Minchiotti G, Di Nocera P P
International Institute of Genetics and Biophysics, Consiglio Nazionale Delle Ricerche, Naples, Italy.
Mol Cell Biol. 1991 Oct;11(10):5171-80. doi: 10.1128/mcb.11.10.5171-5180.1991.
Drosophila melanogaster F elements are mobile, oligo(A)-terminated DNA sequences that likely propagate by the retrotranscription of RNA intermediates. Plasmids bearing DNA segments from the left-hand region of a full-length F element fused to the CAT gene were used as templates for transient expression assays in Drosophila Schneider II cultured cells. Protein and RNA analyses led to the identification of two promoters, Fin and Fout, that transcribe in opposite orientations. The Fin promoter drives the synthesis of transcripts that initiate around residue +6 and are directed toward the element. Fin, that probably controls the formation of F transposition RNA intermediates and gene products, is internal to the transcribed region. Sequences important for accumulation of Fin transcripts are included within the +1 to +30 interval; an additional regulatory element may coincide with a heptamer located downstream of this region also found in the 5' end regions of F-like Drosophila retrotransposons. Analysis of the template activity of 3' deletion derivatives indicates that the level of accumulation of Fin RNA is also dependent upon the presence of sequences located within the +175 to +218 interval. The Fout promoter drives transcription in the opposite orientation with respect to Fin. Fout transcripts initiate at nearby sites within the +92 to +102 interval. Sequences downstream of these multiple RNA start sites are not required for the activity of the Fout promoter. Deletions knocking out the Fin promoter do not impair Fout transcription; conversely, initiation at the Fin promoter still takes place in templates that lack the Fout promoter. At a low level, both promoters are active in cultured cells.
果蝇黑腹果蝇F元件是可移动的、以寡聚(A)结尾的DNA序列,可能通过RNA中间体的逆转录进行传播。携带来自全长F元件左手区域与CAT基因融合的DNA片段的质粒被用作果蝇施耐德II培养细胞中瞬时表达测定的模板。蛋白质和RNA分析导致鉴定出两个启动子,Fin和Fout,它们以相反的方向转录。Fin启动子驱动转录本的合成,这些转录本在大约+6位点开始并指向元件。Fin可能控制F转座RNA中间体和基因产物的形成,位于转录区域内部。对于Fin转录本积累重要的序列包含在+1至+30区间内;一个额外的调控元件可能与位于该区域下游的七聚体一致,该七聚体也存在于F样果蝇逆转座子的5'末端区域。对3'缺失衍生物模板活性的分析表明,Fin RNA的积累水平也取决于+175至+218区间内序列的存在。Fout启动子相对于Fin以相反的方向驱动转录。Fout转录本在+92至+102区间内的附近位点起始。这些多个RNA起始位点下游的序列对于Fout启动子的活性不是必需的。敲除Fin启动子的缺失不会损害Fout转录;相反,在缺乏Fout启动子的模板中,Fin启动子处的起始仍然会发生。在低水平下,两个启动子在培养细胞中都是活跃的。