A miRNA-Based Prognostic Model to Trace Thyroid Cancer Recurrence.

Papillary thyroid carcinomas (PTCs) account for most endocrine tumors; however, screening and diagnosing the recurrence of PTC remains a clinical challenge. Using microRNA sequencing (miR-seq) to explore miRNA expression profiles in PTC tissues and adjacent normal tissues, we aimed to determine which miRNAs may be associated with PTC recurrence and metastasis. Public databases such as TCGA and GEO were utilized for data sourcing and external validation, respectively, and miR-seq results were validated using quantitative real-time PCR (qRT-PCR). We found miR-145 to be significantly downregulated in tumor tissues and blood. Deregulation was significantly related to clinicopathological features of PTC patients including tumor size, lymph node metastasis, TNM stage, and recurrence. In silico data analysis showed that miR-145 can negatively regulate multiple genes in the TC signaling pathway and was associated with cell apoptosis, proliferation, stem cell differentiation, angiogenesis, and metastasis. Taken together, the current study suggests that miR-145 may be a biomarker for PTC recurrence. Further mechanistic studies are required to uncover its cellular roles in this regard.

Targeted disruption of the histone lysine 79 methyltransferase Dot1L in nephron progenitors causes congenital renal dysplasia.

The epigenetic regulator Dot1, the only known histone H3K79 methyltransferase, has a conserved role in organismal development and homoeostasis. In yeast, Dot1 is required for telomeric silencing and genomic integrity. In Drosophila, Dot1 (Grappa) regulates homoeotic gene expression. Dysregulation of DOT1L (human homologue of Dot1) causes leukaemia and is implicated in dilated cardiomyopathy. In mice, germline disruption of Dot1L and loss of H3K79me2 disrupt vascular and haematopoietic development. Targeted inactivation of Dot1L in principal cells of the mature collecting duct affects terminal differentiation and cell type patterning. However, the role of H3K79 methylation in mammalian tissue development has been questioned, as it is dispensable in the intestinal epithelium, a rapidly proliferating tissue. Here, we used lineage-specific Cre recombinase to delineate the role of Dot1L methyltransferase activity in the mouse metanephric kidney, an organ that develops via interactions between ureteric epithelial (Hoxb7) and mesenchymal (Six2) cell lineages. The results demonstrate that Dot1LHoxb7 is dispensable for ureteric bud branching morphogenesis. In contrast, Dot1LSix2 is critical for the maintenance and differentiation of Six2+ progenitors into epithelial nephrons. Dot1LSix2 mutant kidneys exhibit congenital nephron deficit and cystic dysplastic kidney disease. Molecular analysis implicates defects in key renal developmental regulators, such as Lhx1, Pax2 and Notch. We conclude that the developmental functions of Dot1L-H3K79 methylation in the kidney are lineage-restricted. The link between H3K79me and renal developmental pathways reaffirms the importance of chromatin-based mechanisms in organogenesis.

Mdm4 controls ureteric bud branching via regulation of p53 activity.

The antagonism between Mdm2 and its close homolog Mdm4 (also known as MdmX) and p53 is vital for embryogenesis and organogenesis. Previously, we demonstrated that targeted disruption of Mdm2 in the Hoxb7+ ureteric bud (Ub) lineage, which gives rise to the renal collecting system, causes renal hypodysplasia culminating in perinatal lethality. In this study, we examine the unique role of Mdm4 in establishing the collecting duct system of the murine kidney. Hoxb7Cre driven loss of Mdm4 in the Ub lineage (UbMdm4-/-) disrupts branching morphogenesis and triggers UB cell apoptosis. UbMdm4-/- kidneys exhibit abnormally dilated Ub tips while the medulla is hypoplastic. These structural alterations result in secondary depletion of nephron progenitors and nascent nephrons. As a result, newborn UbMdm4-/- mice have hypo-dysplastic kidneys. Transcriptional profiling revealed downregulation of the Ret-tyrosine kinase pathway components, Gdnf, Wnt11, Sox8, Etv4 and Cxcr4 in the UbMdm4-/- mice relative to controls. Moreover, the expression levels of the canonical Wnt signaling members Axin2 and Wnt9b are downregulated. Mdm4 deletion upregulated p53 activity and p53-target gene expression including Cdkn1a (p21), Gdf15, Ccng1, PERP, and Fas. Germline loss of p53 in UbMdm4-/- mice largely rescues kidney development and terminal differentiation of the collecting duct. We conclude that Mdm4 plays a unique and vital role in Ub branching morphogenesis and collecting system development.

