Transposons are mobile genetic elements that colonize genomes and drive their plasticity in all organisms. DNA transposon-encoded transposases bind to the ends of their cognate transposons and catalyze their movement. In some cases, exaptation of transposon genes has allowed novel cellular functions to emerge. The PiggyMac (Pgm) endonuclease of the ciliate Paramecium tetraurelia is a domesticated transposase from the PiggyBac family. It carries a core catalytic domain typical of PiggyBac-related transposases and a short cysteine-rich domain (CRD), flanked by N- and C-terminal extensions. During sexual processes Pgm catalyzes programmed genome rearrangements (PGR) that eliminate ~ 30% of germline DNA from the somatic genome at each generation. How Pgm recognizes its DNA cleavage sites in chromatin is unclear and the structure-function relationships of its different domains have remained elusive. We provide insight into Pgm structure by determining the fold adopted by its CRD, an essential domain required for PGR. Using Nuclear Magnetic Resonance, we show that the Pgm CRD binds two Zn2+ ions and forms an unusual binuclear cross-brace zinc finger, with a circularly permutated treble-clef fold flanked by two flexible arms. The Pgm CRD structure clearly differs from that of several other PiggyBac-related transposases, among which is the well-studied PB transposase from Trichoplusia ni. Instead, the arrangement of cysteines and histidines in the primary sequence of the Pgm CRD resembles that of active transposases from piggyBac-like elements found in other species and of human PiggyBac-derived domesticated transposases. We show that, unlike the PB CRD, the Pgm CRD does not bind DNA. Instead, it interacts weakly with the N-terminus of histone H3, whatever its lysine methylation state. The present study points to the structural diversity of the CRD among transposases from the PiggyBac family and their domesticated derivatives, and highlights the diverse interactions this domain may establish with chromatin, from sequence-specific DNA binding to contacts with histone tails. Our data suggest that the Pgm CRD fold, whose unusual arrangement of cysteines and histidines is found in all PiggyBac-related domesticated transposases from Paramecium and Tetrahymena, was already present in the ancestral active transposase that gave rise to ciliate domesticated proteins.

中文翻译：

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PiggyMac 富含半胱氨酸结构域的不寻常结构揭示了 PiggyBac 相关转座酶中锌指的多样性

转座子是可移动的遗传元件，可在所有生物体中定殖基因组并驱动其可塑性。DNA 转座子编码的转座酶与其同源转座子的末端结合并催化其运动。在某些情况下，转座子基因的外延使得新的细胞功能出现。纤毛虫草履虫的 PiggyMac (Pgm) 核酸内切酶是来自 PiggyBac 家族的驯化转座酶。它携带一个 PiggyBac 相关转座酶的典型核心催化结构域和一个短的富含半胱氨酸结构域 (CRD)，两侧是 N 端和 C 端延伸。在性过程中，Pgm 会催化程序化基因组重排 (PGR)，从而在每一代从体细胞基因组中消除约 30% 的种系 DNA。Pgm 如何识别染色质中的 DNA 切割位点尚不清楚，其不同结构域的结构-功能关系仍然难以捉摸。我们通过确定其 CRD（PGR 所需的基本域）采用的折叠来深入了解 Pgm 结构。使用核磁共振，我们表明 Pgm CRD 结合两个 Zn2+ 离子，并形成一个不寻常的双核交叉支撑锌指，具有圆形排列的高音谱号折叠，两侧有两个柔性臂。Pgm CRD 结构明显不同于其他几种 PiggyBac 相关转座酶，其中包括来自 Trichoplusia ni 的经过充分研究的 PB 转座酶。相反，Pgm CRD 一级序列中半胱氨酸和组氨酸的排列类似于来自其他物种中发现的piggyBac 样元件的活性转座酶和人类PiggyBac 衍生的驯化转座酶。我们发现，与 PB CRD 不同，Pgm CRD 不结合 DNA。相反，无论其赖氨酸甲基化状态如何，它与组蛋白 H3 的 N 末端的相互作用都很弱。本研究指出了 PiggyBac 家族转座酶及其驯化衍生物中 CRD 的结构多样性，并强调了该结构域可能与染色质建立的多种相互作用，从序列特异性 DNA 结合到与组蛋白尾部的接触。我们的数据表明，Pgm CRD 折叠已经存在于产生纤毛虫驯化蛋白的祖先活性转座酶中，其半胱氨酸和组氨酸的不寻常排列存在于来自草履虫和四膜虫的所有 PiggyBac 相关驯化转座酶中。