PML- and Smyd1-dependent regulatory mechanisms and effects on inflammatory responses of vascular endothelial cells

PML和Smyd1依赖性调节机制及其对血管内皮细胞炎症反应的影响

• 批准号: 418689898
• 负责人: Dr. Janine Berkholz
• 金额: --
• 依托单位: Institut für Physiologie (CCM)
• 依托单位国家: 德国
• 项目类别: Research Grants
• 财政年份: 2019
• 资助国家: 德国
• 起止时间: 2018-12-31 至 2022-12-31
• 项目状态: 已结题
• 来源: https://gepris.dfg.de/gepris/projekt/418689898?language=en
• 关键词: PML; Smyd1; dependent; regulatory; mechanisms

Endothelial cells (ECs) line as a single layer the interior surface of lymphatic and blood vessels. Loss of proper endothelial functions lead to potentially life-threatening conditions. They may emerge as a chronic inflammatory disease with specific anatomical localization, as in the case of arteriosclerosis, but can also arise acute, as in the case of a sepsis or of an ischemic-reperfusion injury, which in contrast to sepsis is typically restricted to single organs. In these, per se very different situations, inflammatory alterations of the endothelium play a pivotal role for the functional integrity of the endothelium of affected blood vessels. Preliminary experiments of the proposed project revealed a protein-protein-interaction of Smyd1 (SET And MYND Domain Containing 1) and PML (Promyelocytic Leukemia Protein) and a PML-dependent SUMOylation of Symd1 in ECs. Overexpression of Smyd1 in ECs induced by gene transfection showed increased expression levels of PML and NFB. Remarkably, the effect of Smyd1 on NFB expression was caused by the histone methyltransferase activity of Smyd1. Overexpression of both, Smyd1 and PML, resulted in an increased expression of the major histocompatibility complex class II transactivator CIITA. An increased endothelial MHC class II expression is associated with autoimmune diseasesand endothelial dysfunction. The expression of PML and Smyd1 in ECs was augmented by treatment with liposaccharide. Hypoxia showed a similar effect. In contrast, atherogenic flow profiles decreased PML and Smyd1 expression. These findings provided experimental evidence that PML and Smyd1 interact with each other to influence inflammatory responses of vascular endothelial cells.In the current proposal, we want to explore in more detail the role of PML and Smyd1 in inflammatory reactions of vascular endothelial cells, blood vessels and organ perfusion. Special stimulation protocols will be applied to mimic inflammatory reactions characteristic for the initiation of arteriosclerosis, sepsis and ischemic-reperfusion injury. The systematic comparison of these stimulation protocols ought to deepen the understanding for varieties and similarities in the investigated endothelial dysfunctions. The underlying cellular mechanisms, which include transcriptional regulation of putative target genes via Smyd1-dependent histone modifications, PML nuclear bodies-mediated SUMOylation of proteins, as well as interaction of both proteins, will be identified and characterized in detail. Implications for endothelial cell function will be experimentally quantified. Finally, translational applications will be prepared by analysis of the obtained findings, the development of Smyd1-inhibitors including the comparison with cardiovascular databases.

内皮细胞（EC）作为单层排列在淋巴管和血管的内表面。适当内皮功能的丧失导致潜在的危及生命的状况。它们可以作为具有特定解剖学定位的慢性炎性疾病出现，如在动脉硬化的情况下，但也可以急性出现，如在脓毒症或缺血-再灌注损伤的情况下，与脓毒症相反，其通常限于单个器官。在这些本身非常不同的情况下，内皮的炎性改变对于受影响血管的内皮的功能完整性起关键作用。该项目的初步实验揭示了Smyd 1（SET和MYND结构域包含1）和PML（早幼粒细胞白血病蛋白）的蛋白质-蛋白质相互作用以及EC中Symd 1的PML依赖性SUMO化。转染Smyd 1基因后，内皮细胞PML和NF κ B B的表达明显增加。值得注意的是，Smyd 1对NF κ B B表达的影响是由Smyd 1的组蛋白甲基转移酶活性引起的。Smyd 1和PML的过度表达导致主要组织相容性复合体II类反式激活因子CIITA的表达增加。内皮MHCII类分子表达增加与自身免疫性疾病和内皮功能障碍有关。脂质体处理可增加内皮细胞PML和Smyd 1的表达。缺氧也有类似的效果。相比之下，致动脉粥样硬化的血流分布降低了PML和Smyd 1的表达。本研究为PML和Smyd 1相互作用影响血管内皮细胞的炎症反应提供了实验证据，我们希望更详细地探讨PML和Smyd 1在血管内皮细胞炎症反应、血管和器官灌注中的作用。将应用特殊的刺激方案来模拟动脉硬化、脓毒症和缺血再灌注损伤起始的炎症反应特征。这些刺激方案的系统比较应该加深对所研究的内皮功能障碍的多样性和相似性的理解。潜在的细胞机制，其中包括通过Smyd 1依赖性组蛋白修饰，PML核小体介导的蛋白质SUMO化，以及这两种蛋白质的相互作用，推定的靶基因的转录调控，将被确定和详细表征。内皮细胞功能的影响将通过实验进行量化。最后，翻译申请将通过分析所获得的结果，Smyd 1抑制剂的开发，包括与心血管数据库的比较来准备。