Histone deacetylases 1 and 2 regulate the transcriptional programs of nephron progenitors and renal vesicles.

Nephron progenitor cells (NPCs) are Six2-positive metanephric mesenchyme cells, which undergo self-renewal and differentiation to give rise to nephrons until the end of nephrogenesis. Histone deacetylases (HDACs) are a group of epigenetic regulators that control cell fate, but their role in balancing NPC renewal and differentiation is unknown. Here, we report that NPC-specific deletion of Hdac1 and Hdac2 genes in mice results in early postnatal lethality owing to renal hypodysplasia and loss of NPCs. HDAC1/2 interact with the NPC renewal regulators Six2, Osr1 and Sall1, and are co-bound along with Six2 on the Six2 enhancer. Although the mutant NPCs differentiate into renal vesicles (RVs), Hdac1/2 mutant kidneys lack nascent nephrons or mature glomeruli, a phenocopy of Lhx1 mutants. Transcriptional profiling and network analysis identified disrupted expression of Lhx1 and its downstream genes, Dll1 and Hnf1a/4a, as key mediators of the renal phenotype. Finally, although HDAC1/2-deficient NPCs and RVs overexpress hyperacetylated p53, Trp53 deletion failed to rescue the renal dysgenesis. We conclude that the epigenetic regulators HDAC1 and HDAC2 control nephrogenesis via interactions with the transcriptional programs of nephron progenitors and renal vesicles.

Regulation of kidney development by the Mdm2/Mdm4-p53 axis.

While p53 activity is required for tumour suppression, unconstrained p53 activity on the other hand is detrimental to the organism, resulting in inappropriate cellular death or proliferation defects. Unimpeded p53 activity is lethal in the developing embryo, underlining the need for maintaining a tight control on p53 activity during this period. The critical role of the negative regulators of p53, Mdm2 and Mdm4, in vertebrate development came to light by fatal disruption of embryogenesis that was observed with Mdm2 and Mdm4 gene deletions in mice. Embryonic lethality was rescued only by superimposing p53 removal. Here we summarize the contribution of the Mdm2/Mdm4-p53 axis that occurs at multiple steps of kidney development. Conditional, cell type-specific deletions reveal distinct functions of these proteins in renal morphogenesis. The severe impact on the renal phenotype from targeted gene deletions underscores the critical role played by the Mdm2/Mdm4-p53 nexus on nephrogenesis, and emphasizes the need to monitor patients with aberrations in this pathway for kidney function defects and associated cardiovascular dysfunction.

Prorenin receptor controls renal branching morphogenesis via Wnt/ß-catenin signaling.

The prorenin receptor (PRR) is a receptor for renin and prorenin, and an accessory subunit of the vacuolar proton pump H+-ATPase. Renal branching morphogenesis, defined as growth and branching of the ureteric bud (UB), is essential for mammalian kidney development. Previously, we demonstrated that conditional ablation of the PRR in the UB in PRRUB-/- mice causes severe defects in UB branching, resulting in marked kidney hypoplasia at birth. Here, we investigated the UB transcriptome using whole genome-based analysis of gene expression in UB cells, FACS-isolated from PRRUB-/-, and control kidneys at birth (P0) to determine the primary role of the PRR in terminal differentiation and growth of UB-derived collecting ducts. Three genes with expression in UB cells that previously shown to regulate UB branching morphogenesis, including Wnt9b, ß-catenin, and Fgfr2, were upregulated, whereas the expression of Wnt11, Bmp7, Etv4, and Gfrα1 was downregulated. We next demonstrated that infection of immortalized UB cells with shPRR in vitro or deletion of the UB PRR in double-transgenic PRRUB-/-/BatGal+ mice, a reporter strain for ß-catenin transcriptional activity, in vivo increases ß-catenin activity in the UB epithelia. In addition to UB morphogenetic genes, the functional groups of differentially expressed genes within the downregulated gene set included genes involved in molecular transport, metabolic disease, amino acid metabolism, and energy production. Together, these data demonstrate that UB PRR performs essential functions during UB branching and collecting duct morphogenesis via control of a hierarchy of genes that control UB branching and terminal differentiation of the collecting duct cells.

Epigenetics mechanisms in renal development.

