Iglesias Neves Henrique, Pereira Tuanny Fernanda, Yagil Ezra, Spira Beny
Departamento de Microbiologia, Instituto de Ciências Biomédicas, Universidade de São Paulo, São Paulo, SP, Brazil.
Department of Biochemistry and Molecular Biology, Tel-Aviv University, Tel-Aviv, Israel.
J Bacteriol. 2015 Apr;197(8):1378-85. doi: 10.1128/JB.02566-14. Epub 2015 Feb 2.
Mutations that cause the constitutive expression of the PHO regulon of Escherichia coli occur either in the pst operon or in the phoR gene, which encode, respectively, a high-affinity Pi transport system and a histidine kinase sensor protein. These mutations are normally selected on glycerol-2-phosphate (G2P) as the carbon source in the presence of excess Pi. The emergence of early PHO-constitutive mutants, which appear after growth for up to 48 h on selective medium, depends on the presence of phoA, which codes for a periplasmic alkaline phosphatase, while late mutants, which appear after 48 h, depend both on phoA and on the ugp operon, which encodes a glycerophosphodiester transport system. The emergence of the late mutants hints at an adaptive mutation process. PHO-constitutive phoR mutants appear only in a host that is mutated in pitA, which encodes an alternative Pi transport system that does not belong to the PHO regulon. The conserved Thr(217) residue in the PhoR protein is essential for PHO repression.
One of the principal ways in which bacteria adapt to new nutrient sources is by acquiring mutations in key regulatory genes. The inability of E. coli to grow on G2P as a carbon source is used to select mutations that derepress the PHO regulon, a system of genes involved in the uptake of phosphorus-containing molecules. Mutations in the pst operon or in phoR result in the constitutive expression of the entire PHO regulon, including alkaline phosphatase, which hydrolyzes G2P. Here we demonstrate that the ugp operon, another member of the PHO regulon, is important for the selection of PHO-constitutive mutants under prolonged nutritional stress and that phoR mutations can be selected only in bacteria lacking pitA, which encodes a secondary Pi transport system.
导致大肠杆菌PHO调节子组成型表达的突变发生在pst操纵子或phoR基因中，它们分别编码一个高亲和力的磷酸盐转运系统和一个组氨酸激酶传感器蛋白。这些突变通常在过量磷酸盐存在的情况下，以甘油-2-磷酸（G2P）作为碳源进行筛选。早期PHO组成型突变体在选择性培养基上生长长达48小时后出现，其出现依赖于编码周质碱性磷酸酶的phoA，而在48小时后出现的晚期突变体则同时依赖于phoA和编码甘油磷酸二酯转运系统的ugp操纵子。晚期突变体的出现暗示了一个适应性突变过程。PHO组成型phoR突变体仅出现在pitA发生突变的宿主中，pitA编码一种不属于PHO调节子的替代性磷酸盐转运系统。PhoR蛋白中保守的苏氨酸（217）残基对于PHO抑制至关重要。
细菌适应新营养源的主要方式之一是在关键调节基因中获得突变。利用大肠杆菌不能以G2P作为碳源生长这一特性来筛选解除PHO调节子抑制的突变，PHO调节子是一个参与含磷分子摄取的基因系统。pst操纵子或phoR中的突变导致整个PHO调节子的组成型表达，包括水解G2P的碱性磷酸酶。在这里，我们证明了PHO调节子的另一个成员ugp操纵子对于在长期营养应激下筛选PHO组成型突变体很重要，并且phoR突变只能在缺乏pitA的细菌中被筛选出来，pitA编码一种次要的磷酸盐转运系统。
