Unravelling the function of circRmst in the development of midbrain dopaminergic neurons

Abstract

Circular RNAs (circRNAs) are covalently closed non-coding RNA (ncRNA) molecules which are formed by an alternative splicing mechanism called backsplicing. Due to recent improvements in RNA sequencing techniques, circRNAs have been discovered to be widely expressed in a tissue-specific and cell type-specific manner and expression patterns differ throughout embryonic development. Because of the relatively high expression of circRNAs in the developing brain, combined with the fact that many other types of ncRNA play a role in brain development, it is interesting to study the role of circRNAs in this process. Previously in this project, multiple circRNAs specific to midbrain dopaminergic (mDA) neurons in the ventral midbrain (vMB) were found. Among them circular rhabdomyosarcoma 2-associated transcript (circRmst) was particularly highly expressed in mDA neurons. circRmst also had a change in expression during development. Therefore, the main aim was to discover the function of circRmst in the development of mDA neurons. Using lentivirus-delivered short hairpin RNAs (shRNA), circRmst was knocked down in embryonic vMB primary cultures, which caused an increase in soma size and complexity, and a decreased amount of mDA neurons. In vivo knockdown of circRmst, induced using in utero electroporation (IUE), led to altered position of mDA neurons. A function of circRmst is proposed in which circRmst could regulate size and migration of mDA neurons by altering the cytoskeleton. Furthermore, circRmst could be involved in mDA neuron differentiation or survivability. This study provides the first evidence that circRmst plays a role in mDA neuron development.