Geisinger Edward, Adhikari Rajan P, Jin Ruzhong, Ross Hope F, Novick Richard P
Department of Microbiology, Skirball Institute, NYU Medical Center, New York, NY 10016, USA.
Mol Microbiol. 2006 Aug;61(4):1038-48. doi: 10.1111/j.1365-2958.2006.05292.x.
RNAIII is a 514 nt regulatory RNA that is the effector molecule of the staphylococcal agr quorum-sensing system, regulating a large set of virulence and other accessory genes at the level of transcription. RNAIII was discovered nearly 20 years ago and we long ago hypothesized that it would function by regulating the synthesis or activity of one or more intermediary transcription factors. We have finally confirmed this hypothesis, showing that Staphylococcus aureus RNAIII regulates the synthesis of a major pleiotropic transcription factor, Rot, by blocking its translation. RNAIII has a complex secondary structure with several stable hairpins that have highly C-rich end loops, unusual in an AT-rich organism. We noted that these loops are complementary to two G-rich stem loops of the rot mRNA translation initiation region (TIR). Pairing of the complementary RNAs would be predicted to occlude the rot Shine-Dalgarno (SD) site and to block rot translation. Through a combination of transcriptional and translational fusions and Northern and Western blot hybridization analyses, we show that RNAIII does, indeed, block rot translation. Through alterations in the C-rich loops of RNAIII and the G-rich loops of rot, we show that the sequences of these loops are critical for inhibition of rot translation and suggest that this inhibition is affected by pairing between the complementary stem loops, followed by the cleavage of rot mRNA. We propose that the RNAIII-rot mRNA interaction plays a key role in agr regulation of staphylococcal virulence.
RNAIII是一种514个核苷酸的调控RNA，是葡萄球菌agr群体感应系统的效应分子，在转录水平上调控大量毒力基因和其他辅助基因。RNAIII于近20年前被发现，我们早就推测它可能通过调控一种或多种中间转录因子的合成或活性来发挥作用。我们最终证实了这一推测，表明金黄色葡萄球菌RNAIII通过阻断主要多效转录因子Rot的翻译来调控其合成。RNAIII具有复杂的二级结构，带有几个稳定的发夹结构，其末端环富含C，这在富含A/T的生物体中并不常见。我们注意到这些环与rot mRNA翻译起始区域（TIR）的两个富含G的茎环互补。预测互补RNA的配对会封闭rot的Shine-Dalgarno（SD）位点并阻断rot的翻译。通过转录和翻译融合以及Northern和Western印迹杂交分析相结合，我们表明RNAIII确实能阻断rot的翻译。通过改变RNAIII富含C的环和rot富含G的环，我们表明这些环的序列对于抑制rot的翻译至关重要，并表明这种抑制作用受互补茎环之间配对的影响，随后是rot mRNA的切割。我们提出RNAIII与rot mRNA的相互作用在葡萄球菌毒力的agr调控中起关键作用。
