Modulation of OPRM1 alternative splicing by morphine and HIV-1 Nef

吗啡和 HIV-1 Nef 对 OPRM1 选择性剪接的调节

• 批准号: 10654016
• 负责人: SULIE L. CHANG
• 金额: $ 34.54万
• 依托单位: TEMPLE UNIV OF THE COMMONWEALTH
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-09-15 至 2026-06-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10654016
• 关键词: Alternative Splicing; Antisense Oligonucleotides; Autopsy; Behavior; Binding; Biochemical; Brain; Brain region; C-terminal; Cell Culture Techniques; Cells; Chronic; Clinical; Consensus; Cyclic AMP; Data; Development; Drug abuse; Elements; Event; Exons; Exposure to; Family; Functional disorder; G Protein-Coupled Receptor Signaling; G-Protein-Coupled Receptors; Gene Activation; HIV; HIV Envelope Protein gp120; HIV Infections; HIV-1; HIV/AIDS; Heroin; Human; Illicit Drugs; In Vitro; Inbred F344 Rats; Incidence; Individual; Mediating; Messenger RNA; Modeling; Molecular; Morphine; Morphine Dependence; Muscarinic M1 Receptor; N-terminal; Neurocognitive; Neuroglia; Neurons; Opiate Addiction; Opioid Receptor; Oral; Pathologic; Pathway interactions; Peripheral Blood Mononuclear Cell; Persons; Phosphotransferases; Physical Dependence; Physical assessment; Physiological; Protein Isoforms; Protein Kinase; RNA; RNA Splicing; Rattus; Recombinants; Regulation; Reporting; Research; Rewards; Role; Self Administration; Signal Pathway; Signal Transduction; Spinal Cord; Tail; Tissues; Transcript; Viral; Withdrawal; brain tissue; exosome; extracellular vesicles; genetic regulatory protein; in vivo; mRNA Precursor; member; nef Protein; novel; opioid abuse; opioid use; response; secretory protein

PROJECT SUMMARY This R01 application is in response to PAS-18-915 entitled “HIV/AIDS High Priority Drug Abuse Research”. Our project will investigate the molecular and functional interactions between HIV and morphine in regulation of alternative pre-mRNA splicing of the opioid receptor M1 (OPRM1) with implications for enhanced opioid dependence observed in the people with HIV (PWH). Clinically used opioids, such as morphine, as well as illicit drugs, such as heroin, activate OPRM1 that is a member of the G protein-coupled receptor (GPCR) family. OPRM1 pre-mRNA undergoes extensive alternative splicing events. To date, 21 isoforms of the human OPRM1 with alternative C-terminal and/or N-terminal regions and 17 isoforms of the rat OPRM1, have been identified. Given the importance of these regions in G protein-coupled receptor (GPCR) signaling, differential regulation of OPRM1 isoforms would have functional consequences. However, characterization of OPRM1 signaling is generalized, and only one isoform (MOR1) has been extensively studied. Our preliminary data suggest that expression of splicing regulatory protein SRSF1 and alternative splicing of MOR-1X is preferentially induced in neuronal cells exposed to morphine. Interestingly, our results also revealed that alternative splicing and expression of MOR-1X isoform is induced in postmortem brain tissues obtained from the PWH. These results suggested that HIV and morphine may impact OPRM1 alternative splicing and synergistically induce MOR-1X isoform expression. The mutually exclusive exon X of OPRM1 pre-mRNA is incorporated into the mature mRNA transcript following exon 3 in the MOR-1X mRNA transcript. This insertion results in substantially longer C-terminal tail having new motifs potentially binding with several cellular kinases that is unique to the MOR-1X isoform. We recently reported that glial cells infected with HIV-1 release Nef protein captured within the extracellular vesicles (Nef-EVs) which are readily taken up by neurons. We further assessed possible role of Nef-EVs and two other HIV secretory proteins, Tat and gp120, in alternative splicing of MOR-1X. Interestingly, while recombinant Tat and gp120 had no visible effects, treatment of neurons with Nef-EVs caused a comparable induction in MOR-1X alternative splicing as did treatment with morphine. Our preliminary results also revealed that co-treatment of neurons with Nef-EVs and morphine synergistically induced MOR-1X alternative splicing. Additionally, alternative splicing of MOR-1X was induced in brain regions involved in the reward pathways of the F344 rat that is the control strain of HIV-1Tg rat that has an additive induction with the exposure to morphine. Taken all together, we hypothesize that HIV-1 contributes to the increased rate of opioid dependence in the PWH by amplifying the rate of MOR-1X alternative splicing induced by morphine. Our proposed studies will reveal a novel synergistic interaction between morphine and HIV on alternative splicing of OPRM1 pre-mRNA leading to preferential expression of MOR-1X isoform with implications in physical dependence of morphine.

