AUTHOR=Wei Yarui , Dong Baozhu , Wu Xiaogang , Zhao Mingmin , Wang Dong , Li Na , Zhang Qian , Zhang Liqun , Zhou Hongyou 

TITLE=RpoZ regulates 2,4-DAPG production and quorum sensing system in Pseudomonas fluorescens 2P24

JOURNAL=Frontiers in Microbiology

VOLUME=14

YEAR=2023

URL=https://www.frontiersin.org/journals/microbiology/articles/10.3389/fmicb.2023.1160913

DOI=10.3389/fmicb.2023.1160913

ISSN=1664-302X

ABSTRACT=<sec><title>Introduction</title><p><italic>Pseudomonas fluorescens</italic> 2P24 was isolated from soil of natural decay associated with wheat take-all and it can effectively control soil-borne diseases caused by a variety of plant pathogens. 2,4-diacetylphloroglucinol (2,4-DAPG), is produced by <italic>P. fluorescens</italic> 2P24 and plays an important role in the prevention and control of plant diseases. To understand the resistant mechanism, in this study, we conducted experiments to explore the regulation role of <italic>rpoZ</italic> in the synthesis of the antibiotic 2,4-DAPG and regulation of QS system.</p></sec><sec><title>Methods</title><p>A random mini-Tn5 mutagenesis procedure was used to screen regulators for <italic>phlA</italic> transcription in stain PM901, which containing a phlA∷lacZ transcriptional fusion reporter plasmid. We identified 12 insertion mutants could significantly change <italic>phlA</italic> gene expression. By analyzing the amino acid sequences of the interrupted gene, we obtained a mutant strain Aa4-29 destroyed the <italic>rpoZ</italic> gene, which encodes the omiga subunit. We constructed the plasmid of <italic>rpoZ</italic> mutant (pBBR-△rpoZ) transformed into competent cells of <italic>P. fluorescens</italic> 2P24 by electro-transformation assay. The strains of <italic>P. fluorescens</italic> 2P24/pBBR, 2P24-△rpoZ/pBBR, 2P24-△rpoZ/pBBR-rpoZ were used to evaluate the regulation role of <italic>rpoZ</italic> in 2,4-DAPG production and quorum sensing system.</p></sec><sec><title>Results</title><p>According to β-galactosidase activity, we found that <italic>rpoZ</italic> positively regulated the expression of <italic>phlA</italic> (a synthesis gene of 2,4-DAPG) and <italic>PcoI</italic> (a synthesis gene of PcoI/PcoR QS signal system) at the transcriptional level. The production of 2,4-DAPG antibiotic and signal molecule AHL was influenced by <italic>rpoZ</italic>. Further, <italic>rpoZ</italic> was involved in regulating <italic>rsmA</italic> expression. <italic>RpoZ</italic> also has a certain regulatory effect on <italic>rpoS</italic> transcription, but no effect on the transcription of <italic>phlF</italic>, <italic>emhABC</italic> and <italic>emhR</italic>. According to the biocontrol assay, <italic>P. fluorescens</italic> 2P24 strains with <italic>rpoZ</italic> showed obvious antagonism ability against the <italic>Rhizoctonia solani</italic> in cotton, while the mutant strain of <italic>rpoZ</italic> lost the biocontrol effect. <italic>RpoZ</italic> had a significant effect on the swimming and biofilm formation in <italic>P. fluorescens</italic> 2P24.</p></sec><sec><title>Conclusion</title><p>Our data showed that <italic>rpoZ</italic> was an important regulator of QS system, 2,4-DAPG in <italic>P. fluorescens</italic> 2P24. This may imply that <italic>P. fluorescens 2P24</italic> has evolved different regulatory features to adapt to different environmental threats.</p></sec>