Decoding pleiotropic causal genetic risk variants of periodontitis using massively parallel reporter assays

• Applicants: Professor Dr. Martin Kircher; Professor Dr. Arne Schäfer
• Subject Area: Dentistry, Oral Surgery ; General Genetics and Functional Genome Biology
• Term: since 2023
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 517993632

Severe periodontitis is a progressive inflammatory disease of the oral cavity and represents the sixth-most prevalent disease condition in the world. The observed phenotypic variation is largely attributed to genetic variability between different individuals, and the specific biological consequences of genetic variants form the genetic architecture that influences and shapes disease manifestations. We carried out a series of genome-wide association studies and identified numerous suggestive single nucleotide polymorphisms (SNPs) associated with periodontitis. Some are in strong linkage disequilibrium with genome-wide significant associations (p < 5 x 10-8) with other common complex diseases and traits, indicating pleiotropic effects of these loci. Pleiotropic loci have strong implications for identification of molecular targets for drug development, future genetic risk-profiling, and classification of diseases. We developed plasmid- and lentivirus-based massively parallel reporter assays (MPRAs) that enable us to test the associated alleles at tens of thousands of candidate regulatory sequences (CRS) for their regulatory function in parallel. Here, we propose to use MPRAs to identify the putative causal variants (PCVs) of nine pleiotropic periodontitis associations that are specifically shared with other diseases and traits such as coronary heart disease, bone mineral density, plasminogen concentration and activity, as well as gut microbiome composition. Subsequently, the identified functional pleiotropic CRS will be validated using luciferase reporter genes. We will computationally predict underlying transcription factor binding affinities at the functional PCVs, and we will biologically validate their allele-specific binding affinities using antibody electrophoretic mobility shift assays. Furthermore, we will determine the target genes of the functional PCVs by CRISPR-dCas9 gene activation/inhibition or chromatin conformation capture methods. If the molecular genetic causes of complex diseases are not understood, we believe that there cannot be long-term gain in translational medicine. Prerequisite for the discovery of disease relevant signaling cascades and of their interactions between different diseases is the identification and characterization of the hitherto unknown causal variants that underlie these associations, and particularly of those that are shared with different diseases. These signaling pathways together with the corresponding regulatory chromatin elements also provide suitable anchor points for future translational medicine. Therefore, we consider this project, which is currently unique in dental research, to have significance beyond oral medicine.

• DFG Programme: Research Grants