Uridylation of miRNAs by ZCCHC6 Regulates IL-6 Expression in Arthritis

 描述(申请人提供)：据估计，美国有2700万25岁及以上的人患有骨关节炎(OA)，这是一种关节软骨的退行性疾病，其特征是软骨基质丢失，主要是胶原和蛋白多糖，导致关节组织破坏和功能丧失。在45岁之前，患骨性关节炎的男性比女性多；45岁以后，女性更常见。促炎细胞因子IL-6最近引起了人们的兴趣，因为它在骨性关节炎患者的滑液和血清中水平升高，并具有 上调基质金属蛋白酶13的表达，抑制II型胶原的表达。成熟的miRNAs是一种非编码RNA，它抑制靶mRNAs的翻译和稳定性，现在已知的是在3‘端(3’NTA)上添加非模板核苷酸(主要是尿苷)。负责miRNAs 3‘端修饰的酶最近才在癌细胞和免疫细胞中被发现，但尚未在软骨或软骨细胞中被描述。我们的初步数据表明，(1)ZCCHC6-一种核苷酸转移酶-在受损的骨性关节炎软骨中高表达；(2)调节人软骨细胞中IL-6的表达。已知ZCCHC6可尿苷化mRNAs，因此提示ZCCHC6介导的miRNA3‘端修饰可能在骨关节炎中IL-6的转录后调节中发挥重要作用，并可能参与其他炎症介质的转录后调节。我们将追求4个特定的目标来验证基本假设，即ZCCHC6尿苷酸化软骨细胞中的成熟miRNAs来调节IL-6的表达和OA的炎症。具体目标--1.研究将检验这一假设，即“以miRNAs为靶标的细胞因子在骨关节炎软骨中被尿苷基化”。具体目标--2.我们将检验“miRNA尿化可抑制人骨关节炎软骨细胞中IL-6的表达”这一假设。具体目标--3.我们将验证“ZCCHC6的表达对人骨关节炎软骨细胞中IL-6的表达是必不可少的”这一假设。具体目标--4.在人类骨性关节炎的动物模型中，我们将检验这一假设，即ZCCHC6在疾病的诱导和发展过程中介导miRNA尿苷化和调节IL-6的表达。从这些研究中获得的知识将是重要的，因为通过了解miRNA3‘末端修饰如何影响骨关节炎中炎症介质的表达，可能会导致有效治疗骨关节炎的新生物标记物和疗法的开发。

 DESCRIPTION (provided by applicant): Abstract An estimated 27 million Americans age 25 and older have osteoarthritis (OA), a degenerative disease of articular cartilage characterized by loss of the cartilage matrix, mainly collagen and proteoglycans, leading to joint tissue destruction and loss of function. Before age 45, more men than women have osteoarthritis; after age 45, it is more common in women. The pro-inflammatory cytokine IL-6 has recently gained interest due to its elevated levels in synovial fluid and sera from OA patients and its capacity to upregulate the expression of MMP-13 and inhibit the expression of type-II collagen. Mature miRNAs are non-coding RNAs that inhibit the translation and stability of target mRNAs and are now known to undergo addition of non-template nucleotides (largely uridines) to the 3' ends (3'NTAs). The enzymes responsible for 3' end modifications of miRNAs have only recently been identified in cancer and immune cells but have not yet been described in cartilage or chondrocytes. Our preliminary data show that (1) ZCCHC6-a nucleotidyl transferase-is highly expressed in damaged OA cartilage; and (2) regulates IL-6 expression in human chondrocytes. ZCCHC6 is known to uridylate mRNAs thus suggesting that ZCCHC6-mediated miRNA 3'end modifications could play a significant role in the post-transcriptional regulation of IL-6, and possibly of other inflammatory mediators, in OA. We will pursue 4 specific aims to test the fundamental hypothesis that ZCCHC6 uridylates mature miRNAs in chondrocytes to regulate IL-6 expression and inflammation in OA. Specific Aim-1. Studies will test the hypothesis that "cytokine targeting miRNAs are uridylated in OA cartilage". Specific Aim-2. We will test the hypothesis that "miRNA uridylation abrogates repression of IL-6 expression in human OA chondrocytes". Specific Aim-3. We will test the hypothesis that "expression of ZCCHC6 is essential for IL-6 expression in human OA chondrocytes". Specific Aim-4. In an animal model of human OA we will test the hypothesis that "ZCCHC6 mediates miRNA uridylation and regulate IL-6 expression during the induction and progression of the disease". Knowledge gained from these studies will be important since by understanding how miRNA 3'end modifications affect the expression of inflammatory mediators in OA may lead to the development of novel biomarkers and therapies for the effective treatment of OA.