Expression of the pyr Operon of Lactobacillus plantarum Is Regulated by Inorganic Carbon Availability through a Second Regulator, PyrR 2 , Homologous to the Pyrimidine-Dependent Regulator PyrR 1

Abstract

ABSTRACT Inorganic carbon (IC), such as bicarbonate or carbon dioxide, stimulates the growth of Lactobacillus plantarum . At low IC levels, one-third of natural isolated L. plantarum strains are nutritionally dependent on exogenous arginine and pyrimidine, a phenotype previously defined as high-CO 2 -requiring (HCR) prototrophy. IC enrichment significantly decreased the amounts of the enzymes in the pyrimidine biosynthetic pathway encoded by the pyrR 1 BCAa 1 Ab 1 DFE operon, as demonstrated by proteomic analysis. Northern blot and reverse transcription-PCR experiments demonstrated that IC levels regulated pyr genes mainly at the level of transcription or RNA stability. Two putative PyrR regulators with 62% amino acid identity are present in the L. plantarum genome. PyrR 1 is an RNA-binding protein that regulates the pyr genes in response to pyrimidine availability by a mechanism of transcriptional attenuation. In this work, the role of PyrR 2 was investigated by allelic gene replacement. Unlike the pyrR 1 mutant, the ΔpyrR 2 strain acquired a demand for both pyrimidines and arginine unless bicarbonate or CO 2 was present at high concentrations, which is known as an HCR phenotype. Analysis of the IC- and pyrimidine-mediated regulation in pyrR 1 and pyrR 2 mutants suggested that only PyrR 2 positively regulates the expression levels of the pyr genes in response to IC levels but had no effect on pyrimidine-mediated repression. A model is proposed for the respective roles of PyrR 1 and PyrR 2 in the pyr regulon expression.