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. A better understanding of the mechanisms by which gene-environment interactions induce congenital defects is necessary to advance preventive and therapeutic strategies for organ dysgenesis. We previously demonstrated that bradykinin B2 receptor-null (BdkrB2-/-) embryos exposed to gestational salt stress develop renal dysgenesis as a result of p53-mediated apoptosis of nephron progenitors and repression of the terminal differentiation program. A physiological cross-talk operates between the tumor suppressor protein, p53, and the BdkrB2 during renal organogenesis. Thus, while BdkrB2 is a target for p53-mediated transcriptional activation, BdkrB2 is required to prevent the phosphorylation of p53 on Ser23 by Checkpoint kinase 1 (Chk1). p-p53Ser23 is the central mediator of renal dysgenesis in BdkrB2-/- mutant mice. The Overall hypothesis to be tested in this Competitive Renewal application is that activation of the Chk1-p53 pathway, which mediates the susceptibility of BdkrB2 mutants to renal dysgenesis, is cell autonomous, and that embryonic salt stress acts epigenetically to repress the terminal differentiation program in a genetically susceptible host.
Specific Aim 1 tests the hypothesis that BdkrB2 receptors mediate nephroprotection in the ureteric bud lineage. To this end, we have developed conditional BdkrB2lox/lox mice, which will be used to produce progeny lacking the receptor from the ureteric bud or metanephric mesenchyme by Cre-mediated excision. We anticipate that loss of BdkrB2 from the collecting duct is sufficient to recapitulate the conventional BdkrB2-null phenotype by activating the Chk1- p53 pathway.
Specific Aim 2 tests the hypothesis that exposure of BdkrB2-null embryos to salt stress is associated with progressive epigenetic silencing of Pax-2, a transcription factor that is required for normal epithelial nephron differentiation. Genetic crosses to reporter mice will reveal the spatiotemporal dynamics of Pax2 gene transcription in Bdkrb2-mutants. Characterization of the histone code at the Pax2 promoter will determine that progressive transcriptional repression in BdkrB2-/- kidneys is accompanied by histone deacetylase-1 recruitment, promoter histone H3 deacetylation and lysine 4 (H3K4) demethylation and H3K9/27 methylation, followed by hypermethylation of CpG islands. We postulate that these modifications in the histone code inhibit transcription factor binding to the Pax2 promoter. Finally, we will determine if these epigenetic effects on gene expression can be rescued by conditional inactivation of HDAC1.
Specific Aim 3 tests the hypothesis that p-p53Ser23, which accumulates in dysplastic BdkrB2-/- kidneys, represses Hepatocyte Nuclear Factor-1 (HNF1?) gene transcription. These studies will demonstrate that repression of HNF1? contributes to epithelial-mesenchymal transition and thus to dysplasia. We propose that replacement of the endogenous p53 gene with a non-phosphorytable version of p53 (p53S/A23) rescues HNF1? expression and terminal differentiation in salt-stressed BdkrB2-/- embryos.
Congenital malformations of the kidney and urinary tract account for up to 40% of chronic kidney failure in children less than 4 years of age. In addition to mutations in the DNA sequence, abnormal kidney development may result from alterations in the epigenetic code. Stable alterations in the epigenetic code resulting from transient environmental exposures can permanently disrupt the fetal gene expression program. Thus, understanding how a defined genetic mutation (e.g., BdkrB2-/-) interacts with a defined embryonic stressor (gestational salt) to alter the regulation of kidney development may open new avenues to the treatment or prevention of kidney and urinary tract malformations.
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|Yan, Lei; Yao, Xiao; Bachvarov, Dimcho et al. (2014) Genome-wide analysis of gestational gene-environment interactions in the developing kidney. Physiol Genomics 46:655-70|
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|Chen, Shaowei; Bellew, Christine; Yao, Xiao et al. (2011) Histone deacetylase (HDAC) activity is critical for embryonic kidney gene expression, growth, and differentiation. J Biol Chem 286:32775-89|
|Hilliard, Sylvia; Aboudehen, Karam; Yao, Xiao et al. (2011) Tight regulation of p53 activity by Mdm2 is required for ureteric bud growth and branching. Dev Biol 353:354-66|
|Song, Renfang; Spera, Melissa; Garrett, Colleen et al. (2010) Angiotensin II AT2 receptor regulates ureteric bud morphogenesis. Am J Physiol Renal Physiol 298:F807-17|
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