Lactobacillus brevis responds to flavonoids through KaeR, a LysR‐type of transcriptional regulator

• Published in: Molecular microbiology 2011-09, Vol.81 (6), p.1623-1639
• Main Authors: Pande, Santosh G., Pagliai, Fernando A., Gardner, Christopher L., Wrench, Algevis, Narvel, Raed, Gonzalez, Claudio F., Lorca, Graciela L.
• Format: Article
• Language: English
• Subjects: Artificial Gene Fusion; Bacillus subtilis; Bacillus subtilis - drug effects; Bacillus subtilis - genetics; Bacillus subtilis - metabolism; Bacterial Proteins - metabolism; Bacteriology; Base Sequence; beta-Galactosidase - genetics; beta-Galactosidase - metabolism; Binding sites; Biological and medical sciences; Deoxyribonuclease I - metabolism; DNA Footprinting; DNA, Bacterial - genetics; DNA, Intergenic; Electrophoretic Mobility Shift Assay; Fundamental and applied biological sciences. Psychology; Gene expression; Gene Expression Profiling; Gene Expression Regulation, Bacterial; Genes, Reporter; Kaempferols - metabolism; Lactobacillus brevis; Lactobacillus brevis - drug effects; Lactobacillus brevis - genetics; Lactobacillus brevis - metabolism; Microbiology; Miscellaneous; Molecular Sequence Data; Molecules; Phytochemicals; Protein Binding; Real-Time Polymerase Chain Reaction; Transcription Factors - metabolism
• ISSN: 0950-382X; 1365-2958
• DOI: 10.1111/j.1365-2958.2011.07796.x

Summary The ability of transcription factors to respond to flavonoids as signal molecules was investigated in Lactobacillus brevis. Through in vitro screening of a small library of flavonoids, LVIS1989 (KaeR), a LysR‐type transcriptional regulator (LTTR), was identified as responsive to kaempferol. The modulation of KaeR activity by flavonoids was characterized in vivo and in vitro. DNase I footprint assays identified the binding of KaeR at two distinctive sites, one in the intergenic region between LVIS1988 and kaeR (−39 to +2) and another within LVIS1988 (−314 to −353, from kaeR translational start point). EMSA assays revealed that both binding sites are required for KaeR binding in vitro. Furthermore, KaeR–DNA interactions were stabilized by the addition of kaempferol (20 µM). In vivo qRT‐PCR experiments performed in L. brevis confirmed that the divergently transcribed genes LVIS1988, LVIS1987 and LVIS1986 and kaeR are upregulated in the presence of kaempferol, indicating the role of KaeR as a transcriptional activator. Transcriptional lacZ fusions using Bacillus subtilis as a surrogate host showed that expression of kaeR and LVIS1988 were induced by the presence of the flavonoid. These results indicate that KaeR belongs to a small and poorly understood group of LTTRs that are positively autoregulated in the presence of a ligand.