Repository of DUTH

Μεταβολές στην έκφραση αναδυόμενων υποψήφιων γονιδίων πριν και μετά την εμφάνιση τικ σε παιδιά με το Σύνδρομο Gilles de la Tourette (Master thesis)

Κωνσταντίνου, Γεώργιος/ Konstantinou, Georgios

Tourette Syndrome (TS) is a neurodevelopmental, multifactorial disorder characterized by chronic vocal and motor tics which typically manifest during childhood or early adolescence. The origin of its clinical phenotype is not yet clearly understood. Several scientists have attempted to address its pathophysiology by focusing either on the genetic basis of the disorder or on environmental responses that trigger pathways associated with the clinical presentation of the syndrome. A significant aid in this effort has been the Genome-Wide Association Studies (GWAS), through which genomic variations and chromosomal loci are identified that may be associated with TS etiology and predisposition. The aim of the thesis was to perform a differential gene expression analysis (from peripheral whole blood total RNA) of the genes DALRD3, WDR6, NCKIPSD, CELSR3, and PCDH9. According to the most recent (unpublished) GWAS findings, carried out by Prof. Paschou’s group and colleagues from the USA and Europe, the aforementioned genes have emerged as interesting top-hits potentially implicated in TS, either through gene-based analysis or via functional annotation. The present study was conducted using total RNA samples previously available in the lab, through the EMTICS consortium biobank maintained at DUTh, which was built between 2013 and 2018. Samples were analyzed using either one-step or two-step quantitative real-time polymerase chain reaction (RT-qPCR). The study groups include pediatric individuals recruited through the EMTICS consortium during 2013-2017, who were sampled for DNA and RNA analysis at baseline (no tics, visit 01 or V1) and upon the manifestation of tics (tic onset, visit Ex), as well as an age-matched control group of healthy individuals. Subsequently, expression levels were compared with genotypes of tophit SNVs corresponding to the chromosomal regions of these genes, which were highlighted in the latest GWAS. Genotyping analysis of particular SNPs, which emerged as top hits from the most recent (unpublished) GWAS analysis, was performed either by extracting genotyping data from the Illumina GWAS platform or by Sanger sequencing. Relative quantification of DALRD3 expression using two-step RT-qPCR protocol revealed significantly elevated expression at baseline (V1), followed by a notable reduction at tic onset (Ex), with statistical validation through Wilcoxon test (p = 0.015) and t-test (p = 0.0095). Differential gene expression analysis on whole-transcriptome level (microarray data analysis) confirmed upregulation of DALRD3 alongside WDR6, NCKIPSD, and CELSR3 at V1 and Ex compared to controls, but failed to identify significant intra-group differences. Expression analysis of PCDH9 showed no statistically significant differences between groups across all applied methods. Correlation analysis between genotype and expression failed to produce conclusive results due to sample size limitations and technical constraints in paired genotyping and transcript quantification.Despite the limitations of this study, consistently elevated DALRD3 expression in TS cases compared to controls emerged as an interesting finding that warrants further investigation in a larger cohort, considering how little information is available regarding the molecular functions of DALRD3 in human tissues.