de Rouvroit C Lambert, Sluiters C, Cornelis G R
Microbial Pathogenesis Unit, International Institute of Cellular and Molecular Biology (ICP) and Faculté de Médecine, Université Catholique de Louvain, UCL 54.90, B-1200 Brussels, Belgium.
Mol Microbiol. 1992 Feb;6(3):395-409. doi: 10.1111/j.1365-2958.1992.tb01483.x.
Upon incubation at 37°C in the absence of Ca ions, pathogenic yersiniae release high amounts of pYV plasmid-encoded proteins called Yops, involved in pathogenesis. Yersinia enterocolitica also express two outer membrane proteins, an adhesin called YadA and a lipoprotein called YIpA. The production of the Yops is co-ordinately regulated by a 20kb region of the plasmid referred to as the 'Ca dependence region' and containing at least four loci called virA, virB, virC, and virF. The 8.5kb virC region, involved in the specific transport of the Yops, is a single operon containing 13 open reading frames called yscA to yscM. Gene virF encodes a key transcriptional activator of the yop, yadA and ylpA genes. It is only transcribed at 37°C and its expression is modulated by a chromosome-encoded histone-like protein called YmoA. We show here that virF also controls the virC operon. By contrast, VirF is not essential for the induction of virA and virB. The VirF protein binds specifically to yop promoters. In particular, it protects the region spanning nucleotides -64 to -34 of yopH. In order to analyse the role of temperature in the induction of the yop regulon, we constructed Y. enterocolitica strains expressing virF from the tac promoter. In spite of the fact that virF was transcribed at 25°C, neither the Yops nor YadA were expressed at that temperature. This poor response to VirF at 25°C was at least partially due to a weak and slow transcription of the genes controlled by virF. Surprisingly, when cloned on pACYC184, gene yadA was expressed even in absence of VirF, but remained thermodependent. Hence temperature and virF are both required for the induction of the yop regulon. Among other possible roles, temperature could modify the structure of either the activator itself or the yop promoter. The fact that VirF binds in vitro to yop promoters at 25°C rules out the first hypothesis. In order to test the second hypothesis, we studied, in vivo, the activity of the yopH promoter in ymoA mutants. The yopH promoter became active in the absence of VirF, indicating that yop promoter activity depends upon chromatin structure. We conclude from these two observations that, in vivo, temperature is required to modify the DNA structure of the yop promoters in order to allow the action of the transcriptional activator.
在无钙离子存在的情况下于37°C孵育时，致病性耶尔森菌会释放大量由pYV质粒编码的、参与致病过程的蛋白质Yops。小肠结肠炎耶尔森菌还表达两种外膜蛋白，一种名为YadA的黏附素和一种名为YIpA的脂蛋白。Yops的产生由质粒上一个20kb的区域协同调控，该区域被称为“钙依赖性区域”，包含至少四个基因座，即virA、virB、virC和virF。8.5kb的virC区域参与Yops的特异性转运，是一个单一操纵子，包含13个开放阅读框，称为yscA至yscM。基因virF编码yop、yadA和ylpA基因的关键转录激活因子。它仅在37°C转录，其表达受一种名为YmoA的染色体编码的类组蛋白调控。我们在此表明virF也控制virC操纵子。相比之下，VirF对于virA和virB的诱导并非必需。VirF蛋白特异性结合yop启动子。特别是，它保护yopH基因中跨越核苷酸 -64至 -34的区域。为了分析温度在yop调节子诱导中的作用，我们构建了从tac启动子表达virF的小肠结肠炎耶尔森菌菌株。尽管virF在25°C转录，但在该温度下Yops和YadA均未表达。在25°C时对VirF的这种不良反应至少部分是由于virF控制的基因转录微弱且缓慢。令人惊讶的是，当克隆到pACYC184上时，即使在没有VirF的情况下基因yadA也会表达，但仍受热依赖性调控。因此，温度和virF对于yop调节子的诱导都是必需的。在其他可能的作用中，温度可能会改变激活因子本身或yop启动子的结构。VirF在25°C时能在体外结合yop启动子这一事实排除了第一种假设。为了检验第二种假设，我们在体内研究了ymoA突变体中yopH启动子的活性。yopH启动子在没有VirF的情况下变得活跃，表明yop启动子活性取决于染色质结构。我们从这两个观察结果得出结论，在体内，需要温度来改变yop启动子的DNA结构，以便转录激活因子发挥作用。
