Periodic Reporting for period 4 - Reg-Seq (Functional genomics of non-coding mutations in regulatory regions of four metabolic tissues, and their involvement into type 2 diabetes through large-scale sequencing)
Période du rapport: 2021-11-01 au 2022-04-30
Type 2 diabetes (T2D) is a complex systemic metabolic disorder which has developed into a global major health problem responsible for early morbidities and mortality. T2D is a multifactorial disease, resulting from the progressive alteration of insulin secretion from beta cells in pancreatic islets, on a background of resistance to insulin action in various organs and tissues, including liver, skeletal muscle and adipose tissue.
T2D heritability is estimated around 70%. Genome-wide association studies identified more than 250 T2D-associated common variants, but altogether they explain less than 15% of T2D inheritance.
Rare loss-of-function coding variants may contribute to T2D risk.
Moreover, recent studies suggested the putative impact on T2D risk of functional non-coding variants within ac-tive enhancers of pancreatic islets; but tissue-specific regulatory regions have remained largely unexplored by genetic epidemiology studies.
I intend to make progress in T2D genetic epidemiology and physiology by extending genetic investigation to frequent and rare mutations in coding and non-coding regulatory regions on a whole-genome basis. In the Reg-Seq project, I propose the next-generation sequencing (NGS) of most relevant DNA regulatory sequences in large well-phenotyped prospective populations, followed by a comprehensive functional experiments when genetic associations are identified.
This project should lead to the identification of new functional genetic markers and pathways involved in T2D risk and development, and should help to stratify the T2D population for precision medicine, which is one of the major goals of the EU-Horizon 2020 programme. Further elucidation of the missing heritability should enable a major advance in the prediction of T2D risk, which is still very poor for a genuine clinical translation.
T2D heritability is estimated around 70%. Genome-wide association studies identified more than 250 T2D-associated common variants, but altogether they explain less than 15% of T2D inheritance.
Rare loss-of-function coding variants may contribute to T2D risk.
Moreover, recent studies suggested the putative impact on T2D risk of functional non-coding variants within ac-tive enhancers of pancreatic islets; but tissue-specific regulatory regions have remained largely unexplored by genetic epidemiology studies.
I intend to make progress in T2D genetic epidemiology and physiology by extending genetic investigation to frequent and rare mutations in coding and non-coding regulatory regions on a whole-genome basis. In the Reg-Seq project, I propose the next-generation sequencing (NGS) of most relevant DNA regulatory sequences in large well-phenotyped prospective populations, followed by a comprehensive functional experiments when genetic associations are identified.
This project should lead to the identification of new functional genetic markers and pathways involved in T2D risk and development, and should help to stratify the T2D population for precision medicine, which is one of the major goals of the EU-Horizon 2020 programme. Further elucidation of the missing heritability should enable a major advance in the prediction of T2D risk, which is still very poor for a genuine clinical translation.
During the period covered by this report, I have optimized several technologies/experiments required for achieving the different milestones of my Reg-Seq ERC proposal:
1/ I have implemented the workflow for targeted next-generation sequencing of specific genomic regions in 10,000 DNA samples. The workflow includes the preparation of DNA samples (quantity/concentration), the preparation of libraries (Roche technologies) through different automates, the sequencing of libraries (Illumina next-generation sequencing), the computer analyses (alignment of reads and detection/annotation of variants), the curation of data and the statistical analyses.
2/ I have developed a new technology enabling the analysis of spatial interactions between a specific genomic locus of interest and the rest of the genome.
3/ I have implemented chromatin immunoprecipitation (ChIP) sequencing (ChIP-seq) targeting specific histone marks in specific cell lines, along with the accurate computer analyses.
4/ In the same context, I have implemented genome editing (CRISPR/Cas9) in in specific cell lines.
1/ I have implemented the workflow for targeted next-generation sequencing of specific genomic regions in 10,000 DNA samples. The workflow includes the preparation of DNA samples (quantity/concentration), the preparation of libraries (Roche technologies) through different automates, the sequencing of libraries (Illumina next-generation sequencing), the computer analyses (alignment of reads and detection/annotation of variants), the curation of data and the statistical analyses.
2/ I have developed a new technology enabling the analysis of spatial interactions between a specific genomic locus of interest and the rest of the genome.
3/ I have implemented chromatin immunoprecipitation (ChIP) sequencing (ChIP-seq) targeting specific histone marks in specific cell lines, along with the accurate computer analyses.
4/ In the same context, I have implemented genome editing (CRISPR/Cas9) in in specific cell lines.
In addition to the different milestones related to the development of new technologies, my main goal is to find key regulatory regions involved in beta cell function, through the identification of rare functional DNA variants enriched in patients with type 2 diabetes.