Evolutionary Analysis and Comparative Genomics of Protein Superfamilies

1)阿拉文德博士和他的团队进行了研究，以确定新的核酸修饰酶。核酸中的修饰碱基提供了一层信息，指导着生物功能，超过了传统碱基的编码能力。最近，依赖于2-氧戊二酸和铁(II)的双加氧酶超家族的成员被预测并随后被证实催化核酸中碱基的氧化修饰。该超家族可以修饰从小分子到生物聚合物的各种底物。其中，两个不同的家族，即AlkB和动质体碱基J结合蛋白(JBP)催化核酸中碱基的原位羟化。利用对基因组结构和蛋白质结构域结构的序列、结构和上下文信息的敏感计算分析，他们报告了五个不同的依赖于2-氧戊二酸和铁(II)的双加氧酶家族，他们预测这些双加氧酶参与核酸修饰。在DNA修饰家族中，他们发现，动质体基J结合蛋白的双加氧酶结构域属于一个更大的家族，其中包括以人类癌基因Tet1为原型的Tet蛋白，以及担子菌真菌、绿藻、异叶变形鞭毛虫和噬菌体的蛋白。他们提出的证据表明，其中一些蛋白质可能参与了胞嘧啶5-甲基的氧化修饰，导致5-羟甲基胞嘧啶的形成。担子菌真菌如漆树属和鸡腿霉的Tet/JBP同源物显示出大量的谱系特异性扩增，并与编码一个新的和独特的转座酶家族的基因紧密连锁，该家族是大漩涡样HMG家族的成员。他们提出，这些真菌成员是移动转座子的一部分。这是首次报道一种真核转座元件，它编码自己的DNA修饰酶，具有潜在的调节作用。通过对其他特性不佳的DNA修饰酶的更广泛的分析，他们还表明，催化腺嘌呤N6-氨甲酰甲基化的噬菌体Mu妈妈样蛋白也与不同的细菌转座酶家族有关，这表明转座元件对DNA的修饰可能比之前所认为的更普遍。在本研究中确定的其他依赖于2-氧戊二酸和铁(II)的双加氧酶家族中，有一个在藻类中发现，预计主要由RNA修饰酶组成，并在蛋白质结构域结构中显示出惊人的多样性，表明存在可能具有独特适应作用的RNA修饰。这里提出的结果可能为未来研究意想不到的表观遗传修饰提供手段，如羟甲基胞嘧啶，可能深刻影响我们对基因调控和过程的理解，如DNA去甲基化。 2)DNA胞嘧啶甲基化是逆转座子沉默和哺乳动物发育的关键。在对可以修饰5-甲基胞嘧啶(5mC)的酶的计算机搜索中，Aravind博士确定Tet蛋白是锥体蛋白JBP1和JBP2的哺乳动物同系物，这两种蛋白被认为可以氧化胸腺嘧啶的5-甲基。与他的合作者，哈佛医学院免疫疾病研究所的Anjana Rao博士发现，TET1是急性髓系白血病MLL基因的融合伙伴，是一种依赖于2-氧戊二酸(2OG)和铁(II)的酶，在培养细胞和体外催化5mC转化为5-羟甲基胞嘧啶(HMC)。HMC存在于小鼠胚胎干细胞的基因组中，在RNA干扰介导的TET1缺失后，HMC水平降低。因此，Tet蛋白通过将5mC修饰为HMC，在表观遗传调控中具有潜在的作用。 3)阿拉文博士发起了一个关于染色质蛋白质的项目，该项目导致发现了一种新的组蛋白伴侣，它是HIR-ASF1组蛋白伴侣复合体的一部分。使用蛋白质序列图谱分析方法，他研究了HIR-ASF1复合蛋白质的进化，并表明它们具有比之前所知的更广泛的系统模式。他建立了动物的组蛋白-脱乙酰酶-复合体相互作用蛋白，凯恩/卡宾，是Hir3p的同源基因。它们含有一个保守的核心，由大约30个类似TPR的双螺旋重复序列组成，很可能形成一个超级螺旋支架。他还在所有的Hpc2p同源物中发现了一个保守的结构域，Hun结构域，包括动物ubinuclein/yemanuclein和最近发现的脊椎动物细胞周期调节因子FLJ25778。Hun结构域具有保守的酸性残基的特征模式，根据这一模式，他预测它是一种以前未被识别的组蛋白尾部结合伴侣。通过分析各种高通量数据集，如RNAi击倒、遗传和蛋白质相互作用图以及细胞周期特异性基因表达数据，我们提供了证据，表明Hpc2p同源物可能部署在与脊椎动物细胞周期进展和疟原虫分裂相关的染色质动力学的特定过程中。

1) Dr. Aravind and his group performed research to identify novel nucleic acid modification enzymes. Modified bases in nucleic acids present a layer of information that directs biological function over and beyond the coding capacity of the conventional bases. Recently, members of the 2-oxoglutarate- and iron(II)-dependent dioxygenase super-family, which modify diverse substrates from small molecules to biopolymers, were predicted and subsequently confirmed to catalyze oxidative modification of bases in nucleic acids. Of these, two distinct families, namely the AlkB and the kinetoplastid base J binding proteins (JBP) catalyze in situ hydroxylation of bases in nucleic acids. Using sensitive computational analysis of sequences, structures and contextual information from genomic structure and protein domain architectures, they reported five distinct families of 2-oxoglutarate- and iron(II)-dependent dioxygenase that they predict to be involved in nucleic acid modifications. Among the DNA-modifying families, they showed that the dioxygenase domains of the kinetoplastid base J-binding proteins belong to a larger family that includes the Tet proteins, prototyped by the human oncogene Tet1, and proteins from basidiomycete fungi, chlorophyte algae, heterolobosean amoeboflagellates and bacteriophages. They presented evidence that some of these proteins are likely to be involved in oxidative modification of the 5-methyl group of cytosine leading to the formation of 5-hydroxymethylcytosine. The Tet/JBP homologs from basidiomycete fungi such as Laccaria and Coprinopsis show large lineage-specific expansions and a tight linkage with genes encoding a novel and distinct family of predicted transposases, and a member of the Maelstrom-like