Woodard Lauren E, Downes Laura M, Lee Yi-Chien, Kaja Aparna, Terefe Eyuel S, Wilson Matthew H
Department of Veterans Affairs, Nashville, TN 37212, USA and Department of Medicine, Vanderbilt University, Nashville, TN 37232, USA.
Department of Veterans Affairs, Houston, TX 77030, USA and Department of Medicine, Baylor College of Medicine, Houston, TX 77030, USA.
Nucleic Acids Res. 2017 Jan 9;45(1):353-366. doi: 10.1093/nar/gkw1115. Epub 2016 Nov 28.
Transposons are highly abundant in eukaryotic genomes, but their mobilization must be finely tuned to maintain host organism fitness and allow for transposon propagation. Forty percent of the human genome is comprised of transposable element sequences, and the most abundant cut-and-paste transposons are from the hAT superfamily. We found that the hAT transposase TcBuster from Tribolium castaneum formed filamentous structures, or rodlets, in human tissue culture cells, after gene transfer to adult mice, and ex vivo in cell-free conditions, indicating that host co-factors or cellular structures were not required for rodlet formation. Time-lapsed imaging of GFP-laced rodlets in human cells revealed that they formed quickly in a dynamic process involving fusion and fission. We delayed the availability of the transposon DNA and found that transposition declined after transposase concentrations became high enough for visible transposase rodlets to appear. In combination with earlier findings for maize Ac elements, these results give insight into transposase overproduction inhibition by demonstrating that the appearance of transposase protein structures and the end of active transposition are simultaneous, an effect with implications for genetic engineering and horizontal gene transfer.
转座子在真核生物基因组中高度丰富，但其移动必须得到精细调控，以维持宿主生物体的适应性并促进转座子的传播。人类基因组的40%由转座元件序列组成，最丰富的剪切粘贴型转座子来自hAT超家族。我们发现，来自赤拟谷盗的hAT转座酶TcBuster在基因转移至成年小鼠后，以及在无细胞条件下的体外实验中，在人类组织培养细胞中形成了丝状结构或小杆，这表明小杆的形成不需要宿主辅助因子或细胞结构。对人类细胞中带有绿色荧光蛋白的小杆进行延时成像显示，它们在一个涉及融合和裂变的动态过程中迅速形成。我们延缓了转座子DNA的可利用性，发现当转座酶浓度高到足以出现可见的转座酶小杆时，转座作用会下降。结合早期关于玉米Ac元件的研究结果，这些结果通过证明转座酶蛋白结构的出现与活跃转座的结束是同时发生的，深入揭示了转座酶过量产生抑制现象，这一效应对基因工程和水平基因转移具有重要意义。
