Structure and Function of JmjC Histone Demethylases

描述（由申请人提供）：组蛋白的共价修饰，真核细胞中基因活性的基本调节剂，通过各种酶促反应重塑染色质结构。一些修饰的可逆过程，如乙酰化和去乙酰化，被很好地表征。然而，甲基化和去甲基化是否可逆地促进基因调控仍然存在争议。最近的研究表明，甲基化和去甲基化被普遍用于翻译后修饰组蛋白以调节基因活性。除了LSD1（一种核胺氧化酶同系物）被发现具有组蛋白赖氨酸去甲基化酶的功能外，我们和其他人还发现一些含有JmjC结构域的蛋白是组蛋白去甲基化酶。具体来说，我们发现JMJD2蛋白家族的成员是组蛋白去甲基酶，作用于H3-K9和H3-K36的三甲基。此外，该家族的一些成员对二甲基也有活性。为了了解这些蛋白的结构和功能之间的关系，我们确定了JMJD2A蛋白的催化核心结构。该结构揭示了一些新的结构特征，如新的JmjN结构域、JmjC结构域、c端结构域和锌指基序。这些独特的结构特征创造了一个潜在的催化中心。该结构还揭示了一个特征基元，其中包括辅助因子的结构决定因素，如Fe（II）和a-酮戊二酸，这是非含血红素加氧酶的标志。然而，仍有一些主要问题有待解决，如催化核心识别底物肽的结构基础、甲基特异性的结构决定因素、调节酶活性的机制以及不同含JmjC结构域组蛋白去甲基化酶之间的关系。此外，只有一小部分含有JmjC结构域的蛋白质被表征。我们认为在含有JmjC结构域的蛋白家族中存在未表征的组蛋白去甲基化酶。来自JMJD2A的结构信息应该有助于揭示这些新的组蛋白去甲基化酶。为了回答这些问题并理解含JmjC结构域的组蛋白去甲基化酶如何起作用，提出了三个具体目标。具体目标1 -表征JMJD2家族成员的结构和功能，并鉴定新的组蛋白去甲基化酶；特定目的2 -确定不同底物多肽存在下JMJD2A催化核的高分辨率结构；具体目标3 -确定整个JMJD2A蛋白和其他含JmjC结构域蛋白的催化核心的高分辨率结构，并揭示调节JMJD2A和其他相关蛋白活性的机制。

DESCRIPTION (provided by applicant): Covalent modifications of histone proteins, essential regulators of the activity of genes in eukaryotic cells, remodel the chromatin structure via a variety of enzymatic reactions. The reversible processes of some modifications, such as acetylation and deacetylation, are well characterized. Whether methylation and demethylation reversibly contribute to gene regulation, however, remains controversial. Recent studies have shown that methylation and demethylation are universally used to posttranslationally modify histones for the regulation of gene activity. In addition to LSD1, a nuclear amine oxidase homolog, which was found to function as a histone lysine demethylase, we and others found that some JmjC domain-containing proteins are histone demethylases. Specifically, we found that members of the JMJD2 protein family are histone demethylases that act on trimethyl groups of H3-K9 and H3-K36. Moreover, some members of this family also have activity for dimethyl groups. To understand the relationships between the structures and functions of these proteins, we have determined the structure of the catalytic core of the JMJD2A protein. From this structure, several novel structure features were revealed, such as the novel JmjN domain, the JmjC domain, the C-terminal domain, and a zinc finger motif. These unique structural features create a potential catalytic center. The structure also revealed a characteristic signature motif, which includes structural determinants for cofactors such as Fe(II) and a-ketoglutarate, a hallmark of non-heme containing oxygenases. There, however, are major questions that remained to be answered, such as the structural basis of the recognition of the substrate peptides by the catalytic core, the structural determinants of the specificity for the methyl groups, the mechanisms that regulate the activity of the enzyme, and the relationships among different JmjC domain-containing histone demethylases. Furthermore, only a small percentage of JmjC domain-containing proteins have been characterized. We believe that there are uncharacterized histone demethylases within the family of JmjC domain-containing proteins. Structural information derived from JMJD2A should be helpful in revealing these new histone demethylases. To answer these questions and to understand how the JmjC domain-containing histone demethylases function, three specific aims are proposed. Specific aim 1 - To characterize the structures and functions of the JMJD2 family members and to identify new histone demethylases; Specific aim 2 - To determine the high-resolution structures of the catalytic core of JMJD2A in the presence of different substrate peptides; Specific aim 3 - To determine the high-resolution structures of the entire JMJD2A protein and of the catalytic cores of other JmjC domain-containing proteins and to reveal the mechanisms that regulate the activity of JMJD2A and other related proteins.