Our previous studies demonstrated that the regulation of the DNA methylation pattern involves the poly(ADP-ribosyl)ation (PARylation) process, a post-translational modification of proteins catalysed by the poly(ADP-ribose) polymerase (PARP) enzymes. DNA methylation can influence the genetic susceptibility to type 2 diabetes mellitus (T2DM) and the progression of the disease. We have recently shown that the PARylation process is over-activated in patients with T2DM, particularly in those with poor glycaemic control. In PBMC from diabetic patients, PARylation was found to be linked to dysregulation of DNA methylation pattern via activation of the DNA de-methylation cascade.
In the present project, we aim to test if the observed changes in DNA methylation pattern are effectively induced by PARP-1 over-activation in T2DM and if this interplay may be driven by a common element: prolonged increases in reactive oxygen species (ROS) production, key factors in the pathogenesis of T2DM and obesity-related diseases. For this reason, we will use cultured cells exposed to high glucose as a valuable tool for defining the role of PARylation in the DNA methylation process by modulating its activity through specific inhibitors and silencing of PARP enzymes. As ROS production has been reported to stimulate PARP activity in response to high glucose, we aim to understand if enhanced intracellular glucose transport can lead to ROS overproduction and subsequent PARP activation by monitoring the production of ROS in our cellar system.
Further step will be to measure the oxidative status of both PBMC and plasma in subjects affected by T2DM and see how it correlates with PARylation/ DNA demethylation interplay.
The understanding of the DNA methylation/de-methylation mechanism is important, since restoring the control of DNA de-methylation cycle promises beneficial effects. In this context, the use of PARP inhibitors may be therapeutically relevant.
HIGHLIGHTS
Relevance
This project aims to collect initial evidence that may form an understanding for the mechanistic basis of the emerging connection between defective glucose metabolism and epigenetic modifications of chromatin in diabetes. We highlight the ability of the PARylation process to alter the epigenetic profile via DNA de-methylation in a HG milieu, thereby exerting epigenetic control functions in T2DM.
The hyperglycaemia-driven over-production of ROS and oxidative stress are the major biochemical abnormalities underlying diabetes pathogenesis and its complications and have been proposed as the primary mediators of 'glucotoxicity'. More recently, it was found that glucotoxicity generates permanent changes of the expression of genes associated with the control of glucose homeostasis through epigenetic mechanisms. This finding offers an elegant explanation of the phenomenon of "metabolic memory", which accounts for the correlation between transient/chronic poor glycaemic control and the irreversible progression of diabetic complications. In this context, we propose the ROS-mediated activation of PARP1 exerting epigenetic control on diabetes-related gene expression.
Far-reaching, relevant clinical implications of this research include the use of PARs as indicators of peripheral DNA damage and cellular stress. PAR might represent new biomarkers for the follow-up monitoring of the diabetic patient. In addition, PARP inhibitors might be exploited as new therapeutic drugs in the management of diabetes by delaying some complications, thereby improving the patient outcome.
Innovativeness
PARylation may be considered a key process able to transduce the insults due to poor glycaemic control into variations of the epigenetic code, paving the way to i) the exploitation of PAR detection as marker of chronic inflammation associated to T2DM, ii) the use of PARP inhibitors in diabetes management.
Originality
PARylation could constitute a link between oxidative stress, typical of a condition of poor glycaemic control, and epigenetic deregulation.
Feasibility
The project will combine expertise in PARylation and DNA methylation with expertise in clinical medicine of diabetes. The proponents have the required background knowledge and possess the basic equipment to address the experimental aims of the project.
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