Periodic Reporting for period 1 - NOJUNKDNA (A strategy to investigate cell type-specific effects of non-coding regulatory elements to be applied in neurodegeneration research)
Reporting period: 2018-05-01 to 2020-04-30
In the last 20 years, many risk factors have been identified that link genetic variants to disease susceptibility. While more than a few hundred of these commonly called “risk genes” are known, efforts to translate these insights into improved diagnostics or therapy have lead virtually nowhere. One of the reasons is that the vast majority of genetic variants linked to complex neurodegenerative disorders are surprisingly often found in non-coding regions of the genome and hence do not overlap with protein-coding genes. It is hypothesized that these non-coding variants are linked to a dysregulated gene expression landscape, which are inherently cell type-specific. However, it is currently unknown for many of these variants which cell type they exert their effect in and how strong this effect is. Without this knowledge, efforts to develop targeted therapies are tainted by increased an increased risk of off-target effects. Since cell types always work in networks, a much better understanding of the molecular cascades and their interplay that ultimately lead to the neurodegenerative phenotype is needed in order for the scientific community to develop better diagnostic and therapeutic approaches.
The overarching goal of this project is to develop a method suitable to monitor and functionally interrogate these regions in a cell type-specific way. This is done in vivo with the help of mouse models.
This project officially ended in May 2020. Preliminary results suggest that our approach is expedient, feasible and translatable to disciplines outside of the neurodegeneration research field. Currently, we are collecting more data, after which the results will be disseminated to the scientific community.
- Creation and maintenance of specific mouse colonies (using an intercross of cell type-specific reporter mice and a mouse model of Alzheimer's disease)
- Writing an animal experimental protocol for issuance of a permit pertaining to the work on the above-mentioned animals
- Bioinformatic analysis of hundreds of variants associated with neurodegeneration, including testing their conservation between mouse and human; translating their conserved sequence from the human genome to the mouse genome in order to test human variants in mice; overlaying translated variants with publically available open chromatin data to pre-filter for cell type specific effects; sliding window analysis of the variants and association with the genome sequence;
- Creation of a randomly barcoded reporter plasmid library from scratch using the human-to-mouse translated regions described above
- Sequencing of the reporter library to determine the association of barcode to translated region
- Preparation of the plasmid library to be packaged into viral vectors
- Training in animal surgery in preparation for in vivo injections
Further experiments are still ongoing. Unfortunately, the Covid-19 outbreak has virtually halted all lab work for us and our collaborators, leading to a drastic slow-down in the completion of this project. As soon as results become available, they will be disseminated to the scientific community.