Genetic manipulation of ureteric bud tip progenitors in the mammalian kidney through an Adamts18 enhancer driven tet-on inducible system
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1.
University of Southern California
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2.
ETH Zurich
Abstract
The ureteric epithelial progenitor (UEP) population within the embryonic kidney generates the arborized epithelial network of the kidney’s collecting system and plays a critical role in the expansion and induction of the surrounding nephron progenitor pool. Adamts18 shows UEP- restricted expression in the kidney and progenitor tip-restricted expression in several other organs undergoing branching epithelial growth. Adamts18 is encoded by 23 exons. Genetic removal of genomic sequence spanning exons 1 to 3 led to a specific loss of Adamts18 expression in UEPs, suggesting this region may encode a UEP-specific enhancer. Intron 2 (3 kb) was shown to have enhancer activity driving expression of the doxycycline inducible tet-on transcriptional regulator (rtTA) in an Adamts18en-rtTA transgenic mouse strain. Crossing Adamts18en-rtTA mice to a doxycycline dependent GFP reporter mouse enabled the live imaging of embryonic kidney explants. This facilitated the analysis of ureteric epithelial branching events at the cellular level. Ablation of UEPs at the initiation of ureteric bud outgrowth through the doxycycline-mediated induction of Diphtheria Toxin A (DTA) generated a range of phenotypes from complete kidneys agenesis, to duplex kidneys with double ureters. The latter outcome points to the potential of regulative processes to restore UEPs. In contrast, overexpression of YAP prior to ureteric bud outgrowth led to a complete failure of kidney development. Elevating YAP levels at later stages retarded branching growth. A similar phenotype was observed with the overexpression of MYC within the branch-tip localized UEP population. These experiments showcase the utility of the Adamts18en-rtTA transgenic model to the investigation of cellular and molecular events specific to branch tip progenitors within the mammalian kidney complementing existing CRE-dependent genetic tools. Further, the illustrative examples point to areas where new insight may be gained into the regulation of UEP programs.
Copyright and License
© 2019 Elsevier Inc. All rights reserved.
Acknowledgement
We would like to be thank Dr. Ben Stanger for graciously providing the TRE-YAP mice. We thank Seth Ruffins for his assistance with microscopy and imaging analysis programs. We would like to thank Gohar Saribekyan and Junji Watanabe for paraffin sectioning, and Richard Lopez and Himmat Sahi for their assistance with data collection. We would like to thank Riana Parvez for generating the volume projections.
Funding
Research in the A.P.M. laboratory was funded by a grant from the National Institutes of Health [DK054364]. E.A.R. was supported by a graduate student training fellowship from the National Institutes of Health [5T32HD060549] and by the National Institute of Diabetes and Digestive and Kidney Diseases of the National Institutes of Health [F31DK107216]. These sources of funding were not involved in the study design, data collection, data analysis, writing the report, or the decision to submit for publication.
Contributions
E.A.R., N.O.L., O.M., and A.P.M planned experiments, collected data, and analyzed results; E.A.R. assembled the figures; and E.A.R. and A.P.M. wrote the manuscript with inputs from other authors.
Data Availability
Data not available / Data will be made available on request
Supplemental Material
Suppleentary data is attached.
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Additional details
- National Institute of Diabetes and Digestive and Kidney Diseases
- F31DK107216
- National Institutes of Health
- DK054364
- National Institutes of Health
- 5T32HD060549
- Caltech groups
- Division of Biology and Biological Engineering
- Publication Status
- Published