Synaptic localization and restricted diffusion of a Drosophila neuronal synaptobrevin - Green fluorescent protein chimera in vivo

Fluorescent markers for subcellular compartments in Drosophila neurons shou ld allow one to combine genetic mutant analysis with visualization of subce llular structures in vivo. Here we describe an analysis of two markers whic h may be used to observe different compartments of live Drosophila synapses . Soluble jellyfish green fluorescent protein (GFP) expressed at high level s in neurons diffuses freely in the neuronal cytosol as evidenced by confoc al microscopy and fluorescence recovery from photobleaching experiments. Th us, the distribution pattern of soluble GFP in motor axons and larval motor terminals indicates the expected distribution For diffusible presynaptic m olecules. In contrast to GFP, a neurally expressed neuronal synaptobrevin-G FP chimera (n-syb GFP) is transported down axons and specifically localized to nerve terminals. We demonstrate that n-syb GFP labels synaptic-vesicle membrane at larval motor terminals by documenting its restriction to presyn aptic varicosities, its colocalization with synaptic vesicle antigens, and its redistribution in Drosophila shi(ts1) mutant nerve terminals transientl y depleted of synaptic vesicles. Surprisingly, n-syb GFP expressed in muscl e is concentrated at the subsynaptic reticulum (SSR), postsynaptic infoldin gs of muscle plasma membrane. We suggest, using different membrane markers, that this apparent postsynaptic enrichment simply reflects a concentration of plasma membrane in the SSR, rather than a selective targeting of n-syb GFP to postsynaptic sites. Utilities and implications of these studies are demonstrated or discussed.