Axonal targeting of the serotonin transporter in cultured rat dorsal raphe neurons is specified by SEC24C-dependent export from the endoplasmic reticulum.
Export of the serotonin transporter (SERT) from the endoplasmic reticulum (ER) is mediated by the SEC24C isoform of the coatomer protein-II complex. SERT must enter the axonal compartment and reach the presynaptic specialization to perform its function, i.e., the inward transport of serotonin. Refilling of vesicles is contingent on the operation of an efficient relay between SERT and the vesicular monoamine transporter-2 (VMAT2). Here, we visualized the distribution of both endogenously expressed SERT and heterologously expressed variants of human SERT in dissociated rat dorsal raphe neurons to examine the role of SEC24C-dependent ER export in axonal targeting of SERT. We conclude that axonal delivery of SERT is contingent on recruitment of SEC24C in the ER. This conclusion is based on the following observations. (1) Both endogenous and heterologously expressed SERT were delivered to the extensive axonal arborizations and accumulated in bouton-like structures. (2) In contrast, SERT-(607)RI(608)-AA, in which the binding site of SEC24C is disrupted, remained confined to the microtubule-associated protein 2-positive somatodendritic compartment. (3) The overexpression of dominant-negative SEC24C-D(796)V/D(797)N (but not of the corresponding SEC24D mutant) redirected both endogenous SERT and heterologously expressed yellow fluorescent protein-SERT from axons to the somatodendritic region. (4) SERT-K(610)Y, which harbors a mutation converting it into an SEC24D client, was rerouted from the axonal to the somatodendritic compartment by dominant-negative SEC24D. In contrast, axonal targeting of the VMAT2 was disrupted by neither dominant-negative SEC24C nor dominant-negative SEC24D. This suggests that SERT and VMAT2 reach the presynaptic specialization by independent routes.
Institute of Pharmacology, Center of Physiology and Pharmacology, Medical University of Vienna, A-1090 Vienna, Austria.|UCD Conway Institute of Biomolecular and Biomedical Research, University College Dublin, Belfield, Dublin 4, Ireland.
Therese R
TR
Montgomery
Institute of Pharmacology, Center of Physiology and Pharmacology, Medical University of Vienna, A-1090 Vienna, Austria.
Thomas
T
Steinkellner
Institute of Pharmacology, Center of Physiology and Pharmacology, Medical University of Vienna, A-1090 Vienna, Austria.
Sonja
S
Sucic
Institute of Pharmacology, Center of Physiology and Pharmacology, Medical University of Vienna, A-1090 Vienna, Austria.
Florian
F
Koban
Department of Molecular Genetics, Medical University of Vienna, A-1090 Vienna, Austria.
Stefan
S
Schüchner
Department of Molecular Genetics, Medical University of Vienna, A-1090 Vienna, Austria.
Egon
E
Ogris
Institute of Pharmacology, Center of Physiology and Pharmacology, Medical University of Vienna, A-1090 Vienna, Austria.
Harald H
HH
Sitte
Institute of Pharmacology, Center of Physiology and Pharmacology, Medical University of Vienna, A-1090 Vienna, Austria.
Michael
M
Freissmuth
D000818:Animals; D001369:Axons / Q000378:metabolism*; D002478:Cells, Cultured; D050483:Dopamine Plasma Membrane Transport Proteins / Q000378:metabolism; D004721:Endoplasmic Reticulum / Q000378:metabolism*; D005260:Female; D006801:Humans; D008297:Male; D051379:Mice; D008822:Mice, Transgenic; D009154:Mutation / Q000235:genetics; D009474:Neurons / Q000378:metabolism / Q000648:ultrastructure*; D021381:Protein Transport / Q000235:genetics / Q000502:physiology; D050783:Proto-Oncogene Proteins c-ets / Q000235:genetics; D011903:Raphe Nuclei / Q000166:cytology*; D051381:Rats; D050486:Serotonin Plasma Membrane Transport Proteins / Q000235:genetics; D013574:Synaptosomes / Q000378:metabolism; D014365:Tryptophan Hydroxylase / Q000378:metabolism; D033921:Vesicular Transport Proteins / Q000235:genetics / Q000378:metabolism*
D050483:Dopamine Plasma Membrane Transport Proteins; C086924:Elk3 protein, mouse; D050783:Proto-Oncogene Proteins c-ets; D050486:Serotonin Plasma Membrane Transport Proteins; C493231:Slc6a4 protein, mouse; D033921:Vesicular Transport Proteins; D014365:Tryptophan Hydroxylase; C504569:tph2 protein, rat
axonal delivery; raphe neurons; serotonin transporter; somatodendritic targeting; vesicular monoamine transporter-2