Robin S, Chambeyron S, Brun C, Bucheton A, Busseau I
Institut de Génétique Humaine, CNRS, 141 Rue de la Cardonille, 34396 Montpellier Cedex 5, France.
Mol Genet Genomics. 2002 Aug;267(6):829-34. doi: 10.1007/s00438-002-0716-z. Epub 2002 Jul 9.
I factors are non-LTR retrotransposons of Drosophila melanogaster that transpose at high frequency in the germline of females resulting from appropriate crosses, allowing in vivo studies of the retrotransposition process. Reverse transcription of a full-length RNA intermediate is thought to occur at the site of integration, using a 3' hydroxyl group generated by endonucleolytic cleavage of the genomic DNA to prime synthesis of the first cDNA strand. This target-primed reverse transcription (TPRT) process is mediated by endonuclease and reverse transcriptase activities encoded by the element. We have designed a molecularly tagged, endonuclease-defective I element that can be mobilised with high efficiency by constructs that express the product of the I factor ORF2 in trans. This indicates that the endonuclease activity required for retrotransposition of the I factor can be provided in trans. Using this system, we show that the endonuclease domain of the R1Bm retrotransposon from Bombyx mori cannot functionally replace that of the I factor.
I因子是黑腹果蝇的非LTR逆转座子，在适当杂交产生的雌性生殖系中高频转座，从而能够对逆转座过程进行体内研究。全长RNA中间体的逆转录被认为发生在整合位点，利用基因组DNA内切核酸酶切割产生的3'羟基来起始第一条cDNA链的合成。这种靶标引发的逆转录（TPRT）过程由该元件编码的内切核酸酶和逆转录酶活性介导。我们设计了一种分子标记的、内切核酸酶缺陷的I元件，它可以被反式表达I因子ORF2产物的构建体高效动员。这表明I因子逆转座所需的内切核酸酶活性可以反式提供。利用这个系统，我们发现家蚕R1Bm逆转座子的内切核酸酶结构域不能在功能上替代I因子的内切核酸酶结构域。
