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Rôle du facteur de transcription HNF1B dans la tubulogénèse rénale chez la souris

Abstract : Mammalian kidney is an essential excretory organ that regulates fluid balance, osmolarity and pH. It also ensures blood filtration to excrete metabolism end products and drugs. The initiation of definitive mammalian kidney development is marked by the emergence of the ureteric bud (UB) from the Wolffian Duct (WD). The UB subsequently undergoes a complex and stereotyped process of branching to give rise to the entire urinary collecting duct (CD) system and the ureter. As the UB undergoes branching morphogenesis, the tip cells control multiple events including mesenchymal-to-epithelial conversion and subsequent formation of regionalized nephrons, the filtering units of the kidney. The POUhomeodomain transcription factor HNF1B plays a critical role in the early differentiation of various organs including pancreas, liver and kidney. In humans, HNF1B heterozygous mutations cause the complex syndrome known as Renal Cysts and Diabetes, characterized by kidney, genital tracts and pancreas abnormalities as well as early onset of Diabetes. Our lab has previously shown that HNF1B is involved in early mouse kidney development for UB timing outgrowth and branching, as well as for induction of nephrogenesis. However, Hnf1b is also expressed during branching in UB and CDs, and at every nephrogenesis steps, suggesting a later and specific role during both processes. Given the reciprocal interactions between the UB and the metanephric mesenchyme, to discriminate the specific Hnf1b functions in these compartments, we inactivated this gene individually in those two tissues, through the use of appropriate Cre-recombinase mouse lines. Hnf1b-specific inactivation in nephron progenitors, using the Wnt4-EGFPCre mouse line, leads to mutant newborns death soon after birth, with mild hypoplastic kidneys that lack all nephron segments but exhibit rather correct UB branching. Mutant renal vesicles develop and polarize normally but fail to progress to correctly patterned “S” shaped bodies (SSB). This is associated with strong downregulation of the Notch pathway components Lfng, Dll1 and Jag1 and the Irx1/2 factors, which are potential regulators of proximal and Henle’s loop segment fates. Moreover, HNF1B is recruited in vivo to the regulatory sequences of most of these genes. Overexpression of a HNF1B dominant-negative construct in Xenopus embryos causes downregulation specifically of proximal and intermediate pronephric segment markers. Our results show that HNF1B is required for the acquisition of a proximointermediate segment fate in vertebrates, thus uncovering a previously unappreciated function of a novel SSB subcompartment in global nephron segmentation and further differentiation. By removal of Hnf1b from the WD and the UB using the Hoxb7-Cre mouse line, we observe multiple urogenital tract abnormalities in part due to an early mosaic Hoxb7-Cre activity. Analyzing kidney tree architecture, we found that tips number and length are massively reduced and UB branching is severely miss-patterned in mutants. Combining the Hnf1b mosaic deletion with a reporter line expressing a membrane-associated fluorescent protein EGFP or Tomato, we developped an original genetic approach that allows visualize the behavior of recombined and WT cells within the UB branches. By time lapse on organotypic kidney cultures we observe, at different stages, that cells lacking Hnf1b become excluded from the UB tips. This suggests that HNF1B may be required cell autonomously at the tip domain for the branching process. Moreover, collecting duct cell polarity appears impaired in mutants and do not further maturate properly becoming either dilated or over cystic both in vivo and in vitro [...]
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https://tel.archives-ouvertes.fr/tel-03118983
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Submitted on : Friday, January 22, 2021 - 5:10:12 PM
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  • HAL Id : tel-03118983, version 1

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Audrey Desgrange. Rôle du facteur de transcription HNF1B dans la tubulogénèse rénale chez la souris. Biologie du développement. Université Pierre et Marie Curie - Paris VI, 2015. Français. ⟨NNT : 2015PA066741⟩. ⟨tel-03118983⟩

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