Abstract
We expressed the synaptic vesicle proteins SV2, synaptotagmin, and synaptophysin in CHO fibroblasts to investigate the targeting information contained by each protein. All three proteins entered different cellular compartments. Synaptotagmin was found on the plasma membrane. Both SV2 and synaptophysin were sorted to small intracellular vesicles, but synaptophysin colocalized with early endosomal markers, while SV2 did not. SV2-containing vesicles did not have the same sedimentation characteristics as authentic synaptic vesicles, even though transfected SV2 was sorted from endosomal markers. We also created cell lines expressing both SV2 and synaptotagmin, both synaptotagmin and synaptophysin, and lines expressing all three synaptic vesicle proteins. In all cases, the proteins maintained their distinct compartmentalizations, were not found in the same organelle, and did not created synaptic vesicle-like structures. These results have important implications for models of synaptic vesicle biogenesis.
Highlights
We expressed the synaptic vesicle proteins SV2, synaptotagmin, and synaptophysin in CHO fibroblasts to investigate the targeting information contained by each protein
This result shows that nonneuronal cells can recognize and sort synaptic vesicle proteins, and further suggests that synaptic vesicles are endocytosed through conventional coated pit recycling with endocytic markers like the transferrin receptor, and are later restricted to a distinct vesicle population
To investigate the targeting information contained in the SV2 and synaptotagmin primary sequences, we expressed both proteins in CHO fibroblasts
Summary
We expressed the synaptic vesicle proteins SV2, synaptotagmin, and synaptophysin in CHO fibroblasts to investigate the targeting information contained by each protein. Synaptophysin is sorted to a small, endocytic vesicle population when expressed in most nonneuroendocrine cell lines (Leube et al, 1989; Johnston et al, 1989; Clift-O'Grady et al, 1990; Cameron et al, 1991; Linstedt and Kelly, 1991) This result shows that nonneuronal cells can recognize and sort synaptic vesicle proteins, and further suggests that synaptic vesicles are endocytosed through conventional coated pit recycling with endocytic markers like the transferrin receptor, and are later restricted to a distinct vesicle population. Synaptophysin may contribute endosomal targeting signals to the synaptic vesicle, expression of synaptophysin does not direct the formation of a small, uniform synaptic vesicle-like organdie. If these complexes exist in vivo, they could be extremely important in all aspects of synaptic vesicle function, including biogenesis and recycling
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