Project Details
Projekt
Integrative craniofacial genomics - From GWAS to function
Applicant
Professorin Dr. Kerstin Ludwig
Subject Area
Human Genetics
Developmental Biology
Developmental Biology
Term
from 2016 to 2023
Project identifier
Deutsche Forschungsgemeinschaft (DFG) - Project number 289822512
Nonsyndromic cleft lip with or without cleft palate (nsCL/P) is a common human birth defect resulting from molecular disturbances during craniofacial development. Recent breakthroughs in the identification of genetic risk factors for nsCL/P have been achieved, mainly via genome-wide association studies. Sixteen risk loci are established to date, and the majority of these map to non-coding regions outside known genes. Although this suggests that the identified variants have a regulatory effect, understanding of the underlying biological mechanisms remains limited, mainly due to little access to relevant biological material.The main aim of this Emmy-Noether application is to advance understanding of embryonic craniofacial development using nsCL/P as a model trait. The project will (i) identify novel susceptibility regions via the integration of genome-wide genetic- and functional datasets, and (ii) characterize known and newly identified risk loci using genetics, bioinformatics and functional approaches. Functional data will include the analysis of different histone modification marks, transcription factor binding profiles, and expression levels during human and murine embryonic development. For this purpose, valuable datasets are currently being generated by the proposed collaboration partners and will be complemented by data from international consortia (e.g., FaceBase). This approach will lead to the discovery of putative causal regulatory elements and candidate genes. The identified regions will be re-sequenced in large cohorts of 1,500 nsCL/P patients and 1,500 controls using a novel targeted next-generation sequencing protocol, i.e., molecular inversion probes. Establishing this technology in the Emmy-Noether lab will allow time- and cost-effective sequencing of large cohorts, and can be extended to the analysis of both somatic variation and other phenotypes. Subsequently, identified variants will be confirmed by Sanger sequencing, and co-segregation or de novo status will be assessed. At a cohort-level, burden analyses will provide insights into the general overrepresentation of rare variants in patients, within functionally relevant regions. For confirmed variants, functional analyses will be conducted in vivo using the zebrafish as a model organism. Candidate regulatory elements will be inserted into one-stage embryos using Tol2-mediated transposition, and fluorescence patterns will be monitored up to five days post-fertilization. In addition, candidate genes will be knocked-down using either morpholinos or Crispr/Cas9- technology.The results of this Emmy-Noether group will provide novel insights into the molecular pathways underlying craniofacial development. More generally, they will facilitate understanding of the regulatory architecture of non-coding regions, and their role in embryonic development and disease.
DFG Programme
Independent Junior Research Groups
International Connection
Netherlands, United Kingdom, USA
