Both IIC and IID Components of Mannose Phosphotransferase System Are Involved in the Specific Recognition between Immunity Protein PedB and Bacteriocin-Receptor Complex

<div><p>Upon exposure to exogenous pediocin-like bacteriocins, immunity proteins specifically bind to the target receptor of the mannose phosphotransferase system components (man-PTS IIC and IID), therefore preventing bacterial cell death. However, the specific recognition of immunity proteins and its associated target receptors remains poorly understood. In this study, we constructed hybrid receptors to identify the domains of IIC and/or IID recognized by the immunity protein PedB, which confers immunity to pediocin PA-1. Using <i>Lactobacillus plantarum</i> man-PTS EII mutant W903, the IICD components of four pediocin PA-1-sensitive strains (<i>L</i>. <i>plantarum</i> WQ0815, <i>Leuconostoc mesenteroides</i> 05–43, <i>Lactobacillus salivarius</i> REN and <i>Lactobacillus acidophilus</i> 05–172) were respectively co-expressed with the immunity protein PedB. Well-diffusions assays showed that only the complex formed by LpIICD from <i>L</i>. <i>plantarum</i> WQ0815 with pediocin PA-1 could be recognized by PedB. In addition, a two-step PCR approach was used to construct hybrid receptors by combining LpIIC or LpIID recognized by PedB with the other three heterologous IID or IIC compounds unrecognized by PedB, respectively. The results showed that all six hybrid receptors were recognized by pediocin PA-1. However, when IIC or IID of <i>L</i>. <i>plantarum</i> WQ0815 was replaced with any corresponding IIC or IID component from <i>L</i>. <i>mesenteroides</i> 05–43, <i>L</i>. <i>salivarius</i> REN and <i>L</i>. <i>acidophilus</i> 05–172, all the hybrid receptors could not be recognized by PedB. Taken altogether, we concluded that both IIC and IID components of the mannose phosphotransferase system play an important role in the specific recognition between the bacteriocin-receptor complex and the immunity protein PedB.</p></div

Sensitivity of <i>L</i>. <i>plantarum</i> W903 derivatives harboring gene <i>IICD</i> or <i>IICD</i> and <i>pedB</i> to pediocin PA-1.

<p>Sensitivity of <i>L</i>. <i>plantarum</i> W903 derivatives harboring gene <i>IICD</i> or <i>IICD</i> and <i>pedB</i> to pediocin PA-1.</p

PCR primers for amplifying genes <i>IIC</i> and <i>IID</i>.

<p>PCR primers for amplifying genes <i>IIC</i> and <i>IID</i>.</p

Both IIC and IID Components of Mannose Phosphotransferase System Are Involved in the Specific Recognition between Immunity Protein PedB and Bacteriocin-Receptor Complex - Fig 4

<p><b>Multiple sequence alignments (A) and phylogenetic clustering (B) of IIC and IID proteins from <i>L</i>. <i>plantarum</i>, <i>L</i>. <i>mesenteroides</i>, <i>L</i>. <i>salivarius</i>, <i>L</i>. <i>acidophilus</i> and <i>P</i>. <i>acidilactici</i>.</b> Transmembrane helix (TMhelix), extracellular and intracellular regions were determined by using TMHMM v. 2.0 software. An asterisk, two dots, and one dot indicated decreasing degrees of conservation. The conserved motifs GGQGxxG and GG[D/K]FxxxG in the extracellular loop region are indicated by a grey background. The residues from <i>L</i>. <i>plantarum</i> and <i>P</i>. <i>acidilactici</i> in the intracellular regions are indicated by boxes. Sequence alignments and phylogenetic trees were constructed by using MUSCLE v. 3.8.31 software with default settings (<a href="http://www.ebi.ac.uk/Tools/msa/muscle/" target="_blank">http://www.ebi.ac.uk/Tools/msa/muscle/</a>) [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0164973#pone.0164973.ref027" target="_blank">27</a>].</p

Administration of L9 ameliorated AHR enhanced by exposure of PM_2.5.

<p>(A) AHR to increasing doses of Mch. (B) The maximal response to Mch challenge. Each value is expressed as mean ± SEM. n = 5–8. ## <i>P</i> < 0.005, ### <i>P</i> < 0.001 vs. OVA+PM_2.5.</p

Experimental setup of the PM_2.5 exposure enhanced mouse model of asthma.

<p>Experimental setup of the PM_2.5 exposure enhanced mouse model of asthma.</p

Analysis of Metal and PAH composition in PM_2.5.

<p>Analysis of Metal and PAH composition in PM_2.5.</p

Outline of the cloning procedure.

<p>Outline of the cloning procedure.</p

Oral administration of <i>Lactobacillus paracasei</i> L9 attenuates PM_2.5-induced enhancement of airway hyperresponsiveness and allergic airway response in murine model of asthma

<div><p>This study investigated allergy immunotherapy potential of <i>Lactobacillus paracasei</i> L9 to prevent or mitigate the particulate matter 2.5 (PM_2.5) enhanced pre-existing asthma in mice. Firstly, we used a mouse model of asthma (a 21-day ovalbumin (OVA) sensitization and challenge model) followed by PM_2.5 exposure twice on the same day of the last challenge. PM_2.5 was collected from the urban area of Beijing and underwent analysis for metals and polycyclic aromatic hydrocarbon contents. The results showed that PM_2.5 exposure enhanced airway hyper-responsiveness (AHR) and lead to a mixed Th2/ IL-17 response in asthmatic mice. Secondly, the PM_2.5 exposed asthmatic mice were orally administered with L9 (4×10⁷, 4×10⁹ CFU/mouse, day) from the day of first sensitization to the endpoint, for 20 days, to investigate the potential mitigative effect of L9 on asthma. The results showed that L9 ameliorated PM_2.5 exposure enhanced AHR with an approximate 50% decrease in total airway resistance response to methacholine (48 mg/ml). L9 also prevented the exacerbated eosinophil and neutrophil infiltration in bronchoalveolar lavage fluid (BALF), and decreased the serum level of total IgE and OVA-specific IgG1 by 0.44-fold and 0.3-fold, respectively. Additionally, cytokine production showed that L9 significantly decreased T-helper cell type 2 (Th2)–related cytokines (IL-4, -5, -13) and elevated levels of Th1 related IFN-γ in BALF. L9 also reduced the level of IL-17A and increased the level of TGF-β. Taken together, these results indicate that L9 may exert the anti-allergic benefit, possibly through rebalancing Th1/Th2 immune response and modulating IL-17 pro-inflammatory immune response. Thus, L9 is a promising candidate for preventing PM exposure enhanced pre-existing asthma.</p></div

PM_2.5 exposure exacerbated airway inflammatory cell infiltration in mice with OVA induced pre-existing asthma.

<p>(A) Histopathological examination of lung tissue inflammatory cell infiltration in mice. Representative photos of H&E-stained lung sections (original magnification 20x). (B) Cell population of macrophage, eosinophil, neutrophil and lymphocyte in BALF. Each value is expressed as mean ± SEM. n = 5–8. * <i>P</i>< 0.05, ** <i>P</i> < 0.005, *** <i>P</i> < 0.001 vs. CON; ⁺ <i>P</i> < 0.05 vs. OVA+FILTER.</p