Regulatory links between transcription initiation and co-transcriptional processes in neurons

• Applicant: Dr. Valerie Hilgers
• Subject Area: Developmental Biology ; General Genetics and Functional Genome Biology
• Term: since 2022
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 500335138

Neurons are highly polarized cells of complex and dynamic architecture. The formation and maintenance of neural cells and neural circuits require the coordinated expression of genes at each step of RNA metabolism: from transcription, processing, localized transport and translation, to degradation. To achieve this level of complexity, neurons employ mechanisms that increase RNA regulatory potential. The extent of neuronal RNA diversity, and the high number of mRNA isoforms and non-coding RNAs present in neurons, have only been appreciated in recent years with the emergence of new transcriptomics technologies. The regulatory links between transcriptional, co-transcriptional and post-transcriptional processes are still poorly understood. Neurological diseases are virtually always associated with RNA regulatory dysfunction; to understand the molecular processes underlying such pathologies, we need to gain mechanistic and functional insight into how RNA diversity is regulated. The highly conserved RNA-binding protein ELAV is expressed in all neurons across the animal kingdom. While ELAV proteins are prominent for their role in numerous human neurological diseases and are required for neuronal differentiation, their molecular function and associated mechanisms are not well understood. Our previous work in Drosophila uncovered that ELAV determines neuronal transcript signatures such as neuron-specific alternative splicing and neuron-specific alternative polyadenylation. Moreover, we showed for the first time, a functional link between regulation of transcription initiation and alternative polyadenylation. In this proposal, we intend to discover how distinct, neuron-specific transcriptional events are co-regulated: circular RNA (circRNA) formation, alternative splicing, and alternative polyadenylation. Our preliminary data indicate that the three processes are mechanistically linked and controlled by ELAV at transcription initiation. First, employing several complementary ultra-long-read-sequencing techniques in Drosophila brain tissue, we will define entire mRNA isoforms from 5’end to 3’end, and uncover regulatory links between transcription initiation, splicing and transcription termination in vivo. Functional genetics and RNA biochemistry studies will reveal the molecular mechanisms of how ELAV dictates isoform choice in a neuron-specific manner. Second, we will determine how ELAV modulates circRNA synthesis and homeostasis in vivo, using functional transcriptomics in sorted elav mutant neurons, as well as ELAV binding studies. Finally, we will integrate results to obtain a global understanding of the mechanisms that co-regulate transcriptional processes to generate the neuron-specific RNA landscape.

• DFG Programme: Research Grants