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Publication
Single-quantum-dot tracking reveals altered membrane dynamics of an
attention-deficit/hyperactivity-disorder-derived dopamine transporter coding
variant.
Authors Kovtun O, Sakrikar D, Tomlinson ID, Chang JC, Arzeta-Ferrer X, Blakely RD,
Rosenthal SJ
Submitted By Submitted Externally on 7/24/2015
Status Published
Journal ACS chemical neuroscience
Year 2015
Date Published 4/15/2015
Volume : Pages 6 : 526 - 534
PubMed Reference 25747272
Abstract The presynaptic, cocaine- and amphetamine-sensitive dopamine (DA) transporter
(DAT, SLC6A3) controls the intensity and duration of synaptic dopamine signals
by rapid clearance of DA back into presynaptic nerve terminals. Abnormalities in
DAT-mediated DA clearance have been linked to a variety of neuropsychiatric
disorders, including addiction, autism, and attention deficit/hyperactivity
disorder (ADHD). Membrane trafficking of DAT appears to be an important, albeit
incompletely understood, post-translational regulatory mechanism; its
dysregulation has been recently proposed as a potential risk determinant of
these disorders. In this study, we demonstrate a link between an ADHD-associated
DAT mutation (Arg615Cys, R615C) and variation on DAT transporter cell surface
dynamics, a combination only previously studied with ensemble biochemical and
optical approaches that featured limited spatiotemporal resolution. Here, we
utilize high-affinity, DAT-specific antagonist-conjugated quantum dot (QD)
probes to establish the dynamic mobility of wild-type and mutant DATs at the
plasma membrane of living cells. Single DAT-QD complex trajectory analysis
revealed that the DAT 615C variant exhibited increased membrane mobility
relative to DAT 615R, with diffusion rates comparable to those observed after
lipid raft disruption. This phenomenon was accompanied by a loss of transporter
mobilization triggered by amphetamine, a common component of ADHD medications.
Together, our data provides the first dynamic imaging of single DAT proteins,
providing new insights into the relationship between surface dynamics and
trafficking of both wild-type and disease-associated transporters. Our approach
should be generalizable to future studies that explore the possibilities of
perturbed surface DAT dynamics that may arise as a consequence of genetic
alterations, regulatory changes, and drug use that contribute to the etiology or
treatment of neuropsychiatric disorders.




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