CtsR, the Master Regulator of Stress-Response in Oenococcus oeni, Is a Heat Sensor Interacting With ClpL1

Abstract

Oenococcus oeni is a lactic acid bacterium responsible for malolactic fermentation of wine. While many stress response mechanisms implemented by O. oeni during wine adaptation have been described, little is known about their regulation. CtsR is the only regulator of stress response genes identified to date in O. oeni. Extensively characterized in Bacillus subtilis, the CtsR repressor is active as a dimer at 37°C and degraded at higher temperatures by a proteolytic mechanism involving two adapter proteins, McsA and McsB, together with the ClpCP complex. The O. oeni genome does not encode orthologs of these adapter proteins and the regulation of CtsR activity remains unknown. In this study, we investigate CtsR function in O. oeni by using antisense RNA silencing in vivo to modulate ctsR gene expression. Inhibition of ctsR gene expression by asRNA leads to a significant loss in cultivability after heat shock (58%) and acid shock (59%) highlighting the key role of CtsR in the O. oeni stress response. Regulation of CtsR activity was studied using a heterologous expression system to demonstrate that O. oeni CtsR controls expression and stress induction of the O. oeni hsp18 gene when produced in a ctsR-deficient B. subtilis strain. Under heat stress conditions, O. oeni CtsR acts as a temperature sensor and is inactivated at growth temperatures above 33°C. Finally, using an E. coli bacterial two-hybrid system, we showed that CtsR and ClpL1 interact, suggesting a key role for ClpL1 in controlling CtsR activity in O. oeni.

Effect of High Hydrostatic Pressure on Stress-Related dnaK, hrcA, and ctsR Expression Patterns in Selected Lactobacilli Strains

“…It has been reported that the ctsR mutant L. plantarum strain was more resistant to oxidative stress and more sensitive to ethanol and heat stresses than its wild strain, which most likely is related to the pleiotropic character of the gene transcription [11,14]. A possible explanation of these results might be associated with the complexity of the stress-related mechanisms in LAB strains [24], which we also visualized in PPI network analysis. Many transcriptional regulators, such as HrcA, CtsR, Fur, MarR, and MeR family are differentially expressed in response to the heat shock [25,26].…”

Heterologous Expression of Argininosuccinate Synthase From Oenococcus oeni Enhances the Acid Resistance of Lactobacillus plantarum

“…The citrate metabolism affects the acid tolerance of O. oeni , owing to its end products (Augagneur et al, 2007), with Margalef-Català et al (2016) finding that the genes involved in glutamine and glutamate metabolism were upregualted of O. oeni under stress. Darsonval et al (2018) verified that the CtsR is the master regulator of stress-response in O. oeni . In addition, the arginine catabolism through the arginine deiminase (ADI) pathway is known to enhance acid tolerance by converting arginine into an alkaline product and by increasing external pH (Tonon et al, 2001; Arena and Manca de Nadra, 2005; Araque et al, 2016).…”

Oenococcus oeni, a species born and moulded in wine: a critical review of the stress impacts of wine and the physiological responses

“…Based on the results of Constantini et al (2015) and their own results, Margalef-Català et al (2016a) suggest that translational regulation is an important factor in O. oeni based on the induction of ribosomal proteins. Posttranscriptional regulation has been observed for ClpL (Beltramo et al 2004) and in a recent study, authors found that CtsR and ClpL1 interact, suggesting that ClpL1 has an important role in regulation (Darsonval et al 2018). In this study, they demonstrated that CtsR is a heat-sensor which becomes inactivated above 33 C. The heterologous expression of ctsR into L. plantarum conferred significant acidethanol resistance, which increased the expression of ctsR, clp and hsp genes (Zhao et al 2019b), which could be due to a high sensitivity and high response of the O. oeni CtsR to stress.…”

“…Based on the results of Constantini et al () and their own results, Margalef‐Català et al () suggest that translational regulation is an important factor in O. oeni based on the induction of ribosomal proteins. Post‐transcriptional regulation has been observed for ClpL (Beltramo et al ) and in a recent study, authors found that CtsR and ClpL1 interact, suggesting that ClpL1 has an important role in regulation (Darsonval et al ). In this study, they demonstrated that CtsR is a heat‐sensor which becomes inactivated above 33°C.…”

“…It is expected that higher temperature will significantly expatiate the effect of ethanol stress, as it does for yeast (Piper 1995), and since they both have an effect on membrane fluidity (Tourdot-Maréchal et al 2000). The recent study showing that at least some O. oeni have a strong heat-stress response at 33 C (Darsonval et al 2018) indicates the potentially significant implication that temperature around 30-35 C can have, in particular among other stressors. With the implication of global climate change leading to rising temperature, the high temperature around MLF as well as the increased concentration of ethanol in the final wines (Jones et al 2005), this subject will be of increasing importance (Mozell and Thac 2014).…”