Appreciation for the role of epigenetic modifications in the diagnosis and treatment of diseases is fast gaining attention. Treatment of chronic kidney disease stemming from diabetes or hypertension as well as Wilms tumor will all profit from knowledge of the changes in the epigenomic landscapes. To do so, it is essential to characterize the epigenomic modifiers and their modifications under normal physiological conditions. The transcription factor Pax2 was identified as a major epigenetic player in the early specification of the kidney. Notably, the progenitors of all nephrons that reside in the cap mesenchyme display a unique bivalent histone signature (expressing repressive epigenetic marks alongside activation marks) on lineage-specific genes. These cells are deemed poised for differentiation and commitment to the nephrogenic lineage. In response to the appropriate inducing signal, these genes lose their repressive histone marks, which allow for their expression in nascent nephron precursors. Such knowledge of the epigenetic landscape and the resultant cell fate or behavior in the developing kidney will greatly improve the overall success in designing regenerative strategies and tissue reprogramming methodologies from pluripotent cells.

Histone deacetylase 1 and 2 regulate Wnt and p53 pathways in the ureteric bud epithelium.

Histone deacetylases (HDACs) regulate a broad range of biological processes through removal of acetyl groups from histones as well as non-histone proteins. Our previous studies showed that Hdac1 and Hdac2 are bound to promoters of key renal developmental regulators and that HDAC activity is required for embryonic kidney gene expression. However, the existence of many HDAC isoforms in embryonic kidneys raises questions concerning the possible specificity or redundancy of their functions. We report here that targeted deletion of both the Hdac1 and Hdac2 genes from the ureteric bud (UB) cell lineage of mice causes bilateral renal hypodysplasia. One copy of either Hdac1 or Hdac2 is sufficient to sustain normal renal development. In addition to defective cell proliferation and survival, genome-wide transcriptional profiling revealed that the canonical Wnt signaling pathway is specifically impaired in UB(Hdac1,2-/-) kidneys. Our results also demonstrate that loss of Hdac1 and Hdac2 in the UB epithelium leads to marked hyperacetylation of the tumor suppressor protein p53 on lysine 370, 379 and 383; these post-translational modifications are known to boost p53 stability and transcriptional activity. Genetic deletion of p53 partially rescues the development of UB(Hdac1,2-/-) kidneys. Together, these data indicate that Hdac1 and Hdac2 are crucial for kidney development. They perform redundant, yet essential, cell lineage-autonomous functions via p53-dependent and -independent pathways.

Epigenetic States of nephron progenitors and epithelial differentiation.

In mammals, formation of new nephrons ends perinatally due to consumption of mesenchymal progenitor cells. Premature depletion of progenitors due to prematurity or postnatal loss of nephrons due to injury causes chronic kidney disease and hypertension. Intensive efforts are currently invested in designing regenerative strategies to form new nephron progenitors from pluripotent cells, which upon further differentiation provide a potential source of new nephrons. To know if reprogramed renal cells can maintain their identity and fate requires knowledge of the epigenetic states of native nephron progenitors and their progeny. In this article, we summarize current knowledge and gaps in the epigenomic landscape of the developing kidney. We now know that Pax2/PTIP/H3K4 methyltransferase activity provides the initial epigenetic specification signal to the metanephric mesenchyme. During nephrogenesis, the cap mesenchyme housing nephron progenitors is enriched in bivalent chromatin marks; as tubulogenesis proceeds, the tubular epithelium acquires H3K79me2. The latter mark is uniquely induced during epithelial differentiation. Analysis of histone landscapes in clonal metanephric mesenchyme cell lines and in Wilms tumor and normal fetal kidney has revealed that promoters of poised nephrogenesis genes carry bivalent histone signatures in progenitors. Differentiation or stimulation of Wnt signaling promotes resolution of bivalency; this does not occur in Wilms tumor cells consistent with their developmental arrest. The use of small cell number ChIP-Seq should facilitate the characterization of the chromatin landscape of the metanephric mesenchyme and various nephron compartments during nephrogenesis. Only then we will know if stem and somatic cell reprogramming into kidney progenitors recapitulates normal development.

Elevated expression of long intergenic non-coding RNA HOTAIR in a basal-like variant of MCF-7 breast cancer cells.

