Role of lysine methyltransferase SMYD2-dependent methylation and identification of new Smyd2 related pathways in pancreatic ductal adenocarcinoma

翻译后修饰可以显着改变蛋白质的性质，从而在一个大的网络中调节细胞过程，因为它已被证明用于调节染色质结构的组蛋白修饰。赖氨酸甲基转移酶SMYD2被认为是甲基化组蛋白，因此可能有助于表观遗传密码。SMYD2还负责甲基化其他细胞蛋白，如肿瘤抑制因子p53和RB。SMYD2在许多人类肿瘤中过表达，其中包括胰腺导管腺癌（PDAC）。PDAC通常诊断得很晚，治愈率很低。我提出的研究项目的目标是了解SMYD2在PDAC中的作用，并确定SMYD2靶点，这可能对肿瘤发生和肿瘤维持有相当大的影响。我将使用一个小鼠模型，其中PDAC是由致癌Kras激活诱导的。Kras在超过90%的PDAC中发生突变。通过对Smyd2基因敲除的分析，探讨Smyd2蛋白对肿瘤发生的影响。这些实验将通过分析SMYD 2耗竭的人胰腺细胞系来补充，以分析对肿瘤维持的影响。由于被SMYD2甲基化的底物还不完全清楚，并且在小鼠中的第一个实验表明Smyd2对肿瘤发生具有独立于p53的作用，因此我将使用两种不同的策略，这将使我能够在体内分析SMYD2靶点。第一种策略是基于甲基化蛋白的富集，其将根据表达SMYD2或SMYD2的催化失活形式的细胞而被差异标记。第二种策略是基于工程SMYD2，它专门使用SAM类似物，并将炔基而不是甲基转移到底物上。炔基将用于分离SMYD2的特定底物。将检测鉴定的靶标的致癌或肿瘤抑制功能。了解SMYD2的作用并确定SMYD2的新底物，这是PDAC肿瘤发生和维持的关键因素，将阐明对胰腺肿瘤细胞至关重要的基本机制，并将确定新的途径作为PDAC和其他相关癌症类型治疗和诊断的新起点。

Post-translational modifications can change the properties of proteins significantly and thus regulate cellular processes in a large network, as it has been shown for histone modifications regulating chromatin structure. The lysine methyltransferase SMYD2 is thought to methylate histones and might therefore contribute to the epigenetic code. SMYD2 is also responsible for methylating other cellular proteins, such as the tumor suppressors p53 and RB. SMYD2 is overexpressed in many human tumors, among these pancreatic ductal adenocarcinoma (PDAC). PDAC is often diagnosed very late and curability is very low. The goal of my proposed research project is to understand the role of SMYD2 in PDAC and to identify SMYD2 targets, which may have considerable effect on tumorigenesis and tumor maintenance. I will use a mouse model, in which PDAC is induced by activation of oncogenic Kras. Kras is mutated in more than 90% of all PDACs. Analysing the knockout of Smyd2 in this system, I will test the influence of the mouse Smyd2 protein on tumorigenesis. These experiments will be complemented by the analysis of human pancreatic cell lines depleted for SMYD2 to analyse the effect on tumor maintenance. Since the substrates that are methylated by SMYD2 are not yet completely known and first experiments in mice show that Smyd2 has effects on tumorigenesis independent on p53, I will use two different strategies, which will allow me to analyse SMYD2 targets in vivo. The first strategy is based on the enrichment of methylated proteins, which will be differentially labelled dependent on cells expressing SMYD2or a catalytically inactive form of SMYD2. The second strategy is based on an engineered SMYD2, which specifically uses a SAM analog and will transfer an alkyne group instead of a methylgroup to the substrate. The alkyne group will be used to isolate the specific substrates of SMYD2. The identified targets will be tested for their oncogenic or tumor suppressive function. Understanding the role of SMYD2 and identifying new substrates of SMYD2, which are key players in PDAC tumorigenesis and maintenance, will illuminate basic mechanisms that are crucial for pancreatic tumor cells and will identify novel pathways as a new starting point for the therapy and diagnosis of PDAC and other related cancer types.