项目摘要 此R01的应用是对PAS-18-915的响应，标题为“ HIV/AIDS高优先药物滥用研究”。 我们的项目将调查HIV和吗啡之间的分子和功能相互作用。 阿片受体M1（OPRM1）的替代性mRNA剪接，对增强阿片类药物的影响 在艾滋病毒（PWH）患者中观察到的依赖性。临床使用的阿片类药物，例如吗啡以及 非法药物（例如海洛因）激活G蛋白偶联受体（GPCR）的成员OPRM1 家庭。 OPRM1前MRNA经历了广泛的替代剪接事件。迄今为止，人类的21种同工型 具有替代C末端和/或N末端区域的OPRM1以及大鼠OPRM1的17种同工型 确定。鉴于这些区域在G蛋白偶联受体（GPCR）信号中的重要性，差异 OPRM1同工型的调节将产生功能后果。但是，OPRM1的表征 信号传导已广泛化，只有一种同工型（MOR1）被广泛研究。我们的初步数据 表明剪接调节蛋白SRSF1和MOR-1X的替代剪接的表达是 有趣的是，我们的结果也表明 MOR-1X同工型的替代剪接和表达是在验尸脑组织中诱导的 PWH。这些结果表明，艾滋病毒和吗啡可能会影响oprm1替代剪接和 协同诱导MOR-1X同工型表达。 OPRM1前MRNA的互斥外显子X是 在MOR-1X mRNA转录本中外显子3后，掺入成熟的mRNA转录本中。这个插入 导致与几个细胞激酶可能结合的新基序的较长的C末端尾巴 这是MOR-1X同工型独有的。我们最近报道说，感染了HIV-1释放NEF的神经胶质细胞 在细胞外蔬菜（NEF-EV）中捕获的蛋白质很容易被神经元吸收。我们进一步 评估了NEF-EV和其他两种HIV秘密蛋白TAT和GP120的可能作用 MOR-1X。有趣的是，尽管重组TAT和GP120没有明显影响，但对神经元的治疗 NEF-EVS与吗啡治疗一样，引起MOR-1X替代剪接的可比诱导。我们的 初步结果还表明，神经元与NEF-EVS和吗啡的共同治疗协同作用 诱导MOR-1X替代剪接。此外，在大脑区域诱导了MOR-1X的替代剪接 参与F344大鼠的奖励途径，这是HIV-1TG大鼠的控制菌株，具有加性 接触吗啡的诱导。一起，我们假设HIV-1有助于 通过扩增MOR-1X替代剪接的速率，PWH中阿片类药物的依赖性率提高了 由吗啡。我们提出的研究将揭示吗啡与HIV之间的新型协同相互作用 OPRM1前MRNA的替代剪接，导致MOR-1X同工型的首选表达 吗啡的物理依赖性。

项目成果

iPSC-derived three-dimensional brain organoid models and neurotropic viral infections.

• DOI: 10.1007/s13365-023-01133-3
• 发表时间: 2023-04
• 期刊: JOURNAL OF NEUROVIROLOGY
• 影响因子: 3.2
• 作者: Swingler, Michael;Donadoni, Martina;Bellizzi, Anna;Cakir, Senem;Sariyer, Ilker K.
• 通讯作者: Sariyer, Ilker K.

Modulation of OPRM1 Alternative Splicing by Morphine and HIV-1 Nef.

• DOI: 10.1007/s11481-021-10009-4
• 发表时间: 2022-06
• 期刊: Journal of neuroimmune pharmacology : the official journal of the Society on NeuroImmune Pharmacology
• 作者: Donadoni M;Huang W;Yarandi SS;Burdo TH;Chang SL;Sariyer IK
• 通讯作者: Sariyer IK