Institute of Biotechnology, Life Sciences Center, Vilnius University, Vilnius, Lithuania.
Nature. 2021 Nov;599(7886):692-696. doi: 10.1038/s41586-021-04058-1. Epub 2021 Oct 7.
Transposition has a key role in reshaping genomes of all living organisms. Insertion sequences of IS200/IS605 and IS607 families are among the simplest mobile genetic elements and contain only the genes that are required for their transposition and its regulation. These elements encode tnpA transposase, which is essential for mobilization, and often carry an accessory tnpB gene, which is dispensable for transposition. Although the role of TnpA in transposon mobilization of IS200/IS605 is well documented, the function of TnpB has remained largely unknown. It had been suggested that TnpB has a role in the regulation of transposition, although no mechanism for this has been established. A bioinformatic analysis indicated that TnpB might be a predecessor of the CRISPR-Cas9/Cas12 nucleases. However, no biochemical activities have been ascribed to TnpB. Here we show that TnpB of Deinococcus radiodurans ISDra2 is an RNA-directed nuclease that is guided by an RNA, derived from the right-end element of a transposon, to cleave DNA next to the 5'-TTGAT transposon-associated motif. We also show that TnpB could be reprogrammed to cleave DNA target sites in human cells. Together, this study expands our understanding of transposition mechanisms by highlighting the role of TnpB in transposition, experimentally confirms that TnpB is a functional progenitor of CRISPR-Cas nucleases and establishes TnpB as a prototype of a new system for genome editing.
转座在所有生物的基因组重排中起着关键作用。IS200/IS605 和 IS607 家族的插入序列是最简单的移动遗传元件之一，仅包含转座及其调控所需的基因。这些元件编码 tnpA 转座酶，这是移动所必需的，并且通常携带一个辅助的 tnpB 基因，该基因对于转座是可有可无的。虽然 TnpA 在 IS200/IS605 转座子的转座中的作用已有详细记录，但 TnpB 的功能在很大程度上仍然未知。有人认为 TnpB 在转座的调控中起作用，尽管尚未建立这种作用的机制。生物信息学分析表明，TnpB 可能是 CRISPR-Cas9/Cas12 核酸酶的前身。然而，尚未赋予 TnpB 任何生化活性。在这里，我们表明来自 Deinococcus radiodurans ISDra2 的 TnpB 是一种 RNA 指导的核酸酶，由来自转座子的右端元件的 RNA 引导，以切割 DNA 紧邻 5'-TTGAT 转座子相关基序。我们还表明，TnpB 可以被重新编程以切割人类细胞中的 DNA 靶位点。总之，这项研究通过强调 TnpB 在转座中的作用，扩展了我们对转座机制的理解，实验证实了 TnpB 是 CRISPR-Cas 核酸酶的功能性前体，并确立了 TnpB 作为一种新的基因组编辑系统的原型。
