Uncovering the transcriptional landscape of astrocytes highlights glial actin dynamics as important for neuronal remodeling

Abstract

Developmental neuronal remodeling is an evolutionary conserved mechanism to sculpt the mature nervous system. It often involves pruning of exuberant connections as a mechanism to refine neural circuits. Understanding the molecular mechanisms that regulate developmental remodeling may shed light on processes like axon elimination during development and disease. Interestingly, glia were found to be involved in many aspects of neuronal remodeling. However, the full extent of these glia-neuron interactions remains unknown. To identify glial molecules that affect Drosophila mushroom body (MB) axon pruning, we performed RNA-seq analysis of larval and adult astrocytes. Since pruning occurs at early pupal stages, we focused on genes specifically enriched in third instar larva astrocytes. We screened 21 genes by knocking down their expression in glia and visualizing MB remodeling in parallel. Thus far, we identified 8 candidates which seem to be required in glia for proper MB axon pruning. For example, Arpc1, which belongs to the evolutionary conserved Arp2/3 complex that governs F-actin polymerization, is required in astrocytes for MB remodeling. This led us to test other Arp2/3 complex proteins, as well as different formin nucleators, and indeed we found that glial expression of these genes was required for MB axon pruning. Interestingly, interfering with glial F-actin dynamics did not affect their migration or gross morphology. However, we found that astrocyte specific knock down of Arpc1 induced defects in the astrocytes’ ability to infiltrate the axonal bundle at the onset of pruning. Remarkably, decreasing axonal adhesion suppressed the pruning defect phenotype caused by astrocytic Arpc1 knockdown, suggesting that actin dependent astrocytic infiltration is a key step in axon pruning. In summary, we have identified a yet unknown actin-based mechanism by which glia affect neuronal remodeling. Furthermore, this study offers a unique opportunity to dissect the mechanisms of neuron-glia interactions during developmental remodeling.