Key Points Annotated lncRNA 1810058I24Rik encodes a conserved mitochondrial micropeptide. Macrophages fail to activate the Nlrp3 inflammasome in the absence of Mm47. Genetic deletion of Mm47 in macrophages affects only Nlrp3 and not Aim2 or Nlrc4. Functional peptides encoded by short open reading frames are emerging as important mediators of fundamental biological processes. In this study, we identified a micropeptide produced from a putative long noncoding RNA (lncRNAs) that is important in controlling innate immunity. By studying lncRNAs in mice macrophages, we identified lncRNA 1810058I24Rik, which was downregulated in both human and murine myeloid cells exposed to LPS as well as other TLR ligands and inflammatory cytokines. Analysis of lncRNA 1810058I24Rik subcellular localization revealed that this transcript was localized in the cytosol, prompting us to evaluate its coding potential. In vitro translation with 35S-labeled methionine resulted in translation of a 47 aa micropeptide. Microscopy and subcellular fractionation studies in macrophages demonstrated endogenous expression of this peptide on the mitochondrion. We thus named this gene mitochondrial micropeptide-47 (Mm47). Crispr–Cas9–mediated deletion of Mm47, as well as small interfering RNA studies in mice primary macrophages, showed that the transcriptional response downstream of TLR4 was intact in cells lacking Mm47. In contrast, Mm47-deficient or knockdown cells were compromised for Nlrp3 inflammasome responses. Activation of Nlrc4 or Aim2 inflammasomes were intact in cells lacking Mm47. This study therefore identifies, to our knowledge, a novel mitochondrial micropeptide Mm47 that is required for the activation of the Nlrp3 inflammasome. This work further highlights the functional activity of short open reading frame–encoded peptides and underscores their importance in innate immunity.

中文翻译：

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激活 Nlrp3 炎性体需要线粒体微肽

Key Points Annotated lncRNA 1810058I24Rik 编码一个保守的线粒体微肽。在没有 Mm47 的情况下，巨噬细胞无法激活 Nlrp3 炎症小体。巨噬细胞中 Mm47 的基因缺失仅影响 Nlrp3，而不影响 Aim2 或 Nlrc4。由短的开放阅读框编码的功能性肽正在成为基本生物过程的重要介质。在这项研究中，我们鉴定了一种由推定的长链非编码 RNA (lncRNA) 产生的微肽，该微肽对控制先天免疫很重要。通过研究小鼠巨噬细胞中的 lncRNA，我们确定了 lncRNA 1810058I24Rik，它在暴露于 LPS 以及其他 TLR 配体和炎性细胞因子的人和鼠骨髓细胞中均下调。lncRNA 1810058I24Rik 亚细胞定位的分析表明，该转录物定位于细胞质中，促使我们评估其编码潜力。用 35S 标记的甲硫氨酸进行体外翻译导致翻译了 47 个氨基酸的微肽。巨噬细胞中的显微镜和亚细胞分离研究证明了该肽在线粒体上的内源性表达。因此，我们将该基因命名为线粒体微肽-47 (Mm47)。Crispr-Cas9 介导的 Mm47 缺失，以及小鼠原代巨噬细胞中的小干扰 RNA 研究表明，在缺乏 Mm47 的细胞中，TLR4 下游的转录反应是完整的。相比之下，Mm47 缺陷或敲低细胞受到 Nlrp3 炎症小体反应的影响。Nlrc4 或 Aim2 炎症小体的激活在缺乏 Mm47 的细胞中是完整的。因此，本研究确定，据我们所知，一种新的线粒体微肽 Mm47，它是 Nlrp3 炎症小体激活所必需的。这项工作进一步突出了短开放阅读框编码肽的功能活性，并强调了它们在先天免疫中的重要性。