Serotonin transporter (SERT) is responsible for lowering serotonin (5-HT) levels in the vicinity of synapses that release this neurotransmitter. In this process, 5-HT is transported into the cell from which it was released, where it is available for repackaging into synaptic vesicles. Drugs that inhibit SERT increase the level of synaptic 5-HT and lengthen the duration of its action. These drugs include abused substances such as cocaine, and therapeutic drugs such as antidepressants. Some drugs that are substrates for SERT, such as 3,4-methylenedioxymethamphetamine (a.k.a. "ecstasy") act through SERT to release 5-HT from neurons. From the profound behavioral consequences of these drugs, it is clear that regulation of SERT activity is likely to be a key event in normal brain physiology. Five years ago we found a mutant form of the transporter in patients with obsessive compulsive disorder. This mutant is apparently activated in a constitutive fashion by a pathway normally involving cyclic GMP (cGMP). We have since discovered that the defect represents a defect in the ability to remove a phosphate group at position 276 in the SERT molecule. This position is phosphorylated by a cGMP-dependent enzyme, PKG. As part of the long term goal to understand the structure and function of neurotransmitter transporters, this proposal outlines plans to investigate the association of enzymes that phosphorylate and dephosphorylate SERT, along with other components of the cGMP signaling pathway. The mechanism by which SERT conformation is affected by phosphorylation will also be investigated. All of these studies are directed to testing the hypothesis that production of cGMP activates a signaling pathway that phosphorylates and dephosphorylates SERT with the consequent increase and decrease in turnover rate. We will also test the prediction that conformational changes underlie the effects that 5-HT transport has on SERT interaction with regulatory proteins.

Public Health Relevance

This proposal concerns a mechanism for regulation of serotonin transporter, a target for drugs of abuse and antidepressants. Defects in regulation were found in a mutant form of this transporter associated with psychiatric disorders.
We aim to understand the structural and mechanistic events that are responsible for this regulation at a molecular level.

Agency
National Institute of Health (NIH)
Institute
National Institute on Drug Abuse (NIDA)
Type
Research Project (R01)
Project #
5R01DA008213-18
Application #
8429477
Study Section
Neurotransporters, Receptors, and Calcium Signaling Study Section (NTRC)
Program Officer
Pilotte, Nancy S
Project Start
1993-05-01
Project End
2015-01-31
Budget Start
2013-02-01
Budget End
2014-01-31
Support Year
18
Fiscal Year
2013
Total Cost
$308,227
Indirect Cost
$121,987
Name
Yale University
Department
Pharmacology
Type
Schools of Medicine
DUNS #
043207562
City
New Haven
State
CT
Country
United States
Zip Code
06520
Tavoulari, Sotiria; Margheritis, Eleonora; Nagarajan, Anu et al. (2016) Two Na+ Sites Control Conformational Change in a Neurotransmitter Transporter Homolog. J Biol Chem 291:1456-71
Zhang, Yuan-Wei; Turk, Benjamin E; Rudnick, Gary (2016) Control of serotonin transporter phosphorylation by conformational state. Proc Natl Acad Sci U S A 113:E2776-83
Rudnick, Gary; Krämer, Reinhard; Blakely, Randy D et al. (2014) The SLC6 transporters: perspectives on structure, functions, regulation, and models for transporter dysfunction. Pflugers Arch 466:25-42
Fenollar-Ferrer, Cristina; Stockner, Thomas; Schwarz, Thomas C et al. (2014) Structure and regulatory interactions of the cytoplasmic terminal domains of serotonin transporter. Biochemistry 53:5444-60
Porton, B; Greenberg, B D; Askland, K et al. (2013) Isoforms of the neuronal glutamate transporter gene, SLC1A1/EAAC1, negatively modulate glutamate uptake: relevance to obsessive-compulsive disorder. Transl Psychiatry 3:e259
Rudnick, Gary (2013) How do transporters couple solute movements? Mol Membr Biol 30:355-9
Schicker, Klaus; Uzelac, Zeljko; Gesmonde, Joan et al. (2012) Unifying concept of serotonin transporter-associated currents. J Biol Chem 287:438-45
Bulling, Simon; Schicker, Klaus; Zhang, Yuan-Wei et al. (2012) The mechanistic basis for noncompetitive ibogaine inhibition of serotonin and dopamine transporters. J Biol Chem 287:18524-34
Wong, Albert; Zhang, Yuan-Wei; Jeschke, Grace R et al. (2012) Cyclic GMP-dependent stimulation of serotonin transport does not involve direct transporter phosphorylation by cGMP-dependent protein kinase. J Biol Chem 287:36051-8
Rudnick, Gary (2011) Cytoplasmic permeation pathway of neurotransmitter transporters. Biochemistry 50:7462-75

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