Proteomic analysis of PACS2 syndrome in a murine model: insights into molecular pathways and calcium regulation

PACS2 Syndrome, resulting from E209K or E210K mutations in the PACS2 protein, is an ultra-rare
disease classified among developmental and epileptic encephalopathies (DEE). Despite patient
heterogeneity, common characteristics include early onset seizures, neurodevelopmental symptoms,
behavioral impairment, and facial dysmorphism. Research on this syndrome is limited due to the
extremely low number of patients and the absence of standardized experimental models. To date,
only three research groups have published data on the impact of the mutated protein on cellular
function, all utilizing in vitro models and selected cell lines. To better understand the disease
pathogenesis, more complex models bridging basic science and clinical studies are required.

Therefore, in this study we aim to insight into total proteome of brain and liver in mice model of
PACS2 Syndrome.

PACS2 Syndrome mouse model was obtain in collaboration with Czech Center of Phenogenomics.
Adult B6 mice were breed in Center of Experimental Medicine (Bialystok, Poland). Following
genotyping confirmation, six wild-type and six E209K heterozygote mice were included in the
study. Mice were sacrificed and chosen brains regions and livers were collected. Due to proteome
analysis, cerebellum, frontal cortex, hippocampus, other brain tissue and liver were homogenate. We
performed proteomic analyzes using the Data Independent Analysis (DIA) mode of data collection.
Tryptically aged proteins were separated on a nano-LC system coupled to Orbitrap Fusion MS,
followed by data analysis using Spectronaut software. The analyzes allowed the identification of
approximately 6,000 proteins differentiating the brain tissue of mice with the PACS2 mutation and
the control group.

PACS2 protein is multifunctional sorting protein strongly associate with mitochondrial function. In
our study we detected different up and down regulation of molecular pathways dependent of
studied tissue. Moreover, we observed changes in expression of proteins associate with calcium
management in cells. This, result may put some light on changes in neurotransmission and
neurodevelopment present in PACS2 Syndrome. Elevated calcium levels in cytoplasm may lead to
glutamate release from neurons and epilepsy. However, although our research provides new data on
the molecular mechanisms of the disease, further analysis is required.