The use of the uromodulin promoter to target production of recombinant proteins into urine of transgenic animals.
A uromodulin promoter has been isolated, sequenced, and used to generate two sets of transgenic mice for expression of the lacZ marker gene and for production of the human recombinant erythropoietin (rhEPO) in urine. We demonstrated that the 5.6-kb fragment of the uromodulin gene containing the 3.7-kb promoter area and, both the first exon and part of the second exon, were sufficient to provide kidney-specific expression of the lacZ gene. Histological analysis of the lacZ expression pattern revealed beta-galactosidase activity specifically in the thick limb of Henle's loop. However, due to random integration of the transgene, ectopic expression was detected in some transgenic lines. Analysis of the EPO-transgenic mice showed that rhEPO was secreted into the urine of founder mice (up to 6 ng/ml). We were able to breed and analyze only two sublines with a very low expression level of rhEPO (up to 260 pg/ml). All of our transgenic mice expressing rhEPO in urine developed disease symptoms similar to polycythemia in humans. These included a considerable increase in red blood cell counts, hemoglobin concentration, and hematocrit concomitant with severe thrombocytopenia, all of which were detected in the rhEPO-expressing mice. Although our model did not prove to be beneficial for commercial production of rhEPO, we concluded that the uromodulin promoter could be useful for expression of other important therapeutic proteins into the urine of transgenic animals.
Zbikowska
Halina M
HM
Plasma Derivative Department, Holland Laboratory, American Red Cross, MD 20855, Rockville, USA.
Soukhareva
Nadia
N
Behnam
Reza
R
Chang
Rosemary
R
Drews
Roman
R
Lubon
Henryk
H
Hammond
David
D
Soukharev
Serguei
S
D000818:Animals; D017931:DNA Primers; D004921:Erythropoietin / Q000097:blood / Q000235:genetics / Q000652:urine*; D005091:Exons; D005260:Female; D005786:Gene Expression Regulation; D006801:Humans; D007668:Kidney / Q000502:physiology; D051379:Mice; D008822:Mice, Transgenic; D009088:Mucoproteins / Q000235:genetics*; D009928:Organ Specificity; D016133:Polymerase Chain Reaction; D011401:Promoter Regions, Genetic; D011994:Recombinant Proteins / Q000652:urine*; D016385:TATA Box; D058949:Uromodulin; D001616:beta-Galactosidase / Q000235:genetics
D017931:DNA Primers; D009088:Mucoproteins; D011994:Recombinant Proteins; C548612:UMOD protein, human; C548613:Umod protein, mouse; D058949:Uromodulin; D004921:Erythropoietin; D001616:beta-Galactosidase