Epigenetic regulation of gene expression is critical to phenotypic maintenance and transition of human breast cancer cells. HOX antisense intergenic RNA (HOTAIR) is a long intergenic non-coding RNA that epigenetically represses gene expression via recruitment of enhancer of zeste homolog 2 (EZH2), a histone methyltransferase. Elevated expression of HOTAIR promotes progression of breast cancer. In the current study we examined the expression and function of HOTAIR in MCF-7-TNR cells, a derivative of the luminal-like breast cancer cell line MCF-7 that acquired resistance to TNF-α-induced cell death. The expression of HOTAIR, markers of the luminal-like and basal-like subtypes, and growth were compared between MCF-7 and MCF-7-TNR cells. These variables were further assessed upon inhibition of HOTAIR, EZH2, p38 MAPK, and SRC kinase in MCF-7-TNR cells. When compared with MCF-7 cells, MCF-7-TNR cells exhibited an increase in the expression of HOTAIR, which correlated with characteristics of a luminal-like to basal-like transition as evidenced by dysregulated gene expression and accelerated growth. MCF-7-TNR cells exhibited reduced suppressive histone H3 lysine27 trimethylation on the HOTAIR promoter. Inhibition of HOTAIR and EZH2 attenuated the luminal-like to basal-like transition in terms of gene expression and growth in MCF-7-TNR cells. Inhibition of p38 and SRC diminished HOTAIR expression and the basal-like phenotype in MCF-7-TNR cells. HOTAIR was robustly expressed in the native basal-like breast cancer cells and inhibition of HOTAIR reduced the basal-like gene expression and growth. Our findings suggest HOTAIR-mediated regulation of gene expression and growth associated with the basal-like phenotype of breast cancer cells.

Genome-wide analysis of gestational gene-environment interactions in the developing kidney.

The G protein-coupled bradykinin B2 receptor (Bdkrb2) plays an important role in regulation of blood pressure under conditions of excess salt intake. Our previous work has shown that Bdkrb2 also plays a developmental role since Bdkrb2(-/-) embryos, but not their wild-type or heterozygous littermates, are prone to renal dysgenesis in response to gestational high salt intake. Although impaired terminal differentiation and apoptosis are consistent findings in the Bdkrb2(-/-) mutant kidneys, the developmental pathways downstream of gene-environment interactions leading to the renal phenotype remain unknown. Here, we performed genome-wide transcriptional profiling on embryonic kidneys from salt-stressed Bdkrb2(+/+) and Bdkrb2(-/-) embryos. The results reveal significant alterations in key pathways regulating Wnt signaling, apoptosis, embryonic development, and cell-matrix interactions. In silico analysis reveal that nearly 12% of differentially regulated genes harbor one or more Pax2 DNA-binding sites in their promoter region. Further analysis shows that metanephric kidneys of salt-stressed Bdkrb2(-/-) have a significant downregulation of Pax2 gene expression. This was corroborated in Bdkrb2(-/-);Pax2(GFP+/tg) mice, demonstrating that Pax2 transcriptional activity is significantly repressed by gestational salt-Bdkrb2 interactions. We conclude that gestational gene (Bdkrb2) and environment (salt) interactions cooperate to impact gene expression programs in the developing kidney. Suppression of Pax2 likely contributes to the defects in epithelial survival, growth, and differentiation in salt-stressed BdkrB2(-/-) mice.

Mdm2 is required for maintenance of the nephrogenic niche.

The balance between nephron progenitor cell (NPC) renewal, survival and differentiation ultimately determines nephron endowment and thus susceptibile to chronic kidney disease and hypertension. Embryos lacking the p53-E3 ubiquitin ligase, Murine double minute 2 (Mdm2), die secondary to p53-mediated apoptosis and growth arrest, demonstrating the absolute requirement of Mdm2 in embryogenesis. Although Mdm2 is required in the maintenance of hematopoietic stem cells, its role in renewal and differentiation of stem/progenitor cells during kidney organogenesis is not well defined. Here we examine the role of the Mdm2-p53 pathway in NPC renewal and fate in mice. The Six2-GFP::Cre(tg/+) mediated inactivation of Mdm2 in the NPC (NPC(Mdm)2(-/-)) results in perinatal lethality. NPC(Mdm)2(-/-) neonates have hypo-dysplastic kidneys, patchy depletion of the nephrogenic zone and pockets of superficially placed, ectopic, well-differentiated proximal tubules. NPC(Mdm2-/-) metanephroi exhibit thinning of the progenitor GFP(+)/Six2(+) population and a marked reduction or loss of progenitor markers Amphiphysin, Cited1, Sall1 and Pax2. This is accompanied by aberrant accumulation of phospho-γH2AX and p53, and elevated apoptosis together with reduced cell proliferation. E13.5-E15.5 NPC(Mdm2-/-) kidneys show reduced expression of Eya1, Pax2 and Bmp7 while the few surviving nephron precursors maintain expression of Wnt4, Lhx1, Pax2, and Pax8. Lineage fate analysis and section immunofluorescence revealed that NPC(Mdm2-/-) kidneys have severely reduced renal parenchyma embedded in an expanded stroma. Six2-GFP::Cre(tg/+); Mdm2(f/f) mice bred into a p53 null background ensures survival of the GFP-positive, self-renewing progenitor mesenchyme and therefore restores normal renal development and postnatal survival of mice. In conclusion, the Mdm2-p53 pathway is essential to the maintenance of the nephron progenitor niche.

