Synaptic plasticity is one of the underlying mechanisms for learning and memory, and dysregulated synaptic pruning is linked to human neurological disorders. Our studies of remodeling C. elegans GABAergic neurons,have revealed a critical role for the actin cytoskeleton in synapse elimination. Dorsal D (DD) GABAergic motor neurons initially form synapses with ventral muscles which then relocate to dorsal muscles during early larval development1. The DD remodeling scheme, however, is normally blocked in Ventral D (VD) GABAergic motor neurons by the COUP-TF/UNC-55 transcription factor.The Miller lab has previously used neuron-specific expression profiling to identify
unc-55-regulated transcripts and global RNAi to show that one of these candidates, ARX-5/p21, a component of the Arp2/3 complex, promotes removal of the presynaptic apparatus in remodeling VD neurons2. Because the Arp2/3 complex nucleates assembly of branched actin networks, this finding suggests that actin polymerization also drives synapse elimination. Here we used cell-specific RNAi to show that ARX-5/p21 is required for removal of presynaptic domains in the native DD remodeling program. For this experiment, we used the
flp-13 promoter to drive co-expression of
arx-5 sense and antisense strands specifically in DD neurons. Two expression plasmids were co-injected, one expressing the
arx-5 sense strand and cytosolic mCherry and the other expressing the
arx-5 antisense strand and nuclear-localized GFP. DD cells showing both cytosolic mCherry (red) and nuclear GFP (green) should express double-stranded RNA (dsRNA) for cell-specific knockdown of
arx-5. We used SNB-1::GFP/Synaptobrevin to visualize DD neuron presynaptic terminals. In wild-type DD neurons, few SNB-1::GFP puncta are retained on the ventral side at the L4 stage due to remodeling. In contrast, DD neurons that express
arx-5 dsRNA retain substantially moreventral SNB-1::GFP fluorescence (p<0.01, N=13) than wild type. Thus, our results suggest that the Arp2/3 complex acts cell-autonomously in DD neurons to promote presynaptic removal.