Abstract
In vitro inhibition of the fungal pathogen Sclerotinia sclerotiorum by Pseudomonas chlororaphis PA23 is reliant upon a LysR-type transcriptional regulator (LTTR) called PtrA. In the current study, we show that Sclerotinia stem rot and leaf infection are significantly increased in canola plants inoculated with the ptrA-mutant compared to the wild type, establishing PtrA as an essential regulator of PA23 biocontrol. LTTRs typically regulate targets that are upstream of and divergently transcribed from the LTTR locus. We identified a short chain dehydrogenase (scd) gene immediately upstream of ptrA. Characterization of a scd mutant revealed that it is phenotypically identical to the wild type. Moreover, scd transcript abundance was unchanged in the ptrA mutant. These findings indicate that PtrA regulation does not involve scd, rather this LTTR controls genes located elsewhere on the chromosome. Employing a combination of complementation and transcriptional analysis we investigated whether connections exist between PtrA and other regulators of biocontrol. Besides ptrA, gacS was the only gene able to partially rescue the wild-type phenotype, establishing a connection between PtrA and the sensor kinase GacS. Transcriptomic analysis revealed decreased expression of biosynthetic (phzA, prnA) and regulatory genes (phzI, phzR, rpoS, gacA, rsmX, rsmZ, retS) in the ptrA mutant; conversely, rsmE, and rsmY were markedly upregulated. The transcript abundance of ptrA was nine-fold higher in the mutant background indicating that this LTTR negatively autoregulates itself. In summary, PtrA is an essential regulator of genes required for PA23 biocontrol that is functionally intertwined with GacS.
Highlights
Public concern over the use of chemical pesticides together with the potential for acquiring resistance to these compounds has led to renewed interest in alternative strategies for management of diseases affecting plants
LysR-type transcriptional regulator (LTTR), which represent the largest family of prokaryotic transcriptional regulators, frequently regulate divergently transcribed genes; targets can be located elsewhere on the chromosome (Schell, 1993; Maddocks and Oyston, 2008)
59 differentially regulated proteins distributed across 16 different COG categories were identified in the ptrA mutant (Klaponski et al, 2014) and genes encoding these proteins are scattered about the chromosome (Loewen et al, 2014)
Summary
Public concern over the use of chemical pesticides together with the potential for acquiring resistance to these compounds has led to renewed interest in alternative strategies for management of diseases affecting plants. Strain PA23 produces an arsenal of compounds including the diffusible antibiotics phenazine 1-carboxylic acid (PCA), 2hydroxyphenazine (2-OH-PHZ), and pyrrolnitrin (PRN) together with degradative enzymes. PRN is the primary antibiotic responsible for biocontrol; phenazines (PHZ) are not essential for fungal suppression, but do play a role in biofilm formation (Selin et al, 2010). Variable expression of genes and gene products required for biocontrol likely contributes to poor performance in the field. It is essential, that molecular mechanisms underlying biocontrol are well understood so that production of the pathogen-suppressive factors can be optimized in the environment
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