Essential role of the Ufm1 E3 ligase in intestinal homeostasis

项目总结： 肠道动态平衡对正常的肠道功能至关重要，而动态平衡的破坏可能导致发病 人类疾病，如慢性炎症，甚至癌症。潘氏细胞在肠道中发挥重要作用 屏障功能与粘膜免疫。最近的研究表明，Paneth的功能受损 细胞可能导致肠道菌群失调，增加炎症性肠病(IBD)的易感性。这个 Ufm1(泛素折叠修饰物1)结合系统是一种新型的类泛素系统，它由Ufm1- 特异的E1酶(Uba5)、E2酶(Ufc1)和可能的E3酶(包含Ufl1的蛋白质复合体) 和Ufbp1)。临床和动物模型研究表明，它可能与许多疾病有关，如 神经功能障碍、缺血性损伤和癌症，但其总体功能和潜在机制仍然 人们对此知之甚少。来自其他人和我们的基因研究最近证明了这一点的重要作用 该系统在胚胎发育和造血中的作用尚不明确，但在其他系统中的作用尚不明确。Ufl1 和Ufbp1是Ufm1 E3连接酶的关键成分，存在于多种组织器官中，并高度 在肠道中表达，但它们在肠道中的作用完全未知。频发肠胃的观察 Ufl1和Ufbp1基因敲除小鼠的出血促使我们产生IEC(肠上皮细胞)特异性 Ufl1和Ufbp1基因敲除小鼠模型。我们发现，IEC特异性的Ufl1或Ufbp1消融导致 潘氏细胞几乎完全丧失，杯状细胞部分丧失，导致对 实验性结肠炎。此外，Ufbp1消融导致内质网应激增加，激活 未折叠蛋白反应(UPR)和细胞凋亡。我们还观察到mRNAs的急剧增加 在急性Ufbp1消融过程中，如溶菌酶等分泌蛋白的变化。我们假设Ufbp1 而超甲基化途径在IRE1介导的ER相关mRNAs降解中起着关键作用， 从而协调蛋白质合成和内质网蛋白负荷，维持潘氏细胞内质网稳态。 Ufbp1缺乏削弱了这种降解机制，导致mRNAs和蛋白质翻译增加， 导致内质网压力升高和潘氏细胞死亡。为了检验这一假设，我们提出了三个具体目标。 目的1：确定Ufbp1在Paneth细胞中的特异性功能；目的2：探讨UPR在Paneth细胞中的作用 目的3：阐明Ufbp1消融诱导细胞凋亡的机制 内质网动态平衡的破坏。这个项目的结果将提供对生理学的关键洞察 这一重要的蛋白质修饰系统在维持肠道功能中的作用和作用机制 动态平衡，并对我们理解潘氏细胞生物学，发展 炎症性疾病的新治疗靶点及其ufmyl化途径的生理功能 以及它与普遍定期审议的相互作用。

Project Summary: Intestinal homeostasis is crucial for normal intestinal functions, and its disruption may lead to the pathogenesis of human diseases such as chronic inflammation and even cancer. Paneth cells play crucial roles in intestinal barrier function and mucosal immunity. Recent studies have demonstrated that impaired function of Paneth cells may lead to dysbiotic intestine and increased susceptibility to inflammatory bowel disease (IBD). The Ufm1 (Ubiquitin-fold modifier 1) conjugation system is a novel ubiquitin-like system that consists of Ufm1- sepcific E1 enzyme (Uba5), E2 enzyme (Ufc1) and a putative E3 enzyme (a protein complex containing Ufl1 and Ufbp1). Clinical and animal model studies implicate its potential involvement in many diseases such as neurological disorders, ischemic injury and cancer, but its overall functions and underlying mechanisms remain poorly understood. Genetic studies from others and us have recently demonstrated the essential role of this system in embryonic development and hematopoiesis, but its role in other systems remains to be defined. Ufl1 and Ufbp1, the key components of Ufm1 E3 ligase, are present in multiple tissues and organs and highly expressed in the intestine, yet their role in the gut is completely unknown. The observation of frequent gut bleeding in Ufl1 and Ufbp1 knockout mice prompted us to generate IEC (intestinal epithelial cell)-specific knockout mouse models of Ufl1 and Ufbp1. We found that IEC-specific ablation of either Ufl1 or Ufbp1 caused nearly complete loss of Paneth cells and partial loss of goblet cells, resulting in increased susceptibility to experimental colitis. Moreover, Ufbp1 ablation led to elevated ER (endoplasmic reticulum) stress, activation of the Unfolded Protein Response (UPR) and apoptosis in IECs. We also observed dramatic increase of mRNAs of secretory proteins such as lysozyme during the course of acute Ufbp1 ablation. We hypothesize that Ufbp1 and the ufmylation pathway plays a pivotal role in IRE1-mediated degradation of ER-associated mRNAs, thereby coordinating protein synthesis and ER protein load and maintaining ER homeostasis of Paneth cells. Ufbp1 deficiency impairs this degradation mechanism and leads to increased mRNAs and protein translation, resulting in elevated ER stress and Paneth cell death. To test this hypothesis, we propose three specific aims. Aim 1: To define Paneth cell-specific function of Ufbp1; Aim 2: To investigate the role of UPR in Paneth cell apoptosis induced by Ufbp1 deficiency; and Aim 3: To elucidate the mechanism of Ufbp1 ablation-induced disruption of ER homeostasis. The outcome of this project will provide critical insight into the physiological function and working mechanism of this important protein modification system in maintaining intestinal homeostasis., and have a significant impact on our understanding of Paneth cell biology, development of novel therapeutic targets for inflammatory disease, and the physiological function of the ufmylation pathway and its interplay with UPR.