Myocardial-specific R-spondin3 drives proliferation of the coronary stems primarily through the Leucine Rich Repeat G Protein coupled receptor LGR4

Coronary artery anomalies are common congenital disorders with serious consequences in adult life. Coronary circulation begins when the coronary stems form connections between the aorta and the developing vascular plexus. We recently identified the WNT signaling modulator R-spondin 3 (Rspo3), as a crucial regulator of coronary stem proliferation. Using expression analysis and tissue-specific deletion we now demonstrate that Rspo3 is primarily produced by cardiomyocytes. Moreover, we have employed CRISPR/Cas9 technology to generate novel Lgr4-null alleles that showed a significant decrease in coronary stem proliferation and thus phenocopied the coronary artery defects seen in Rspo3 mutants. Interestingly, Lgr4 mutants displayed slightly hypomorphic right ventricles, an observation also made after myocardial specific deletion of Rspo3. These results shed new light on the role of Rspo3 in heart development and demonstrate that LGR4 is the principal Rspondin 3 receptor in the heart

Vital functions of WT1 during renal progenitor life

Le développement du rein est un exemple intriguant d’un équilibre délicat entre la prolifération des cellules progénitrices, la différentiation et l’apoptose. Le gène Wt1 est indispensable pour la survie des cellules progénitrices. Le but de cette thèse a été de définir les voies de signalisation activées par Wt1 pendant le développement du rein. En utilisant les souris Wt1 KO, nous avons démontré que WT1 coordonne l’action de deux voies de signalisation opposées : Fgf et Bmp/Smad intervenant dans la survie des cellules progénitrices rénales. Dans une deuxième étude, nous avons analysé le rôle du modificateur épigénétique, le gène Phf19 pendant le développement du rein. Nous avons démontré que l’expression de ce gène est Wt1-dependant et il est exclusivement exprimé dans les cellules progénitrices rénales au cours du développement et que son inactivation dans le rein embryonnaire en culture, conduit à l’apoptose des cellules progénitrices. Nous avons généré des souris knockout de Phf19 par l’approche de CRISPR/Cas9. Dans le cas d’une létalité précoce des embryons homozygotes, nous opterons pour la production du model animal knockout conditionnel et procéderons à la caractérisation de leur profile épigénétique. Cette thèse a permis d’une part, de découvrir deux voies de signalisation antagonistes, régulées par le Wt1et impliquées dans le contrôle de la survie des cellules progénitrices rénales et d’autre part de nous orienter vers le contrôle de la survie et la prolifération de ces cellules par modifications épigénétiques. Ceci nous permettra de contribuer à la connaissance de l’étiologie d’une grande proportion des malformations rénales restant à ce jour inconnues.Kidney organogenesis requires the tight control of proliferation, differentiation and apoptosis of renal progenitor cells. The Wilms’ tumour suppressor Wt1 is required for renal progenitor survival. The aim of this thesis was to elucidate the molecular cause for renal agenesis in Wt1 mutant mouse. Here we demonstrate that lack of Wt1 abolishes FGF and induces BMP/pSMAD signaling within the metanephric mesenchyme. We further show that recombinant BMP4, but not BMP7, induces an apoptotic response within the early kidney that can be suppressed by simultaneous addition of FGFs. These data reveal an unknown sensitivity of early renal progenitors to pSMAD signalling, establishes FGF and pSMAD signalling as antagonistic forces in early kidney development and places WT1 as a key regulator of pro-survival FGF signalling pathway genes. In a second study, we demonstrated, that Phf19, an epigenetic modifier, is essential both for maintaining Wt1 expression in renal progenitor cells and their survival in an ex-vivo culture. We further generated a Phf19 knockout mouse by CRISPR/Cas9. The homozygous embryos will be analyzed to further decipher the contribution of Phf19 to potential kidney malformations and the epigenetic profile of renal progenitor cells will be characterized. Overall, the new insights into the molecular mechanisms controlling the survival of renal progenitor cells, reported in this thesis, provide one more step in our understanding of renal malformations. In addition, our results conducted us toward the epigenetique modifications that could open up promising new avenue of understanding the etiology of an important proportion of renal malformation that remains unknown

