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Articles 1 - 6 of 6
Full-Text Articles in Medicine and Health Sciences
Measuring Plasma Membrane Protein Endocytic Rates By Reversible Biotinylation, Luke Gabriel, Zachary Stevens, Haley Melikian
Measuring Plasma Membrane Protein Endocytic Rates By Reversible Biotinylation, Luke Gabriel, Zachary Stevens, Haley Melikian
Haley Melikian
Plasma membrane proteins are a large, diverse group of proteins comprised of receptors, ion channels, transporters and pumps. Activity of these proteins is responsible for a variety of key cellular events, including nutrient delivery, cellular excitability, and chemical signaling. Many plasma membrane proteins are dynamically regulated by endocytic trafficking, which modulates protein function by altering protein surface expression. The mechanisms that facilitate protein endocytosis are complex and are not fully understood for many membrane proteins. In order to fully understand the mechanisms that control the endocytic trafficking of a given protein, it is critical that the protein s endocytic rate …
Insertion Of Tetracysteine Motifs Into Dopamine Transporter Extracellular Domains, Deanna M. Navaroli, Haley E. Melikian
Insertion Of Tetracysteine Motifs Into Dopamine Transporter Extracellular Domains, Deanna M. Navaroli, Haley E. Melikian
Haley Melikian
The neuronal dopamine transporter (DAT) is a major determinant of extracellular dopamine (DA) levels and is the primary target for a variety of addictive and therapeutic psychoactive drugs. DAT is acutely regulated by protein kinase C (PKC) activation and amphetamine exposure, both of which modulate DAT surface expression by endocytic trafficking. In order to use live imaging approaches to study DAT endocytosis, methods are needed to exclusively label the DAT surface pool. The use of membrane impermeant, sulfonated biarsenic dyes holds potential as one such approach, and requires introduction of an extracellular tetracysteine motif (tetraCys; CCPGCC) to facilitate dye binding. …
Amphetamine-Induced Decreases In Dopamine Transporter Surface Expression Are Protein Kinase C-Independent, Ekaterina Boudanova, Deanna Navaroli, Haley Melikian
Amphetamine-Induced Decreases In Dopamine Transporter Surface Expression Are Protein Kinase C-Independent, Ekaterina Boudanova, Deanna Navaroli, Haley Melikian
Haley Melikian
Amphetamine (AMPH) is a potent dopamine (DA) transporter (DAT) inhibitor that markedly increases extracellular DA levels. In addition to its actions as a DAT antagonist, acute AMPH exposure induces DAT losses from the plasma membrane, implicating transporter-specific membrane trafficking in amphetamine's actions. Despite reports that AMPH modulates DAT surface expression, the trafficking mechanisms leading to this effect are currently not defined. We recently reported that DAT residues 587-596 play an integral role in constitutive and protein kinase C (PKC)-accelerated DAT internalization. In the current study, we tested whether the structural determinants required for PKC-stimulated DAT internalization are necessary for AMPH-induced …
Dopamine Transporter Endocytic Determinants: Carboxy Terminal Residues Critical For Basal And Pkc-Stimulated Internalization, Ekaterina Boudanova, Deanna Navaroli, Zachary Stevens, Haley Melikian
Dopamine Transporter Endocytic Determinants: Carboxy Terminal Residues Critical For Basal And Pkc-Stimulated Internalization, Ekaterina Boudanova, Deanna Navaroli, Zachary Stevens, Haley Melikian
Haley Melikian
Dopamine (DA) reuptake terminates dopaminergic neurotransmission and is mediated by DA transporters (DATs). Acute protein kinase C (PKC) activation accelerates DAT internalization rates, thereby reducing DAT surface expression. Basal DAT endocytosis and PKC-stimulated DAT functional downregulation rely on residues within the 587-596 region, although whether PKC-induced DAT downregulation reflects transporter endocytosis mechanisms linked to those controlling basal endocytosis rates is unknown. Here, we define residues governing basal and PKC-stimulated DAT endocytosis. Alanine substituting DAT residues 587-590 1) abolished PKC stimulation of DAT endocytosis, and 2) markedly accelerated basal DAT internalization, comparable to that of wildtype DAT during PKC activation. Accelerated …
The Acid-Sensitive, Anesthetic-Activated Potassium Leak Channel, Kcnk3, Is Regulated By 14-3-3beta-Dependent, Pkc-Mediated Endocytic Trafficking, Luke Gabriel, Anatoli Lvov, Demetra Orthodoxou, Ann Rittenhouse, William Kobertz, Haley Melikian
The Acid-Sensitive, Anesthetic-Activated Potassium Leak Channel, Kcnk3, Is Regulated By 14-3-3beta-Dependent, Pkc-Mediated Endocytic Trafficking, Luke Gabriel, Anatoli Lvov, Demetra Orthodoxou, Ann Rittenhouse, William Kobertz, Haley Melikian
Haley Melikian
The acid-sensitive neuronal potassium leak channel, KCNK3, is vital for setting the resting membrane potential and is the primary target for volatile anesthetics. Recent reports demonstrate that KCNK3 activity is downregulated by PKC; however, the mechanisms responsible for PKC-induced KCNK3 downregulation are undefined. Here, we report that endocytic trafficking dynamically regulates KCNK3 activity. Phorbol esters and Group I mGluR activation acutely decreased both native and recombinant KCNK3 currents with concomitant KCNK3 surface losses in cerebellar granule neurons and cell lines. PKC-mediated KCNK3 internalization required the presence of both 14-3-3beta and a novel potassium channel endocytic motif, as depleting either 14-3-3beta …
The Plasma Membrane-Associated Gtpase Rin Interacts With The Dopamine Transporter And Is Required For Protein Kinase C-Regulated Dopamine Transporter Trafficking, Deanna M. Navaroli, Zachary H. Stevens, Zeljko Uzelac, Luke Gabriel, Michael J. King, Lawrence M. Lifshitz, Harald H. Sitte, Haley E. Melikian
The Plasma Membrane-Associated Gtpase Rin Interacts With The Dopamine Transporter And Is Required For Protein Kinase C-Regulated Dopamine Transporter Trafficking, Deanna M. Navaroli, Zachary H. Stevens, Zeljko Uzelac, Luke Gabriel, Michael J. King, Lawrence M. Lifshitz, Harald H. Sitte, Haley E. Melikian
Haley Melikian
Dopaminergic signaling and plasticity are essential to numerous CNS functions and pathologies, including movement, cognition, and addiction. The amphetamine- and cocaine-sensitive dopamine (DA) transporter (DAT) tightly controls extracellular DA concentrations and half-life. DAT function and surface expression are not static but are dynamically modulated by membrane trafficking. We recently demonstrated that the DAT C terminus encodes a PKC-sensitive internalization signal that also suppresses basal DAT endocytosis. However, the cellular machinery governing regulated DAT trafficking is not well defined. In work presented here, we identified the Ras-like GTPase, Rin (for Ras-like in neurons) (Rit2), as a protein that interacts with the …