HMG family. They proposed that these fungal members are part of a mobile transposon. This is the first report of a eukaryotic transposable element that encodes its own DNA-modification enzyme with a potential regulatory role. Through a wider analysis of other poorly characterized DNA-modifying enzymes they also showed that the phage Mu Mom-like proteins, which catalyze the N6-carbamoylmethylation of adenines, are also linked to diverse families of bacterial transposases, suggesting that DNA modification by transposable elements might have a more general presence than previously appreciated. Among the other families of 2-oxoglutarate- and iron(II)-dependent dioxygenases identified in this study, one which is found in algae, is predicted to mainly comprise of RNA-modifying enzymes and shows a striking diversity in protein domain architectures suggesting the presence of RNA modifications with possibly unique adaptive roles. The results presented here are likely to provide the means for future investigation of unexpected epigenetic modifications, such as hydroxymethyl cytosine, that could profoundly impact our understanding of gene regulation and processes such as DNA demethylation. 2)DNA cytosine methylation is crucial for retrotransposon silencing and mammalian development. In a computational search for enzymes that could modify 5-methylcytosine (5mC), Dr. Aravind identified TET proteins as mammalian homologs of the trypanosome proteins JBP1 and JBP2, which have been proposed to oxidize the 5-methyl group of thymine. In conjunction with his collaborator Dr. Anjana Rao, Immune Disease Institute, Harvard Medical School it was shown that TET1, a fusion partner of the MLL gene in acute myeloid leukemia, is a 2-oxoglutarate (2OG)- and Fe(II)-dependent enzyme that catalyzes conversion of 5mC to 5-hydroxymethylcytosine (hmC) in cultured cells and in vitro. hmC is present in the genome of mouse embryonic stem cells, and hmC levels decrease upon RNA interference-mediated depletion of TET1. Thus, TET proteins have potential roles in epigenetic regulation through modification of 5mC to hmC. 3)Dr. Aravind initiated a project on chromatin proteins that lead to the discovery of a novel histone chaperone that function as part of the Hir-Asf1 histone chaperone complex. Using protein sequence profile analyses methods he investigated the evolution of Hir-Asf1 complex proteins and showed that they have a much wider phyletic pattern than was previously known. He established the animal histone-deacetylase-complex-interacting proteins, CAIN/CABIN, to be orthologs of Hir3p. They contain a conserved core of around 30 TPR-like bi-helical repeats that are likely to form a super-helical scaffold. He also identified a conserved domain, the HUN domain, in all Hpc2p homologs, including animal ubinuclein/yemanuclein and the recently discovered vertebrate cell-cycle regulator FLJ25778. The HUN domain has a characteristic pattern of conserved acidic residues based on which he predicted that it is a previously unrecognized histone-tail-binding chaperone. By analyzing various high-throughput data sets, such as RNAi knock-downs, genetic and protein interaction maps and cell-cycle-specific gene expression data, we present evidence that Hpc2p homologs might be deployed in specific processes of chromatin dynamics relating to cell-cycle progression in vertebrates and schizogony in Plasmodium.