetyltransferase ◽

Fusion Genes ◽

Specific Expression ◽

Casein Gene ◽

Noncoding Exon ◽

Beta Casein

Previous studies in our laboratory have demonstrated the mammary-specific expression of the entire rat beta-casein gene with 3.5 kilobases (kb) of 5' and 3.0 kb of 3' DNA in transgenic mice (Lee et al., Nucleic Acids Res. 16:1027-1041, 1988). In an attempt to localize sequences that dictate this specificity, lines of transgenic mice carrying two different rat beta-casein promoter-bacterial chloramphenicol acetyltransferase (cat) fusion genes have been established. Twenty and eight lines of transgenic mice carrying two fusion genes containing either 2.3 or 0.5 kb, respectively, of 5'-flanking DNA of the rat beta-casein gene along with noncoding exon I and 0.5 kb of intron A were identified, most of which transmitted the transgenes to their offspring in a Mendelian pattern. CAT activity was detected predominantly in the lactating mammary gland of female transgenic mice but not in the male mammary fat pad. A several-hundred-fold variation in the level of cat expression was observed in the mammary gland of different lines of mice, presumably due to the site of integration of the transgenes. CAT activity was increased in the mammary gland during development from virgin to midpregnancy and lactation. Unexpectedly, the casein-cat transgenes were also expressed in the thymus of different lines of both male and female mice, in some cases at levels equivalent to those observed in the mammary gland, and in contrast to the mammary gland, CAT activity was decreased during pregnancy and lactation in the thymus. Thus, 0.5 kb of 5'-flanking DNA of the rat beta-casein gene along with noncoding exon I and 0.5 kb of intron A are sufficient to target bacterial cat gene expression to the mammary gland of lactating mice.

Download Full-text