Somasekhar G, Szybalski W
Virology. 1987 Jun;158(2):414-26. doi: 10.1016/0042-6822(87)90213-3.
Expression of the late genes of bacteriophage lambda requires, in addition to the host functions, the lambda p'R promoter, the antiterminator sequence qut, and the product of gene Q which interacts with the Q utilization (qut) site. In the absence of the Q function or qut site, the p'R-initiated transcription is blocked by the t'R terminator at the 194th nucleotide downstream of the start point, s'R, producing a short 6 S mRNA. In this study the position and boundaries of the qut site were deduced by constructing plasmids containing various portions of the p'R-qut region, the t'R1 terminator, and the reporter gene galK. We measured galK gene expression in response to the gamma Q gene product supplied in trans by a prophage or Q-expression plasmid. We show that among the lambda proteins, the Q gene product alone is necessary and sufficient for complete qut-mediated transcription antitermination in vivo. These antitermination experiments, employing plasmids that contain different lengths of lambda p'R-qut sequence, identified the right boundary of the qut site, which is located between +4 and +18 (for s'R = +1). The functional left boundary of qut does not extend upstream from the -26th nucleotide of the p'R promoter, as based on the following experiments. The promoter function of the truncated (-26)p'R-s'R-(+18) sequence can be restored by fusion to the complete but qut-less p'R, pp, or PLac promoter; however, no antitermination was observed for such a p-(-26)p'R-s'R-(+18)-t'R-galK plasmid. Thus we conclude that the qut site partially overlaps with the p'R promoter sequence. However, promoters that contain the -10 region of p'R, s'R, and the +1 to +18 qut sequence did mediate Q-dependent antitermination when properly fused to the homologous or heterologous -35 promoter regions. Only those transcripts that start at s'R (+1 or very near to it) and also contain at least the first 18 nucleotides (actually greater than 4 and less than or equal to 18) of 6 S RNA appear to be a target for the Q-qut-mediated transcription antitermination, which acts not only at t'R but also at other Rho-independent or Rho-dependent terminators.
λ噬菌体晚期基因的表达除了需要宿主功能外，还需要λ p'R启动子、抗终止序列qut以及与Q利用（qut）位点相互作用的基因Q产物。在缺乏Q功能或qut位点的情况下，p'R起始的转录在起始点s'R下游第194个核苷酸处被t'R终止子阻断，产生短的6S mRNA。在本研究中，通过构建包含p'R-qut区域、t'R1终止子和报告基因galK不同部分的质粒，推导了qut位点的位置和边界。我们通过原噬菌体或Q表达质粒反式提供的γQ基因产物来测量galK基因的表达。我们表明，在λ蛋白中，仅基因Q产物对于体内完整的qut介导的转录抗终止是必需且充分的。这些抗终止实验使用了包含不同长度λ p'R-qut序列的质粒，确定了qut位点的右边界，其位于+4和+18之间（对于s'R = +1）。基于以下实验，qut的功能性左边界不会从p'R启动子的-26个核苷酸向上游延伸。截短的（-26）p'R-s'R-（+18）序列的启动子功能可以通过与完整但无qut的p'R、pp或PLac启动子融合来恢复；然而，对于这样的p-（-26）p'R-s'R-（+18）-t'R-galK质粒未观察到抗终止现象。因此我们得出结论，qut位点与p'R启动子序列部分重叠。然而，包含p'R的-10区域、s'R和+1至+18 qut序列的启动子在与同源或异源-35启动子区域正确融合时确实介导了Q依赖性抗终止。只有那些从s'R（+1或非常接近它）起始并且还包含6S RNA至少前18个核苷酸（实际上大于4且小于或等于18）的转录本似乎是Q-qut介导的转录抗终止的靶标，该抗终止不仅作用于t'R，还作用于其他Rho非依赖性或Rho依赖性终止子。