The MDM2-p53 pathway: multiple roles in kidney development.

The molecular basis of nephron progenitor cell renewal and differentiation into nascent epithelial nephrons is an area of intense investigation. Defects in these early stages of nephrogenesis lead to renal hypoplasia, and eventually hypertension and chronic kidney disease. Terminal nephron differentiation, the process by which renal epithelial precursor cells exit the cell cycle and acquire physiological functions is equally important. Failure of terminal epithelial cell differentiation results in renal dysplasia and cystogenesis. Thus, a better understanding of the transcriptional frameworks that regulate early and late renal cell differentiation is of great clinical significance. In this review, we will discuss evidence implicating the MDM2-p53 pathway in cell fate determination during development. The emerging central theme from loss- and gain-of-function studies is that tight regulation of p53 levels and transcriptional activity is absolutely required for nephrogenesis. We will also discuss how post-translational modifications of p53 (e.g., acetylation and phosphorylation) alter the spatiotemporal and functional properties of p53 and thus cell fate during kidney development. Mutations and polymorphisms in the MDM2-p53 pathway are present in more than 50 % of cancers in humans. This raises the question of whether sequence variants in the MDM2-p53 pathway increase the susceptibility to renal dysgenesis, hypertension or chronic kidney disease. With the advent of whole exome sequencing and other high throughput technologies, this hypothesis is testable in cohorts of children with renal dysgenesis.

Genome-wide analysis of the p53 gene regulatory network in the developing mouse kidney.

Despite mounting evidence that p53 senses and responds to physiological cues in vivo, existing knowledge regarding p53 function and target genes is largely derived from studies in cancer or stressed cells. Herein we utilize p53 transcriptome and ChIP-Seq (chromatin immunoprecipitation-high throughput sequencing) analyses to identify p53 regulated pathways in the embryonic kidney, an organ that develops via mesenchymal-epithelial interactions. This integrated approach allowed identification of novel genes that are possible direct p53 targets during kidney development. We find the p53-regulated transcriptome in the embryonic kidney is largely composed of genes regulating developmental, morphogenesis, and metabolic pathways. Surprisingly, genes in cell cycle and apoptosis pathways account for <5% of differentially expressed transcripts. Of 7,893 p53-occupied genomic regions (peaks), the vast majority contain consensus p53 binding sites. Interestingly, 78% of p53 peaks in the developing kidney lie within proximal promoters of annotated genes compared with 7% in a representative cancer cell line; 25% of the differentially expressed p53-bound genes are present in nephron progenitors and nascent nephrons, including key transcriptional regulators, components of Fgf, Wnt, Bmp, and Notch pathways, and ciliogenesis genes. The results indicate widespread p53 binding to the genome in vivo and context-dependent differences in the p53 regulon between cancer, stress, and development. To our knowledge, this is the first comprehensive analysis of the p53 transcriptome and cistrome in a developing mammalian organ, substantiating the role of p53 as a bona fide developmental regulator. We conclude p53 targets transcriptional networks regulating nephrogenesis and cellular metabolism during kidney development.

Histone signature of metanephric mesenchyme cell lines.

The metanephric mesenchyme (MM) gives rise to nephrons, the filtering units of the mature kidney. The MM is composed of uninduced (Six2(high)/Lhx1(low)) and induced (Wnt-stimulated, Six2(low)/Lhx1(high)) cells. The global epigenetic state of MM cells is unknown, partly due to technical difficulty in isolating sufficient numbers of homogenous cell populations. We therefore took advantage of two mouse clonal cell lines representing the uninduced (mK3) and induced (mK4) metanephric mesenchyme (based on gene expression profiles and ability to induce branching of ureteric bud). ChIP-Seq revealed that whereas H3K4me3 active region "peaks" are enriched in metabolic genes, H3K27me3 peaks decorate mesenchyme and epithelial cell fate commitment genes. In uninduced mK3 cells, promoters of "stemness" genes (e.g., Six2, Osr1) are enriched with H3K4me3 peaks; these are lost in induced mK4 cells. ChIP-qPCR confirmed this finding and further demonstrated that G9a/H3K9me2 occupy the promoter region of Six2 in induced cells, consistent with the inactive state of transcription. Conversely, genes that mark the induced epithelialized state (e.g., Lhx1, Pax8), transition from a non-permissive to an active chromatin signature in mK3 vs. mK4 cells, respectively. Importantly, stimulation of Wnt signaling in uninduced mK3 cells provokes an active chromatin state (high H3K4me3, low H3K27me3), recruitment of ß-catenin, and loss of pre-bound histone methyltransferase Ezh2 in silent induced genes followed by activation of transcription. We conclude that the chromatin signature of uninduced and induced cells correlates strongly with their gene expression states, suggesting a role of chromatin-based mechanisms in MM cell fate.

