Serotonin-, protein kinase C-, and Hic-5-associated redistribution of the platelet serotonin transporter.
Emerging data indicate the existence of multiple regulatory processes supporting serotonin (5HT) transporter (SERT) capacity including regulated trafficking and catalytic activation, influenced by post-translational modifications and transporter-associated proteins. In the present study, using differential extraction and sedimentation procedures optimized for the purification of cytoskeletal and membrane-skeletal associated proteins, we analyze SERT localization in platelets. We find that most of the plasma membrane SERT is associated with the membrane skeleton. This association can be enhanced by both transporter activation and 5HT2A receptor activation. Inactivation of transport activity by phorbol ester treatment of intact platelets relocates SERT to the cytoskeleton fraction, consequently leading to transporter internalization. The translocation of SERT between these compartments is correlated with changes in the interaction with the LIM domain adaptor protein Hic-5. Co-immunoprecipitation and uptake activity studies suggest that Hic-5 is a determinant of transporter inactivation and relocation to a compartment subserving endocytic regulation. Associations of SERT with Hic-5 are evident in brain synaptosomes, suggesting the existence of parallel mechanisms operating to regulate SERT at serotonergic synapses.
Carneiro
Ana Marin D
AM
Departments of Pharmacology and Psychiatry, Center for Molecular Neuroscience, Vanderbilt University School of Medicine, Nashville, Tennessee 37232-8548, USA.
Blakely
Randy D
RD
D000818:Animals; D001792:Blood Platelets / Q000737:chemistry / Q000378:metabolism*; D002451:Cell Compartmentation; D003598:Cytoskeletal Proteins / Q000032:analysis / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000032:analysis / Q000378:metabolism; D047908:Intracellular Signaling Peptides and Proteins / Q000032:analysis / Q000378:metabolism; D060588:LIM Domain Proteins; D051379:Mice; D011493:Protein Kinase C / Q000032:analysis / Q000378:metabolism; D021381:Protein Transport; D012701:Serotonin / Q000032:analysis / Q000378:metabolism; D050486:Serotonin Plasma Membrane Transport Proteins / Q000032:analysis* / Q000378:metabolism; D013569:Synapses / Q000378:metabolism
D003598:Cytoskeletal Proteins; D004268:DNA-Binding Proteins; D047908:Intracellular Signaling Peptides and Proteins; D060588:LIM Domain Proteins; C493230:SLC6A4 protein, human; D050486:Serotonin Plasma Membrane Transport Proteins; C493231:Slc6a4 protein, mouse; C490351:TGFB1I1 protein, human; C090090:Tgfb1i1 protein, mouse; D012701:Serotonin; D011493:Protein Kinase C