Vglut1 and ZnT3 co-targeting mechanisms regulate vesicular zinc stores in PC12 cells

• Published in: Journal of cell science Vol. 118; no. 9; pp. 1911 - 1921
• Main Authors: Salazar, Gloria, Craige, Branch, Love, Rachal, Kalman, Daniel, Faundez, Victor
• Format: Journal Article
• Language: English
• Published: England 01.05.2005
• Subjects: Animals; Biological Transport; Brain - metabolism; Carrier Proteins - metabolism; Carrier Proteins - physiology; Cation Transport Proteins - metabolism; Cation Transport Proteins - pharmacokinetics; Dose-Response Relationship, Drug; Flow Cytometry; Glutamic Acid - chemistry; Ions; Membrane Proteins - metabolism; Membrane Proteins - physiology; Membrane Transport Proteins - metabolism; Membrane Transport Proteins - physiology; Microscopy, Fluorescence; PC12 Cells; Plasmids - metabolism; Protein Transport; Rats; Subcellular Fractions - chemistry; Synapses - metabolism; Synaptic Vesicles - metabolism; Vesicular Glutamate Transport Protein 1; Zinc - chemistry; Zinc - metabolism
• ISSN: 0021-9533; 1477-9137
• DOI: 10.1242/jcs.02319

The lumenal ionic content of an organelle is determined by its complement of channels and transporters. These proteins reach their resident organelles by adaptor-dependent mechanisms. This concept is illustrated in AP-3 deficiencies, in which synaptic vesicle zinc is depleted because the synaptic-vesicle-specific zinc transporter 3 does not reach synaptic vesicles. However, whether zinc transporter 3 is the only membrane protein defining synaptic-vesicle zinc content remains unknown. To address this question, we examined whether zinc transporter 3 and the vesicular glutamate transporter Vglut1 (a transporter that coexists with zinc transporter 3 in brain nerve terminals) were co-targeted to synaptic-like microvesicle fractions in PC12 cells. Deconvolution microscopy and subcellular fractionation demonstrated that these two transporters were present on the same vesicles in PC12 cells. Vglut1 content in synaptic-like microvesicle fractions and brain synaptic vesicles was partially sensitive to pharmacological and genetic perturbation of AP-3 function. Whole-cell flow-cytometry analysis of PC12 cell lines expressing zinc transporter 3, Vglut1 or both showed that vesicular zinc uptake was increased by Vglut1 expression. Conversely, production of zinc transporter 3 increased the vesicular uptake of glutamate in a zinc-dependent fashion. Our results suggest that the coupling of zinc transporter 3 and Vglut1 transport mechanisms regulates neurotransmitter content in secretory vesicles.