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.
- transgenic mice
- uromodulin promoter
- targeting recombinant proteins into urine
Abbreviations used: rh α1AT, recombinant human α1-antitrypsin; GH, growth hormone; GM-CSF, granulocyte—macrophage colony-stimulating factor; UPII, uroplakin II; PNGase F, peptide N-glycosidase F; PPE, porcine pancreatic elastase; ARC, American Red Cross.
- The Biochemical Society, London ©2002