Key Laboratory of Molecular Epigenetics of Ministry of Education, Northeast Normal University, Changchun, Jilin, China.
School of Life Science, Northeast Normal University, Changchun, Jilin, China.
J Immunol. 2022 May 15;208(10):2376-2389. doi: 10.4049/jimmunol.2200002. Epub 2022 Apr 20.
Proinflammatory cytokines/chemokines are commonly regulated by RNA-binding proteins at posttranscriptional levels. Human Ag R (HuR)/embryonic lethal abnormal vision-like 1 (ELAVL1) is one of the well-characterized RNA-binding proteins that increases the stability of short-lived mRNAs, which encode proinflammatory mediators. HuR employs its nucleocytoplasmic shuttling sequence (HNS) domain, interacting with poly(ADP-ribose) polymerase 1 (PARP1), which accounts for the enhanced poly-ADP-ribosylation and cytoplasmic shuttling of HuR. Also by using its HNS domain, HuR undergoes dimerization/oligomerization, underlying the increased binding of HuR with proinflammatory cytokine/chemokine mRNAs and the disassociation of the miRNA-induced silencing complex from the targets. Therefore, competitively blocking the interactions of HuR with its partners may suppress proinflammatory mediator production. In this study, peptides derived from the sequence of the HuR-HNS domain were synthesized, and their effects on interfering HuR interacting with PARP1 and HuR itself were analyzed. Moreover, cell-penetrating TAT-HuR-HNS3 was delivered into human and mouse cells or administered into mouse lungs with or without exposure of TNF-α or LPS. mRNA levels of proinflammatory mediators as well as neutrophil infiltration were evaluated. We showed that TAT-HuR-HNS3 interrupts HuR-PARP1 interaction and therefore results in a lowered poly-ADP-ribosylation level and decreased cytoplasmic distribution of HuR. TAT-HuR-HNS3 also blocks HuR dimerization and promotes Argonaute 2-based miRNA-induced silencing complex binding to the targets. Moreover, TAT-HuR-HNS3 lowers mRNA stability of proinflammatory mediators in TNF-α-treated epithelial cells and macrophages, and it decreases TNF-α-induced inflammatory responses in lungs of experimental animals. Thus, TAT-HuR-HNS3 is a promising lead peptide for the development of inhibitors to treat inflammation-related diseases.
促炎细胞因子/趋化因子通常在转录后水平上受到 RNA 结合蛋白的调节。人 Ag R（HuR）/胚胎致死异常视觉样 1（ELAVL1）是一种经过充分研究的 RNA 结合蛋白，它可以增加短寿命 mRNA 的稳定性，这些 mRNA 编码促炎介质。HuR 利用其核质穿梭序列（HNS）结构域与聚（ADP-核糖）聚合酶 1（PARP1）相互作用，这解释了 HuR 的增强的多聚 ADP-核糖基化和细胞质穿梭。同样，通过使用其 HNS 结构域，HuR 发生二聚化/寡聚化，从而增加了 HuR 与促炎细胞因子/趋化因子 mRNA 的结合，并使 miRNA 诱导的沉默复合物与靶标分离。因此，竞争性阻断 HuR 与其伙伴的相互作用可能会抑制促炎介质的产生。在这项研究中，合成了源自 HuR-HNS 结构域序列的肽，并分析了它们对干扰 HuR 与 PARP1 和 HuR 自身相互作用的影响。此外，将穿透细胞的 TAT-HuR-HNS3 递送至人源和鼠源细胞中，或在 TNF-α或 LPS 暴露的情况下递送至小鼠肺部。评估了促炎介质的 mRNA 水平以及中性粒细胞浸润情况。我们表明，TAT-HuR-HNS3 中断了 HuR-PARP1 相互作用，从而导致聚 ADP-核糖基化水平降低和 HuR 的细胞质分布减少。TAT-HuR-HNS3 还阻断 HuR 二聚化，并促进 Argonaute 2 基于 miRNA 诱导的沉默复合物与靶标的结合。此外，TAT-HuR-HNS3 降低了 TNF-α处理的上皮细胞和巨噬细胞中促炎介质的 mRNA 稳定性，并降低了实验动物肺部的 TNF-α诱导的炎症反应。因此，TAT-HuR-HNS3 是开发用于治疗炎症相关疾病的抑制剂的有前途的先导肽。
