title: Ctr9, a Protein in the Transcription Complex Paf1, Regulates Dopamine Transporter Activity at the Plasma Membrane.
abstract: Dopamine (DA) is a major regulator of sensorimotor and cognitive functions. The DA transporter (DAT) is the key protein that regulates the spatial and temporal activity of DA release into the synaptic cleft via the rapid reuptake of DA into presynaptic termini. Several lines of evidence have suggested that transporter-interacting proteins may play a role in DAT function and regulation. Here, we identified the tetratricopeptide repeat domain-containing protein Ctr9 as a novel DAT binding partner using a yeast two-hybrid system. We showed that Ctr9 is expressed in dopaminergic neurons and forms a stable complex with DAT in vivo via GST pulldown and co-immunoprecipitation assays. In mammalian cells co-expressing both proteins, Ctr9 partially colocalizes with DAT at the plasma membrane. This interaction between DAT and Ctr9 results in a dramatic enhancement of DAT-mediated DA uptake due to an increased number of DAT transporters at the plasma membrane. We determined that the binding of Ctr9 to DAT requires residues YKF in the first half of the DAT C terminus. In addition, we characterized Ctr9, providing new insight into this protein. Using three-dimensional modeling, we identified three novel tetratricopeptide repeat domains in the Ctr9 sequence, and based on deletion mutation experiments, we demonstrated the role of the SH2 domain of Ctr9 in nuclear localization. Our results demonstrate that Ctr9 localization is not restricted to the nucleus, as previously described for the transcription complex Paf1. Taken together, our data provide evidence that Ctr9 modulates DAT function by regulating its trafficking.
authors.affiliation: INSERM U952, 75005 Paris, France; CNRS UMR 7224, 75005 Paris, France; Université Pierre et Marie Curie, Neuroscience Paris Seine, 75005 Paris, France; Douglas Hospital Research Center, Department of Psychiatry, McGill University, Montreal H4H 1R3 Quebec, Canada.
authors.forename: Stéphanie
authors.initials: S
authors.lastname: De Gois
authors.affiliation: INSERM U952, 75005 Paris, France; CNRS UMR 7224, 75005 Paris, France; Université Pierre et Marie Curie, Neuroscience Paris Seine, 75005 Paris, France.
authors.forename: Patrick
authors.initials: P
authors.lastname: Slama
authors.affiliation: Université Paris Descartes, Sorbonne Paris Cité, 75006 Paris, France; CNRS, UMR 8601, 75006 Paris, France.
authors.forename: Nicolas
authors.initials: N
authors.lastname: Pietrancosta
authors.affiliation: INSERM U952, 75005 Paris, France; CNRS UMR 7224, 75005 Paris, France; Université Pierre et Marie Curie, Neuroscience Paris Seine, 75005 Paris, France.
authors.forename: Amaia M
authors.initials: AM
authors.lastname: Erdozain
authors.affiliation: INSERM U952, 75005 Paris, France; CNRS UMR 7224, 75005 Paris, France; Université Pierre et Marie Curie, Neuroscience Paris Seine, 75005 Paris, France.
authors.forename: Franck
authors.initials: F
authors.lastname: Louis
authors.affiliation: INSERM U952, 75005 Paris, France; CNRS UMR 7224, 75005 Paris, France; Université Pierre et Marie Curie, Neuroscience Paris Seine, 75005 Paris, France.
authors.forename: Caroline
authors.initials: C
authors.lastname: Bouvrais-Veret
authors.affiliation: Hybrigenics, 3-5 Impasse Reille, 75014 Paris, France.
authors.forename: Laurent
authors.initials: L
authors.lastname: Daviet
authors.affiliation: INSERM U952, 75005 Paris, France; CNRS UMR 7224, 75005 Paris, France; Université Pierre et Marie Curie, Neuroscience Paris Seine, 75005 Paris, France; Douglas Hospital Research Center, Department of Psychiatry, McGill University, Montreal H4H 1R3 Quebec, Canada. Electronic address: bruno.giros@mcgill.ca.
authors.forename: Bruno
authors.initials: B
authors.lastname: Giros
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D002460:Cell Line; D050483:Dopamine Plasma Membrane Transport Proteins / Q000032:analysis / Q000378:metabolism*; D006801:Humans; D062105:Molecular Docking Simulation; D008969:Molecular Sequence Data; D009687:Nuclear Proteins / Q000032:analysis / Q000378:metabolism*; D010750:Phosphoproteins / Q000032:analysis / Q000378:metabolism*; D054730:Protein Interaction Domains and Motifs; D021381:Protein Transport; D051381:Rats; D014157:Transcription Factors; D018909:src Homology Domains
chemical_list: C484387:CTR9 protein, human; C000602042:CTR9 protein, rat; D050483:Dopamine Plasma Membrane Transport Proteins; D009687:Nuclear Proteins; D010750:Phosphoproteins; D014157:Transcription Factors
keywords: dopamine transporter; membrane targeting; membrane trafficking; membrane transport; protein-protein interaction; tetratricopeptide domain repeat; transcription factor; uptake activity