Periodic Reporting for period 3 - Neuro-UTR (Mechanism and functional impact of ultra-long 3’ UTRs in the Drosophila nervous system)
Okres sprawozdawczy: 2022-01-01 do 2023-06-30
One particularly striking process is the recently discovered, drastic lengthening of the 3’ untranslated region (3’ UTR) of hundreds of genes, which occurs in neurons from flies to humans. The function of the resulting ultra-long 3’ UTRs is unknown. RNA deregulation plays a central role in neurological diseases; to understand underlying causes, it is essential to study regulatory processes and define the function of these novel 3’ UTRs.
Our study integrates the molecular mechanisms that govern biogenesis and function of ultra-long 3’ UTRs, from nucleus to synapse, in an animal model, the fruit fly Drosophila melanogaster. The results of this research will create a major impact on our understanding of neuronal gene regulation in health and disease.
We generated and optimized a protocol for global aUTR localisation in neurons of adult Drosophila brains, successfully implemented it and validated the purity of synaptosome fractions by electron microscopy. We developed and successfully applied several complementary RNA biochemistry approaches. A preliminary analysis of the data substantiates our work hypothesis that extended 3’UTRs undergo specific posttranscriptional regulation through specific proteins. These important findings on the global regulation of aUTRs in neurons significantly advance our understanding on post-transcriptional regulation of neuronal RNA signatures.
We developed a robust computational approach to quantify alternative poly(A) site usage from traditional mRNA-seq datasets, which are more prevalent than 3’-end sequencing datasets. Hence, our new tool can perform specialised analyses on publicly available datasets, which is a valuable resource for the RNA community.
These methods and their application in the scope of this project will reveal mechanisms of how alternative UTRs regulate gene expression complexity in neurons.
We generated and analysed flies lacking the aUTR of specific genes, and found one particularly interesting disease phenotype, which we are focusing on for functional and molecular characterization. We have made progress in the construction of genetic tools for global up- and down-regulation of aUTRs in vivo. Due to our unexpected finding of the involvement of FNE in aUTR regulation, we adjusted experimental setups and the design of in vivo mutagenesis, which will enable to create better models of aUTR deregulation, and better understand how neuronal UTRs regulate neuronal homeostasis in health and disease.