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首页> 外文期刊>Molecular and Cellular Biology >An in vivo assay for the reverse transcriptase of human retrotransposon L1 in Saccharomyces cerevisiae.
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An in vivo assay for the reverse transcriptase of human retrotransposon L1 in Saccharomyces cerevisiae.

机译：酿酒酵母中人逆转录转座子L1逆转录酶的体内测定。

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摘要

L1 elements constitute a highly repetitive human DNA family (50,000 to 100,000 copies) lacking long terminal repeats and ending in a poly(A) tail. Some L1 elements are capable of retrotransposition in the human genome (Kazazian, H. H., Jr., C. Wong, H. Youssoufian, A. F. Scott, D. G. Phillips, and S.E. Antonarakis, Nature (London) 332:164-166, 1988). Although most are 5' truncated, a consensus sequence of complete L1 elements is 6 kb long and contains two open reading frames (ORFs) (Scott, A. F., B. J. Schmeckpeper, M. Abdelrazik, C. T. Comey, B. O'Hara, J. P. Rossiter, T. Cooley, P. Health, K. D. Smith, and L. Margolet, Genomics 1:113-125, 1987). The protein encoded by ORF2 has reverse transcriptase (RT) activity in vitro (Mathias, S. L., A. F. Scott, H. H. Kazazian, Jr., J. D. Boeke, and A. Gabriel, Science 254:1808-1810, 1991). Because L1 elements are so numerous, efficient methods for identifying active copies are required. We have developed a simple in vivo assay for the activity of L1 RT based on the system developed by Derr et al. (Derr, L. K., J. N. Strathern, and D. J. Garfinkel, Cell 67:355-364, 1991) for yeast HIS3 pseudogene formation. L1 ORF2 displays an in vivo RT activity similar to that of yeast Ty1 RT in this system and generates pseudogenes with unusual structures. Like the HIS3 pseudogenes whose formation depends on Ty1 RT, the HIS3 pseudogenes generated by L1 RT are joined to Ty1 sequences and often are part of complex arrays of Ty1 elements, multiple HIS3 pseudogenes, and hybrid Ty1/L1 elements. These pseudogenes differ from those previously described in that there are base pairs of unknown origin inserted at several of the junctions. In two of three HIS3 pseudogenes studied, the L1 RT appears to have jumped from the 5' end of a Ty1/L1 transcript to the poly(A) tract of the HIS3 RNA.
机译：L1元素构成了高度重复的人类DNA家族（50,000至100,000个拷贝），没有长的末端重复序列并以poly（A）尾部结尾。一些L1元件能够在人类基因组中逆转录（Kazazian，H.H.，Jr.，C.Wong，H.Youssoufian，A.F.Scott，D.G.Phillips和S.E. Antonarakis，Nature（伦敦）332：164-166，1988）。尽管大多数被5'截短，但完整L1元件的共有序列长6 kb，并包含两个开放阅读框（ORF）（Scott，AF，BJ Schmeckpeper，M。Abdelrazik，CT Comey，B.O'Hara，JP Rossiter ，T.Cooley，P.Health，KD Smith和L.Margolet，Genomics 1：113-125，1987）。由ORF2编码的蛋白质在体外具有逆转录酶（RT）活性（Mathias，S.L.，A.F. Scott，H.H. Kazazian，Jr.，J.D. Boeke，and A.Gabriel，Science 254：1808-1810，1991）。由于L1元素数量众多，因此需要用于识别活动副本的有效方法。我们已经根据Derr等人开发的系统开发了一种简单的L1 RT活性体内检测方法。 （Derr，L.K.，J.N.Strathern，和D.J.Garfinkel，Cell 67：355-364，1991）用于酵母HIS3假基因的形成。 L1 ORF2在该系统中显示出类似于酵母Ty1 RT的体内RT活性，并产生具有异常结构的假基因。就像其HIS3假基因的形成取决于Ty1 RT一样，由L1 RT生成的HIS3假基因也与Ty1序列相连，并且通常是Ty1元素，多个HIS3假基因和混合Ty1 / L1元素的复杂数组的一部分。这些假基因与先前描述的假基因的不同之处在于，在几个连接处插入了未知来源的碱基对。在研究的三个HIS3假基因中的两个中，L1 RT似乎已从Ty1 / L1转录本的5'端跳至HIS3 RNA的poly（A）区域。

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