The Adult Adrenal Cortex Undergoes Rapid Tissue Renewal in a Sex-Specific Manner

International audienceEvolution has resulted in profound differences between males and females that extend to non-reproductive organs and are reflected in the susceptibility and progression of diseases. However, the cellular and molecular basis for these differences remains largely unknown. Here we report that adrenal gland tissue renewal is highly active and sexually dimorphic, with female mice showing a 3-fold higher turnover than males. Moreover, in males, homeostasis relies on proliferation of cells within the steroidogenic zone, but females employ an additional stem and/or progenitor compartment situated in the adrenal capsule. Using lineage tracing, sex reversal models, gonadectomy, and dihydrotestosterone treatments, we further show that sex-specific stem cell activity is driven by male hormones that repress recruitment of Gli1+ stem cells from the capsule and cell proliferation. Taken together, our findings provide a molecular and cellular basis for adrenal sex dimorphism that may contribute to the increased incidence of adrenal diseases in females

R-spondin signalling is essential for the maintenance and differentiation of mouse nephron progenitors

During kidney development, WNT/β-catenin signalling has to be tightly controlled to ensure proliferation and differentiation of nephron progenitor cells. Here we show in mice that the signalling molecules RSPO1 and RSPO3 act in a functionally redundant manner to permit WNT/β-catenin signalling and their genetic deletion leads to a rapid decline of nephron progenitors. By contrast, tissue specific deletion in cap mesenchymal cells abolishes mesenchyme to epithelial transition (MET) that is linked to a loss of Bmp7 expression, absence of SMAD1/5 phosphorylation and a concomitant failure to activate Lef1, Fgf8 and Wnt4, thus explaining the observed phenotype on a molecular level. Surprisingly, the full knockout of LGR4/5/6, the cognate receptors of R-spondins, only mildly affects progenitor numbers, but does not interfere with MET. Taken together our data demonstrate key roles for R-spondins in permitting stem cell maintenance and differentiation and reveal Lgr-dependent and independent functions for these ligands during kidney formation

WT1 Maintains Adrenal-Gonadal Primordium Identity and Marks a Population of AGP-like Progenitors within the Adrenal Gland

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WT1 controls antagonistic FGF and BMP-pSMAD pathways in early renal progenitors.

International audienceKidney organogenesis requires the tight control of proliferation, differentiation and apoptosis of renal progenitor cells. How the balance between these cellular decisions is achieved remains elusive. The Wilms' tumour suppressor Wt1 is required for progenitor survival, but the molecular cause for renal agenesis in mutants is poorly understood. Here we demonstrate that lack of Wt1 abolishes fibroblast growth factor (FGF) and induces BMP/pSMAD signalling within the metanephric mesenchyme. Addition of recombinant FGFs or inhibition of pSMAD signalling rescues progenitor cell apoptosis induced by the loss of Wt1. We further show that recombinant BMP4, but not BMP7, induces an apoptotic response within the early kidney that can be suppressed by simultaneous addition of FGFs. These data reveal a hitherto unknown sensitivity of early renal progenitors to pSMAD signalling, establishes FGF and pSMAD signalling as antagonistic forces in early kidney development and places WT1 as a key regulator of pro-survival FGF signalling pathway genes

Retinoic acid signaling is directly activated in cardiomyocytes and protects mouse hearts from apoptosis after myocardial infarction

Retinoic acid (RA) is an essential signaling molecule for cardiac development and plays a protective role in the heart after myocardial infarction (MI). In both cases, the effect of RA signaling on cardiomyocytes, the principle cell type of the heart, has been reported to be indirect. Here we have developed an inducible murine transgenic RA-reporter line using CreER(T2) technology that permits lineage tracing of RA-responsive cells and faithfully recapitulates endogenous RA activity in multiple organs during embryonic development. Strikingly, we have observed a direct RA response in cardiomyocytes during mid-late gestation and after MI. Ablation of RA signaling through deletion of the Aldh1a1/a2/a3 genes encoding RA-synthesizing enzymes leads to increased cardiomyocyte apoptosis in adults subjected to MI. RNA sequencing analysis reveals Tgm2 and Ace1, two genes with well-established links to cardiac repair, as potential targets of RA signaling in primary cardiomyocytes, thereby providing novel links between the RA pathway and heart disease