title: Theoretical insight into the heat shock response (HSR) regulation in Lactobacillus casei and L. rhamnosus.
abstract: The understanding of the heat shock response (HSR) in lactobacilli from a regulatory point of view is still limited, though an increased knowledge on the regulation of this central stress response can lead to improvements in the exploitation of these health promoting microorganisms. Therefore the aim of this in silico study, that is the first to be carried out for members of the Lactobacillus genus, was predicting how HSR influences cell functions in the food associated and probiotic species Lactobacillus casei and Lactobacillus rhamnosus. To this purpose, thirteen whole genomes of these bacteria were analyzed to identify which genes involved in HSR are present. It was found that all the genomes share 25 HSR related genes, including those encoding protein repair systems, HSR repressors, HrcA and CtsR, and the positive regulators of HSR, alternative σ factors σ(32) and σ(24). Two genes encoding a σ(70)/σ(24) factor and a Lon protease, respectively, were found only in some genomes. The localization of the HSR regulators binding sites in genomes was analyzed in order to identify regulatory relationships driving HSR in these lactobacilli. It was observed that the binding site for the HrcA repressor is found upstream of the hrcA-grpE-dnaK-dnaJ and groES-groEL gene clusters, of two hsp genes, clpE, clpL and clpP, while the CtsR repressor binding site precedes the ctsR-clpC operon, clpB, clpE and clpP. Therefore the ClpE-ClpP protease complex is dually regulated by HrcA and CtsR. Consensus sequences for the promoters recognized by the HSR alternative σ factors were defined for L. casei and L. rhamnosus and were used in whole genome searches to identify the genes that are possibly regulated by these transcription factors and whose expression level is expected to increases in HSR. The results were validated by applying the same procedure of promoter consensus generation and whole genome search to an additional 11 species representative of the main Lactobacillus lineages. The composition of the resulting regulons highlighted the existence of relationships between HSR and relevant cell functions, including nutrient utilization, DNA repair, protein synthesis and export of toxic substances. In fact, some of the predicted members of the σ(32) regulon are central regulators ccpA, spxA, cadA, and functional proteins brnQ, ldh, choS, poxL and nagB involved in the tolerance to different stress factors. The analysis of the expression level of these molecular markers of cell protective mechanisms can be used to select the heat shock exposure times and temperatures that maximize the tolerance of L. casei and L. rhamnosus to technological and environmental stress factors.
authors.affiliation: Biotechnology Department, University of Verona, 37134 Verona, Italy. Electronic address: franca.rossi@univr.it.
authors.forename: Franca
authors.initials: F
authors.lastname: Rossi
authors.affiliation: Institute of Food Science (ISA), National Research Council, via Roma, 64, 83100 Avellino, Italy. Electronic address: teresa.zotta@isa.cnr.it.
authors.forename: Teresa
authors.initials: T
authors.lastname: Zotta
authors.affiliation: Department of Food Science, University of Udine, via Sondrio 2/a, 33100 Udine, Italy. Electronic address: lucilla.iacumin@uniud.it.
authors.forename: Lucilla
authors.initials: L
authors.lastname: Iacumin
authors.affiliation: Institute of Food Science (ISA), National Research Council, via Roma, 64, 83100 Avellino, Italy. Electronic address: anna.reale@isa.cnr.it.
authors.forename: Anna
authors.initials: A
authors.lastname: Reale
mesh_terms: D000251:Adenosine Triphosphatases / Q000378:metabolism; D001483:Base Sequence; D016384:Consensus Sequence / Q000235:genetics; D015964:Gene Expression Regulation, Bacterial; D005798:Genes, Bacterial; D018869:Heat-Shock Response / Q000235:genetics*; D007780:Lacticaseibacillus casei / Q000235:genetics*; D052201:Lacticaseibacillus rhamnosus / Q000235:genetics*; D005810:Multigene Family; D011401:Promoter Regions, Genetic; D018085:Regulon / Q000235:genetics; D012808:Sigma Factor / Q000378:metabolism; D013312:Stress, Physiological / Q000235:genetics
chemical_list: D012808:Sigma Factor; D000251:Adenosine Triphosphatases
keywords: CtsR; HSR regulation; HrcA; Promoter consensus; Regulon composition; Stress tolerance; σ(24); σ(32)