Wagner E G, von Heijne J, Nordström K
EMBO J. 1987 Feb;6(2):515-22. doi: 10.1002/j.1460-2075.1987.tb04783.x.
Replication of IncFII plasmids is regulated through the expression of a gene, repA. The RepA protein is rate-limiting for initiation of replication. The main negative control is exerted by a countertranscript, CopA RNA, that binds to the complementary region of the RepA mRNA, thereby inhibiting the formation of the RepA protein. The target region for CopA RNA, CopT, is located upstream of the RepA coding region. An open reading frame for a putative 7k protein overlaps the CopT sequence. Here we show by using lacZ fusions that the 7k gene is expressed. We constructed a translation start mutation in order to abolish formation of the 7k protein. This resulted in a 10-fold decrease in repA expression. The 7k protein produced in trans did not reverse this effect, so the 7k protein per se does not control expression of repA. However, translation of the 7k coding sequence must influence CopA/CopT RNA recognition, since ribosomes will transiently disrupt the target hairpin. We propose here a novel mechanism that affects the level of gene expression: the 7k region of the RepA mRNA is a leader sequence that is involved in expression of the downstream gene; translation of the 7k region competes with a negative control system involving RNA-RNA interaction.
IncFII 质粒的复制通过一个基因 repA 的表达来调控。RepA 蛋白是复制起始的限速因子。主要的负调控由一种反义转录本 CopA RNA 行使，它与 RepA mRNA 的互补区域结合，从而抑制 RepA 蛋白的形成。CopA RNA 的靶区域 CopT 位于 RepA 编码区的上游。一个假定的 7k 蛋白的开放阅读框与 CopT 序列重叠。在这里我们通过使用 lacZ 融合表明 7k 基因是表达的。我们构建了一个翻译起始突变以消除 7k 蛋白的形成。这导致 repA 表达下降了 10 倍。反式产生的 7k 蛋白并没有逆转这种效应，所以 7k 蛋白本身并不控制 repA 的表达。然而，7k 编码序列的翻译必定影响 CopA/CopT RNA 的识别，因为核糖体将暂时破坏靶发夹结构。我们在此提出一种影响基因表达水平的新机制：RepA mRNA 的 7k 区域是一个参与下游基因表达的前导序列；7k 区域的翻译与一个涉及 RNA-RNA 相互作用的负调控系统竞争。
