(a) Larval Drosophila neuromuscular junction. The fruit fly neuromuscular junctions exhibit several key features in common with the excitatory synapses in the vertebrate brain. They utilize glutamate as the major neurotransmitter, and hence have been used as a excellent model to study the molecular mechanisms of synaptic development and functions. Green, motor neuron termini labeled with an anti-neuronal antibody. Magenta, synaptic boutons labeled with anti-Synaptotagmin. Scale bar, 40 μm. (band c) Proximity Ligation Assay in larval neuromuscular junctions. To determine whether DISC1directly interacts with Dysbindin in synaptogenesis, we applied the Proximity Ligation Assay technique. In this assay, the target proteins are localized with specific primary antibodies, which are then detected with secondary antibodies conjugated to oligonucleotide probes. The attached probes can be bridged through hybridization of connector oligonucleotides only when the two proteins are in close proximity to make direct contact. The annealed oligonucleotides are then closed by ligation into circular DNA molecules, which serve as templates for subsequent rolling circle amplification to be visualized by in situ hybridization. Green, motor neuron termini labeled with anti-HRP. Magenta, Proximity Ligation Assay signals between the Drosophila Dysbindin and the human DISC1 protein. Scale bars, 10 μm (b) and 5 μm (c). For more information on this topic, please refer to the article by Furukubo-Tokunaga et al. on pages 1232–1243.
This is a preview of subscription content, access via your institution