Rola białka Mbd3 w procesie epileptogenezy : praca doktorska

Nizińska-Smolińska, Karolina

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Rola białka Mbd3 w procesie epileptogenezy : praca doktorska

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Title: Rola białka Mbd3 w procesie epileptogenezy : praca doktorska

Abstract:

MBD3 (methyl CpG binding domain 3) protein belongs to the MBD family of proteins, responsible for reading the DNA methylation pattern. MBD family proteins bind the methyl-CpG domain and are also involved in heterochromatin formation. Interestingly, MBD3 protein does not have the ability to selectively recognize methyl-CpG islands, however, its characteristic feature is the ability to bind to 5-hydroxymethylcytosine and unmethylated DNA. A study by Bednarczyk and colleagues (Bednarczyk et al. 2016) using a rat model of temporal epilepsy induced by electrical stimulation of the amygdala showed an increase in NuRD complex proteins, including Mbd3 protein, in the brain of epileptic animals. A greater number of regions of DNA to which the NuRD complex, containing the Mbd3 protein, attaches was also observed in stimulated animals, compared to a group of control animals. The aim of the experiments conducted in this dissertation was to investigate whether the Mbd3 protein participates in the processes leading to changes in the seizure threshold. The effects of pentylenetetrazole (PTZ)-induced convulsions on Mbd3 protein levels and Mbd3 mRNA expression in vivo were investigated. An increase in Mbd3 protein levels was demonstrated in the entorhinal cortex and amygdala of rats 4 hours after the induced convulsion.The effects of decreasing and increasing Mbd3 protein levels on seizure threshold in vivo were further evaluated. Commercially designed AAV viral vectors were used to modify Mbd3 levels. It was shown that lowering the level of Mbd3 prolongs the latency time to the onset of a convulsion induced by PTZ injection.Behavioral experiments have shown that downregulation of MBD3 protein increases anxiety in animals. In contrast, overexpression of Mbd3 decreases anxiety responses in animals and increases their excitability and activity in the open field test. The effects of decreasing and increasing Mbd3 protein levels on epileptogenesis were also investigated in a kindling model using PTZ. Elevation of Mbd3 protein levels was shown to accelerate epileptogenesis and the development of tonic-clonic seizures. In order to identify the role of Mbd3 protein in the regulation of gene expression, in vitro experiments were conducted using a model of magnesium deficiency-induced epileptic-like discharges. Overexpression of Mbd3 in vitro was shown to induce changes in gene expression in a time- and state-specific manner in neurons.The data obtained from this project indicate the involvement of the Mbd3 protein in epilepsy pathology.