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Open Access

Truncations in the amino terminus reveal a region key to supporting amphetamine-induced efflux by the human serotonin transporter

  • Sonja Sucic1,
  • Carina Kern1,
  • Dragana Vidovic1,
  • Subhodeep Sarker1,
  • Oliver Kudlacek1,
  • Harald H Sitte1 and
  • Michael Freissmuth1Email author
BMC Pharmacology201111(Suppl 2):A20

https://doi.org/10.1186/1471-2210-11-S2-A20

Published: 5 September 2011

Background

The serotonin transporter (SERT) terminates neurotransmission via reuptake of serotonin from the synaptic cleft. Upon stimulation with amphetamines, SERT switches into an outward transport mode to rapidly release serotonin. We have previously shown that truncation of the first 64 residues of SERT amino terminus leads to loss of amphetamine-induced efflux [1]. This was comparable to the effects of a single point mutation of a juxtamembrane threonine residue at position 81 [1].

Methods

Truncation mutants of SERT amino terminus were generated by removing 22 (Δ22-SERT), 32 (Δ32-SERT) or 42 (Δ42-SERT) amino terminal residues. In addition, alanine scanning mutagenesis was performed along a segment of amino acid residues 32–42. All mutants were pharmacologically characterised in uptake, binding and efflux studies.

Results

Cellular localisation of the mutants examined by confocal microscopy, revealed no differences compared to the wild-type SERT. Functional analysis showed only modest changes in their substrate uptake properties (no significant changes in the Km values and a moderate decrease in the Vmax value of Δ42-SERT). Similarly, there were no marked alterations in the KD and Bmax values of imipramine or in the Ki values of p-chloroamphetamine and ibogaine, determined in radiolabelled imipramine binding assays. However, while amphetamine-induced efflux was unimpaired for Δ22-SERT and to a slight extent decreased for Δ32-SERT, it was completely abolished for Δ42-SERT.

Conclusions

Our results shed new light on the functional role of the amino terminus and point to the segment encompassing residues 32–42 as a region of key importance to supporting amphetamine-induced efflux by SERT.

Declarations

Acknowledgements

This work was supported by SFB35.

Authors’ Affiliations

(1)
Institute of Pharmacology, Center of Physiology and Pharmacology, Medical University of Vienna

References

  1. Sucic S, Dallinger S, Zdrazil B, Weissensteiner R, Jørgensen TN, Holy M, Kudlacek O, Seidel S, Cha JH, Gether U, Newman AH, Ecker GF, Freissmuth M, Sitte HH: The N terminus of monoamine transporters is a lever required for the action of amphetamines. J Biol Chem. 2010, 285: 10924-10938. 10.1074/jbc.M109.083154.PubMed CentralView ArticlePubMedGoogle Scholar

Copyright

© Sucic et al; licensee BioMed Central Ltd. 2011

This article is published under license to BioMed Central Ltd. This is an open access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

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