Impact of HIV-1 Tat protein on cocaine-dopamine transporter interaction

HIV-1 Tat 蛋白对可卡因-多巴胺转运蛋白相互作用的影响

• 批准号: 8690005
• 负责人: Jun Zhu
• 金额: $ 36.62万
• 依托单位: UNIVERSITY OF SOUTH CAROLINA AT COLUMBIA
• 依托单位国家: 美国
• 财政年份: 2013
• 资助国家: 美国
• 起止时间: 2013-07-01 至 2018-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8690005
• 关键词: Address; Affect; Amino Acids; Anti-Retroviral Agents; Binding; Binding Sites; Biological Preservation; Brain; Brain region; Cells; Chinese Hamster Ovary Cell; Cocaine; Computer Simulation; Corpus striatum structure; DNA; Data; Development; Disease; Dopamine; Exposure to; Functional disorder; HIV-1; Human; Impairment; Incidence; Individual; Infection; Lead; Lentivirus Vector; Ligand Binding; Link; Mediating; Molecular; Molecular Target; Mutation; Neurocognitive; Neurons; Neurotransmitters; PC12 Cells; Patients; Physiological; Play; Prevalence; Process; Production; Publishing; Rattus; Recording of previous events; Role; Severities; Site; Site-Directed Mutagenesis; Staging; Structure; Structure-Activity Relationship; Synapses; System; Techniques; Testing; Therapeutic; Transfection; Viral; Viral Proteins; Virus; antiretroviral therapy; attenuation; base; brain cell; conformational conversion; dopamine system; dopamine transporter; drug of abuse; improved; insight; molecular dynamics; mutant; neurotransmission; presynaptic; prevent; psychostimulant; public health relevance; research study; tat Protein; uptake

DESCRIPTION (provided by applicant): HIV-1 associated neurocognitive disorders (HAND) remain highly prevalent in the era of effective antiretroviral therapy. HIV-1 infection within CNS system plays a central role in the development of HAND. Drugs of abuse, such as cocaine, have been shown to increase the incidence and exacerbate the severity of HAND by enhancing viral replication. However, the mechanistic links between cocaine and HAND progression remain undefined. Although changes in many neurotransmitter systems may contribute to HAND, the central dopamine (DA) system plays a crucial role in the development of neurocognitive dysfunction in HAND patients and in the control of psychostimulant action of cocaine. The interplay of HIV-1 Tat protein with cocaine augments synaptic DA level and Tat release within dopaminergic brain regions. Long lasting exposure to elevated DA and Tat eventually lead to DA deficit that potentiates severity and accelerates progression of HAND. Antiretroviral agents cannot prevent the production of HIV-1 viral proteins, such as Tat protein, in proviral-containing brain cells. It is unclear how the DA system is altered in HIV-1 positive cocaine abusers. Therefore, there is a pressing need to define the molecular mechanism(s) by which the impaired DA system by HIV-1 infection affects the progression of HAND in concurrent cocaine abusers. Presynaptic DA transporter (DAT), which is critical for neurocognitive function, is a major molecular target for both Tat and cocaine to impact the DA system. In this application, we hypothesize that Tat, via allosteric binding sites in the DAT, potentiates inhibitory effects of cocaine on DA transport, which is the key to DA system dysfunction occurred in HAND patients. Our proposed experiments will investigate how Tat and cocaine interact with the human DAT through their recognition binding sites on human DAT, thereby leading to dysfunction of the DA system. Our strategy encompasses creating a dynamic 3D computational model to predict potential Tat and cocaine binding pocket residues of human DAT, validating these residues via site-directed mutagenesis, and analyzing the consequent functional changes of the Tat and DAT interaction in neuronal cells and primary neurons. The completion of this application will identify molecular targets on the DAT for developing compounds that specifically block Tat binding site(s) in DAT and stabilize physiological dopaminergic tone, which should be beneficial to the preservation of neurocognitive function in patients with HAND in concurrent cocaine abusers.

描述（由申请人提供）：在有效的抗逆转录病毒治疗时代，HIV-1 相关的神经认知障碍 (HAND) 仍然非常普遍。 CNS 系统内的 HIV-1 感染在 HAND 的发生中发挥着核心作用。滥用药物（如可卡因）已被证明会通过增强病毒复制来增加 HAND 的发病率并加剧其严重程度。然而，可卡因与 HAND 进展之间的机制联系仍不清楚。尽管许多神经递质系统的变化可能导致 HAND，但中枢多巴胺 (DA) 系统在 HAND 患者神经认知功能障碍的发展以及可卡因精神兴奋作用的控制中起着至关重要的作用。 HIV-1 Tat 蛋白与可卡因的相互作用增强了多巴胺能大脑区域内的突触 DA 水平和 Tat 释放。长期持续暴露于升高的 DA 和 Tat 最终会导致 DA 缺乏，从而加剧 HAND 的严重程度并加速其进展。抗逆转录病毒药物不能阻止含有前病毒的脑细胞中产生 HIV-1 病毒蛋白，例如 Tat 蛋白。目前尚不清楚 HIV-1 阳性可卡因滥用者的 DA 系统是如何改变的。因此，迫切需要确定 HIV-1 感染导致的 DA 系统受损影响并发可卡因滥用者 HAND 进展的分子机制。突触前 DA 转运蛋白 (DAT) 对神经认知功能至关重要，是 Tat 和可卡因影响 DA 系统的主要分子靶点。在此应用中，我们假设 Tat 通过 DAT 中的变构结合位点，增强了以下物质的抑制作用： 可卡因对DA转运的影响，这是HAND患者发生DA系统功能障碍的关键。我们提出的实验将研究 Tat 和可卡因如何通过人类 DAT 上的识别结合位点与人类 DAT 相互作用，从而导致 DA 系统功能障碍。我们的策略包括创建动态 3D 计算模型来预测人类 DAT 的潜在 Tat 和可卡因结合口袋残基，通过定点诱变验证这些残基，并分析神经元细胞和原代神经元中 Tat 和 DAT 相互作用的后续功能变化。该申请的完成将确定 DAT 上的分子靶标，以开发专门阻断 DAT 中 Tat 结合位点并稳定生理多巴胺能张力的化合物，这应该有利于保留可卡因滥用者中的 HAND 患者的神经认知功能。