ISC, a Novel Group of Bacterial and Archaeal DNA Transposons That Encode Cas9 Homologs.
ISC, a Novel Group of Bacterial and Archaeal DNA Transposons That Encode Cas9 Homologs.
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DOI：
10.1128/jb.00783-15
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发表时间：
2015-12-28
影响因子：
3.2
通讯作者：
Koonin EV
中科院分区：
文献类型：
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作者：
Kapitonov VV;Makarova KS;Koonin EV
细菌基因组编码Cas9的许多同源物，Cas9是II型CRISPR-Cas系统的效应蛋白。同源区包括富含丝氨酸的螺旋和插入RuvC样核酸酶结构域的HNH核酸酶结构域。然而，这些基因与cas基因或CRISPR无关。在这里，我们表明Cas9同源物代表了一组不同的非自主转座子，我们将其称为ISC（插入序列Cas9样）。我们鉴定了许多不同的全长ISC转座子家族，并证明它们的末端序列（特别是3′末端）与IS 605超家族转座子的末端序列相似，IS 605超家族转座子由TnpA基因编码的Y1酪氨酸转座酶动员，通常也编码含有RuvC样内切酶结构域的TnpB蛋白。ISC和IS 605转座子的末端区域含有可能被Y1转座酶识别的回文结构。来自这两组的转座子被精确地插入特定4-bp靶位点的中间或上游，而没有靶位点重复。我们还确定了自主ISC转座子编码TnPA样Y1转座酶。因此，非自主ISC转座子可以被其他自主ISC转座子的Y1转座酶或更丰富的IS 605转座子的Y1转座酶反式移动。这些发现暗示了一种进化情景，其中ISC转座子可能通过插入编码HNH结构域的移动的II组内含子从IS 605家族转座子进化，并且Cas9随后通过ISC转座子的固定化进化。Cas9内切核酸酶是II型CRISPR-Cas系统的效应子，代表了新一代基因组工程工具。在这里，我们详细描述了一个新的转座因子家族，该家族编码Cas9的可能祖先，并概述了连接这些转座子和Cas9的不同品种的进化情景。
Bacterial genomes encode numerous homologs of Cas9, the effector protein of the type II CRISPR-Cas systems. The homology region includes the arginine-rich helix and the HNH nuclease domain that is inserted into the RuvC-like nuclease domain. These genes, however, are not linked to cas genes or CRISPR. Here, we show that Cas9 homologs represent a distinct group of nonautonomous transposons, which we denote ISC (insertion sequences Cas9-like). We identify many diverse families of full-length ISC transposons and demonstrate that their terminal sequences (particularly 3′ termini) are similar to those of IS605 superfamily transposons that are mobilized by the Y1 tyrosine transposase encoded by the TnpA gene and often also encode the TnpB protein containing the RuvC-like endonuclease domain. The terminal regions of the ISC and IS605 transposons contain palindromic structures that are likely recognized by the Y1 transposase. The transposons from these two groups are inserted either exactly in the middle or upstream of specific 4-bp target sites, without target site duplication. We also identify autonomous ISC transposons that encode TnpA-like Y1 transposases. Thus, the nonautonomous ISC transposons could be mobilized in trans either by Y1 transposases of other, autonomous ISC transposons or by Y1 transposases of the more abundant IS605 transposons. These findings imply an evolutionary scenario in which the ISC transposons evolved from IS605 family transposons, possibly via insertion of a mobile group II intron encoding the HNH domain, and Cas9 subsequently evolved via immobilization of an ISC transposon. IMPORTANCE Cas9 endonucleases, the effectors of type II CRISPR-Cas systems, represent the new generation of genome-engineering tools. Here, we describe in detail a novel family of transposable elements that encode the likely ancestors of Cas9 and outline the evolutionary scenario connecting different varieties of these transposons and Cas9.