Kristín Allison, Stefán Eggertsson, Zophonías Oddur Jónsson and Sigríður Rut Franzdóttir
Introduction: Drosophila melanogaster motor neurons share multiple similarities with their vertebrate counterparts, offering a tactile system to study the molecular cell biology of axons and neuromuscular junctions, NMJs. A conserved protein complex consisting of Pontin and Reptin is required for the assembly and function of numerous multimolecular complexes and cellular processes in eukaryotes. The proteins have been implicated in aggregate clearance as well as in association with RNP granules – both of which are important for neuronal survival and function, with implications for neurodegenerative disorders. The proteins are expressed in the nervous system of various taxa but their roles in neurons have not been described. The aim of this study was to determine the roles of Pontin and Reptin in post-mitotic neurons, focusing on motor neurons and NMJs during larval stages.
Methods: The Gal4/UAS system was used to knock down pontin and reptin expression by RNA interference in Drosophila larval motor neurons. Endogenously tagged Pontin and Reptin lines were generated using CRISPR/Cas9. 3rd instar larval fillets were immunofluorescently stained for confocal and STED super-resolution imaging and analysis.
Results and discussion: Loss of Pontin and Reptin in motor neurons causes synaptic bouton morphology defects. Furthermore, reduced survival and locomotion in adults, combined with accumulation of misfolded proteins, points to defective proteostasis. Pontin and Reptin were found throughout axons and NMJs, indicating that the observed phenotypes may reflect direct axonal roles. We are presently unravelling the molecular and subcellular details of these phenotypes and determining the underlying causative molecular relationships.
