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TRPV5‐promotor driven expression of EGFP produces fluorescence in late DCT, CNT and early CCD
Author(s) -
Praetorius Jeppe,
Andrew Fenton Robert,
Füechtbauer ErnstMartin,
Vind Hofmeister Marlene
Publication year - 2008
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.22.1_supplement.1159.16
Subject(s) - green fluorescent protein , immunolabeling , transgene , microbiology and biotechnology , biology , reabsorption , genetically modified mouse , immunohistochemistry , kidney , gene , genetics , immunology
TRPV5 (ECaC) is an apical Ca 2+ channel of renal late distal convoluted tubules and connecting tubules (DCT2 and CNT, respectively), and is critically involved in regulated Ca 2+ reabsorption. The protein was previously localized to CNT by immunohistochemistry and RT‐PCR analysis. Genetic disruption of TRPV5 leads to hypercalciuria. Transgenic mice were developed that express EGFP driven by the TRPV5 gene promoter 1) to verify and expand the expression‐profile of TRPV5 and 2) to generate a cell line of the renal tubules expressing this protein as a tool for further studies of TRPV5 biology. TRPV5 promoter‐driven EGFP fluorescence was detected in late DCT2 and CNT in isolated living tubules by confocal microscopy. In addition, EGFP was expressed in some cortical collecting ducts (CCD). Robust EGFP signal was confirmed in sections of paraffin embedded kidney, and the expression of EGFP in both late DCT2/CNT and CCD was verified by double‐fluorescence immunolabeling with various tubule markers. In preliminary analysis, TRPV5 protein expression in DCT2/CNT as well as in CCD using anti‐TRPV5 antibodies coincided with the expression of TRPV5 promoter‐driven EGFP. Thus, TRPV5 is expressed in DCT2/CNT and CCD, and an EGFP‐expressing transgenic mouse has been generated enabling easy isolation of TRPV5‐expressing Ca 2+ ‐transporting renal tubules. Supported by the Danish National Research Foundation.