Serotonin Transporter Phosphorylation

描述（由申请人提供）：在多巴胺能突触处，5-羟色胺（5-HT）转运蛋白（SERT）通过将释放的5-HT再摄取到突触前末梢中来决定突触5-HT的浓度和持续时间。因此，突触前质膜中SERT的活性是决定突触能神经传递强度以及行为和生理功能的关键因素。SERT是几种成瘾性和治疗性药物的高亲和力分子靶标，包括可卡因、安非他明、MDMA“摇头丸”、三环类抗抑郁药和SSRIs。在我们之前的资助期间，我们和合作者确定了几种蛋白激酶的急性SERT调节的信号传导机制，并记录了天然和体外模型系统中脂筏介导的胺转运蛋白再分布。然而，SERT磷酸化在5-HT转运调节中的生理相关性，以及负责的位点或结构域还有待研究。该更新申请的主要目标是确定SERT磷酸化在正常5-HT转运调节和与强迫症（OCD）和其他疾病表型的人SERT编码变体相关的失调中的生理作用。具体目标1将检验以下假设：蛋白激酶PKG和p38 MARK在组胺受体（HSR）激活后在特定磷酸化位点/基序上磷酸化SERT并调节5 HT转运。具体目标2将检查hSERT编码变体是否以改变的磷酸化状态存在并且对H3 R/PKG/p38 MARK调节不敏感。具体目标3将测试HSR的体内调节改变SERT磷酸化调节5 HT转运并因此改变突触5 HT水平的假设。本申请中提出的研究将（i）确定影响5-HT转运的生理相关信号/受体;（ii）确定转运蛋白磷酸化和5-HT转运之间的机制联系;（iii）建立负责SERT分布的细胞机制;和（四）提供与hSERT变体相关的失调信息，这些信息可用于开发新的治疗策略，调节SERT功能，治疗精神疾病，如强迫症，抑郁症和药物滥用。

DESCRIPTION (provided by applicant): At serotonergic synapse, serotonin (5-HT) transporters (SERTs) dictate the concentration and duration of synaptic 5-HT by re-uptake of released 5-HT into the presynaptic terminals. Therefore, the activity of SERT in the presynaptic plasma membrane is a critical factor that determines the strength of serotonergic neurotransmission and hence behavioral and physiological functions. SERTs are high-affinity molecular targets for several addictive and therapeutic drugs including cocaine, amphetamine, MDMA "ecstasy", tricyclic antidepressants and SSRIs. In our previous funding period, we, and collaborators identified the signaling mechanisms of acute SERT regulation by several protein kinases, and documented lipid raft-mediated redistribution of amine transporters in native and in vitro model systems. However, the physiological relevance of SERT phosphorylation in 5-HT transport regulation, and the sites or domains responsible have yet to be examined. The major goal of this renewal application is to determine the physiological role of SERT phosphorylation in normal 5-HT transport regulation and in dysregulations associated with human SERT coding variants of obsessive-compulsive disorder (OCD) and other disease phenotypes. Specific Aim 1 will test the hypothesis that protein kinases PKG and p38 MARK phosphorylate SERT on specific phospho-sites/motifs following histamine receptor (HSR) activation and regulate 5HT transport. Specific Aim 2 will examine whether hSERT coding variants exist in altered phosphorylation state and are insensitive to H3R/PKG/p38 MARK modulation. Specific Aim 3 will test the hypotheses that in vivo modulation of HSR alters SERT phosphorylation regulating 5HT transport and hence synaptic 5HT levels. The studies proposed in this application will (i) determine the physiologically relevant signals/receptors influencing 5-HT transport; (ii) define the mechanistic link between transporter phosphorylation and 5-HT transport; (iii) establish the cellular machinery responsible for SERT distribution; and (iv) provide information on dysregulations associated with hSERT variants that could be of use in the development of new therapeutic strategies aimed at regulating SERT function in the treatment of mental illnesses such as OCD, depression and drug abuse.