Autosomal dominant polycystic kidney disease (ADPKD) is caused by mutations in one of two genes, PKD1 or PKD2, whereas it cannot be fully understood in terms of the constrained genetic setting, especially, in families with the same genetic mutations but variable disease severity. Epigenetic regulation as a critical driver of cell fate and survival can occur even in genetically identical humans, which may be an alternative means of explaining PKD-associated alterations. Thus, the roles of epigenetic modulation of gene expression and protein functions in ADPKD should become the focus of scientific investigation. However, in addition to histone deacetylases (HDACs), the roles of DNA and histone methylation and the enzymes that mediate these processes in ADPKD remain largely unexplored. PKD1 is hypermethylated in gene-body regions and its expression is downregulated in ADPKD. By performing whole-genome bisulfite sequencing (WGBS) analysis, we have identified the genome-wide abnormal DNA methylation signatures in ADPKD kidneys compared to those in normal kidneys, suggesting that DNA methylation is one of the key mechanisms underlying cystogenesis. Within the five DNA methyltransferases, DNMT1 is the only enzyme that functions to maintain the DNA methylation patterns in human genome. DNMT1 was upregulated in Pkd1 mutant renal epithelial cells and tissues, implying its role in the maintenance of the abnormal DNA methylation signatures in ADPKD genome. We will investigate the roles and mechanisms of DNMT1 in regulating renal cyst progression in aim 1. Since we identified an interaction between DNMT1 and Smyd2, one of the SET- domain-containing histone (lysine) methyltransferases, it suggested that Smyd2 may be involved in DNMT1 mediated DNA methylation. We will investigate the crosstalk of DNMT1 and Smyd2 in the regulation of DNA methylation and further delineate Smyd2-mediated molecular mechanisms in the regulation of cystogenesis in aim 2, which may address if Smyd2 serves as a recruitment platform for DNMT1 on specific gene methylation, thus highlighting a previously unrecognized direct connection between two key epigenetic repression systems and providing a possible explanation of why the upregulation of DNMT1 in cancer and PKD only targets specific genes but not all genes in patients? genome. Furthermore, we will test if de- methylation of hypermethylated DNA mediated by DNMT1 with Hydralazine and Smyd2 inhibitor delays cyst growth in vivo in aim 3. This is the first study that not only links DNMT1 and DNA methylation to ADPKD but also links the corresponding DNMT1 and Smyd2 signaling together in regulation of DNA methylation and gene expression. In addition, this study will produce information that will be therapeutically relevant with excelling potential for translation.
In this study, we will investigate the functional roles and mechanisms of one of the DNA methyltransferase, DNMT1, in regulating cyst progression and address if a histone methyltransferases, SMYD2, serves as a recruitment platform for DNMT1 on specific gene methylation, thus highlighting a previously unrecognized direct connection between two key epigenetic repression systems. We will then test whether de-methylation of hypermethylated DNA mediated by DNMT1 and/or together SMYD2 with their inhibitors delays cyst growth in vivo. Accomplishing this project will lead to a better understanding of the mechanism of renal cyst formation and will provide novel therapeutic targets for ADPKD treatment.