Histone deacetylases in kidney development: implications for disease and therapy.

Histone deacetylases (HDACs) are an evolutionarily conserved group of enzymes that regulate a broad range of biological processes through removal of acetyl groups from histones as well as non-histone proteins. Recent studies using a variety of pharmacological inhibitors and genetic models of HDACs have revealed a central role of HDACs in control of kidney development. These findings provide new insights into the epigenetic mechanisms underlying congenital anomalies of the kidney and urinary tract (CAKUT) and implicate the potential of HDACs as therapeutic targets in kidney diseases, such as cystic kidney diseases and renal cell cancers. Determining the specific functions of individual HDAC members would be an important task of future research.

Interactions between BdkrB2 and p53 genes in the developing kidney.

Gene-environment interactions are implicated in congenital disorders. Accordingly, there is a pressing need to develop animal models of human disease, which are the product of defined gene-environment interactions. Work from our laboratory demonstrates the presence of genetic interactions between the bradykinin B2 receptor (BdkrB2) and the tumor suppressor protein p53 in the developing kidney. Our studies have shown that the Bdkrb2(-/-) embryos exposed to gestational salt stress develop renal dysgenesis. The underlying mechanism is p53 stabilization and mediated apoptosis and repression of the terminal epithelial differentiation program. We also uncovered a novel functional cross-talk between p53 and BdkrB2. Thus, while BdkrB2 is a target for p53-mediated transcriptional activation, BdkrB2 inactivation results in the upregulation of checkpoint kinase 1 (Chk1) levels, thus potentiating phosphorylation of p53 on Ser23 by Chk1, an essential step in the pathway leading to renal dysgenesis in salt-stressed BdkrB2(-/-) mutant mice. Future studies will now focus on defining how this G-protein-coupled receptor is coupled to the activation of p53, a tumor suppressor gene that is mutated in more than 50% of all human cancers.

A p53-Pax2 pathway in kidney development: implications for nephrogenesis.

Congenital reduction in nephron number (renal hypoplasia) is a predisposing factor for chronic kidney disease and hypertension. Despite identification of specific genes and pathways in nephrogenesis, determinants of final nephron endowment are poorly understood. Here, we report that mice with germ-line p53 deletion (p53(-/-)) manifest renal hypoplasia; the phenotype can be recapitulated by conditional deletion of p53 from renal progenitors in the cap mesenchyme (CM(p53-/-)). Mice or humans with germ-line heterozygous mutations in Pax2 exhibit renal hypoplasia. Since both transcription factors are developmentally expressed in the metanephros, we tested the hypothesis that p53 and Pax2 cooperate in nephrogenesis. In this study, we provide evidence for the presence of genetic epistasis between p53 and Pax2: a) p53(-/-) and CM(p53-/-)embryos express lower Pax2 mRNA and protein in nephron progenitors than their wild-type littermates; b) ChIP-Seq identified peaks of p53 occupancy in chromatin regions of the Pax2 promoter and gene in embryonic kidneys; c) p53 binding to Pax2 gene is significantly more enriched in Pax2 -expressing than non-expressing metanephric mesenchyme cells; d) in transient transfection assays, Pax2 promoter activity is stimulated by wild-type p53 and inhibited by a dominant negative mutant p53; e) p53 knockdown in cultured metanephric mesenchyme cells down-regulates endogenous Pax2 expression; f) reduction of p53 gene dosage worsens the renal hypoplasia in Pax2(+/-) mice. Bioinformatics identified a set of developmental renal genes likely to be co-regulated by p53 and Pax2. We propose that the cross-talk between p53 and Pax2 provides a transcriptional platform that promotes nephrogenesis, thus contributing to nephron endowment.