Study of Arc protein
The aim of this research, starting in 2021, is the structural and functional study of Arc capsid formation. Arc is an early gene expressed upon neuronal activity; its differential expression was shown in several disorders affecting the CNS, such as Alzheimer’s disease, autism spectrum disorder, and schizophrenia. Arc is present in cells in three structural variants: open, closed and capsid forms. The capsid form of Arc is able to bind and transfer mRNA from one cell to another. At the same time, the functional role of different structural variants is not clear. In our study, collaborating with colleagues from the Department of Organic Chemistry and the Department of Biochemistry, we aim to understand the capsid formation and RNA transfer of Arc. In our study, we intend to perform in vivo experiments – on the animal models of dementia and epilepsy – and in vitro experiments as well.
Single-cell mRNA sequencing, neuro-immune interactions
In our single-cell mRNA sequencing work, we investigated the transcriptomic differences between prefrontal pyramidal cells and interneurons, which raise the possibility of selective pharmacological targeting of the two cell types (Ravasz et al., 2021). Accordingly, a number of mRNAs encoding cell surface receptor or ion channel subunits have been identified with at least ten-fold copy number differences between the two cell types. The single-cell sequencing data set, which contains more than 19,000 transcripts, also provided an opportunity to study the neuronal expression of genes related to neuro-immune interactions using bioinformatics tools (in silico analysis). To reveal the functional relevance of neuro-immune interactions, we examine the electrophysiological effects of major cytokines in acute brain slices by patch clamp measurements (ex vivo electrophysiology) and the changes of cortical electrical activity (EEG, in vivo electrophysiology) using LPS-treated mice, a model of acute bacterial infections. The related protein-level changes of the prefrontal cortex are monitored by various molecular methods (Western blot and other immunoassays, proteomic analysis of synaptosome preparations). We conducted our research in collaboration with the associates of University of Pennsylvania.
Ravasz L et al., (2021) Cell Surface Protein mRNAs Show Differential Transcription in Pyramidal and Fast-Spiking Cells as Revealed by Single-Cell Sequencing. Cereb Cortex. 2021 31(2):731-745.
We acquired 2-dimensional differential gel electrophoresis (2D DIGE) method in 2006 in our laboratory; since then, solely our laboratory conducts proteomic experiments with this method on human and animal samples. During this time, we published 21 publications only on this field of science.
In the last few years, we conducted studies from diverse angles mostly on dementia-related diseases (Alzheimer’s disease (AD), vascular dementia).
Chronic cerebral hypoperfusion is the animal model of vascular dementia. We perform proteomic experiments on brain areas that play role in a number of symptoms. Within those brain areas, we analyze the proteome of several cell organelles (synaptosomes, mitochondria, and mitochondria-associated membrane (MAM)) (Tukacs et al., 2020).
We studied and study the most known animal model of AD, the APP/PS1 double transgenic mice from several aspects. The overexpression of amyloid precursor protein (APP) and the accumulation of β-amyloid (Aβ) and tau protein tangles are the hallmarks of AD. In relation to the progression of Aβ-associated processes, we carried out a comprehensive study on the synaptic and non-synaptic mitochondrial proteome changes of 3, 6, and 9 month old APP/PS1 mice. In another study, we compared the MAM proteome of APP/PS1 and C57BL/6 control mice; since the current hypothesis of AD pathology suggests that the MAM has an important role in the early phase of AD.
It is known that synapse loss is the concomitant phenomenon of numerous neurodegenerative disorders, which results in declining cognitive functions. Further, it was shown that one of the members of the complement system, C1q protein, plays an important role in synaptic plasticity; it tags synapsis for phagocytosis by microglia cells. In our experiments, we aimed to better understand the elimination of synaptic interactions by the proteomic study of C1q-tagged and non-tagged synapses (Györffy et al., 2018). Also, we revealed the role of neuropentraxins as the synaptic interaction partners of C1q (Kovács et al., 2021).
We conducted our research in close collaboration with the associates of the Department of Biochemistry and the Proteomics laboratory of ELKH SZBK who performed the MS identification of proteins.
Kovács RÁ et al. (2021) Identification of Neuronal Pentraxins as Synaptic Binding Partners of C1q and the Involvement of NP1 in Synaptic Pruning in Adult Mice. J. Front Immunol. 11:599771.
Tukacs V. et al. (2020) Chronic stepwise cerebral hypoperfusion differentially induces synaptic proteome changes in the frontal cortex, occipital cortex, and hippocampus in rats. Sci Rep.10(1):15999.
Völgyi K. et al. (2018) Early Presymptomatic Changes in the Proteome of Mitochondria-Associated Membrane in the APP/PS1 Mouse Model of Alzheimer's Disease. Mol Neurobiol.;55(10):7839-7857.
Völgyi K. et al. (2017) Mitochondrial Proteome Changes Correlating with β-Amyloid Accumulation. Mol Neurobiol. 54(3):2060-2078.
In our laboratory, besides the above mentioned ex vivo patch clamp method, we perform in vivo electrophysiological experiments on acute and free-moving animals. In many cases, we validated our result from proteomics and single-cell sequencing studies with electrophysiological recordings. An example is presented in the Single-cell sequencing paragraph; where we treat rats and mice with the endotoxin from the cell wall of Gram-negative bacteria, lipopolysaccharide (LPS), and study the effect of immune response in the central nervous system with several electrophysiological techniques (EEG, field potential, pop-spike).
In many cases, we combine electrophysiological recordings with cerebral microdialysis.