Department of Microbiology and Molecular Genetics, McGovern Medical School, The University of Texas Health Science Center at Houston, Houston, Texas, USA.
MD Anderson Cancer Center UTHealth Graduate School of Biomedical Sciences, Houston, Texas, USA.
J Bacteriol. 2022 Sep 20;204(9):e0020822. doi: 10.1128/jb.00208-22. Epub 2022 Aug 25.
The Bacillus anthracis gene, encoding the protective antigen component of anthrax toxin, is part of a bicistronic operon on pXO1 that codes for its own repressor, PagR1. In addition to the operon, PagR1 regulates and , two chromosome genes encoding components of the surface layer, a mounting structure for surface proteins involved in virulence. Genomic studies have revealed a PagR1 paralog, PagR2, encoded by a gene on pXO2. The amino acid sequences of the paralogues are 71% identical and show similarity to the ArsR family of transcription regulators. We determined that the expression of either rPagR1 or rPagR2 in a Δ pXO1/pXO2 (PagR1-PagR2) background repressed the expression of , , , and a newly discovered target, encoding virulence activator AtxA. Despite the redundancy in PagR1 and PagR2 function, we determined that purified rPagR1 bound DNA corresponding to the control regions of all four target genes and existed as a dimer in cell lysates, whereas rPagR2 exhibited weak binding to the DNA of the and promoters, did not bind or promoter DNA, and did not appear as a dimer in cell lysates. A single amino acid change in PagR2, S81Y, designed to match the native Y81 of PagR1, allowed for DNA-binding to the and promoters. Moreover, the S81Y mutation allowed for the detection of PagR2 homomultimers in coaffinity purification experiments. Our results expand our knowledge of the roles of the paralogues in B. anthracis gene expression and provide a potential mechanistic basis for differences in the functions of these repressors. The protective antigen component of the anthrax toxin is essential for the delivery of the enzymatic components of the toxin into host target cells. The toxin genes and other virulence genes of B. anthracis are regulated by multiple -acting regulators that respond to a variety of host-related signals. PagR1, one such -acting regulator, connects the regulation of plasmid-encoded and chromosome-encoded virulence genes by controlling both protective antigen and surface layer protein expression. Whether PagR2, a paralog of PagR1, also functions as a -acting regulator was unknown. This work advances our knowledge of the complex model of virulence regulation in B. anthracis and furthers our understanding of the intriguing evolution of this pathogen.
炭疽杆菌基因，编码炭疽毒素的保护性抗原成分，是编码其自身抑制剂 PagR1 的 pXO1 上双顺反子操纵子的一部分。除了操纵子，PagR1 还调节和，这两个染色体基因编码表面层的组成部分，这是一种与毒力有关的表面蛋白的安装结构。基因组研究揭示了 PagR1 的一个旁系同源物 PagR2，由 pXO2 上的一个基因编码。旁系同源物的氨基酸序列有 71%的同一性，并显示出与 ArsR 家族转录调节剂的相似性。我们确定，在 ΔpXO1/pXO2（PagR1-PagR2）背景下表达 rPagR1 或 rPagR2 抑制了的表达、、、和一个新发现的靶标，编码毒力激活剂 AtxA。尽管 PagR1 和 PagR2 功能存在冗余，但我们确定纯化的 rPagR1 结合了对应于所有四个靶基因控制区的 DNA，并且在细胞裂解物中以二聚体形式存在，而 rPagR2 对和启动子的 DNA 结合较弱，不结合或启动子 DNA，并且在细胞裂解物中不以二聚体形式存在。PagR2 中的单个氨基酸变化 S81Y，旨在与 PagR1 的天然 Y81 匹配，允许 DNA 结合到和启动子。此外，S81Y 突变允许在共亲和纯化实验中检测到 PagR2 同源多聚体。我们的结果扩展了我们对炭疽杆菌基因表达中旁系同源物作用的认识，并为这些抑制剂功能差异提供了潜在的机制基础。炭疽毒素的保护性抗原成分对于将毒素的酶成分递送到宿主靶细胞中是必不可少的。炭疽芽孢杆菌的毒素基因和其他毒力基因受多种多效调节因子调控，这些调节因子对多种宿主相关信号作出反应。PagR1 是这样的一种多效调节因子，通过控制保护性抗原和表面层蛋白的表达来连接质粒编码和染色体编码的毒力基因的调节。PagR1 的旁系同源物 PagR2 是否也作为一种多效调节因子发挥作用尚不清楚。这项工作推进了我们对炭疽杆菌复杂毒力调节模型的认识，并加深了我们对这种病原体引人入胜的进化的理解。
