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Kim, Se Kye,Park, Yun Min,Jung, Kyoung Hwa,Chai, Young Gyu Microbiology Society 2017 Microbiology Vol.163 No.2
<P>Bacillus anthracis, an aetiologic agent of the zoonotic disease anthrax, encodes a putative NlpC/P60 endopeptidase BAS1812. It harbours a signal peptide, three bacterial SH3 domains and an NlpC/P60 family domain. Previous studies showed that BAS1812 is immunogenic in infected hosts and is a potential biomarker for anthrax treatment. To date, however, little information is known about its function and involvement in anthrax pathogenesis. Here we describe the phenotypic effect of BAS1812 deletion in B. anthracis Sterne strain. Transcriptional analysis showed that BAS1812 expression in a host-like environment was enhanced at the end of log phase, started to diminish after entry to stationary phase and increased again late in stationary phase. The constructed BAS1812 mutant showed impaired long-term survival in the stationary growth phase, less resilience to detergent, lesser endospore formation and delayed germination. The mutant also showed diminished ability to degrade peptidoglycan, but its ability to produce anthrax exotoxins was not affected. We hypothesize that BAS1812 is a cell wall hydrolase involved in biological activities related to maintaining cell wall integrity, sporulation and spore germination.</P>
Se Kye Kim,June Bong Lee,Dalmuri Han,Hyung Tae Lee,Seon Mi Wi,Jang Won Yoon 한국예방수의학회 2017 예방수의학회지 Vol.41 No.4
Turmeric is known for its ability to enhance immunity via anti-inflammatory and anti-oxidant effects. Salmonella enterica species contain a large number of pathogenic serotypes that are adapted to a broad range of vertebrate hosts. Our previous study revealed that bioprocessed polysaccharides from the liquid culture Lentinus edodes fungal mycelia containing turmeric (BPP-turmeric) is able to alter chicken macrophage responses and increases chick survival against Salmonella enterica infection. In this study, we examined the immunomodulatory effects of BPP-turmeric on the porcine macrophage 3D4/31 cell line infected with Salmonella enterica subsp. enterica serovar Choleraesuis (S. Choleraesuis) or S. Enteritidis. Our experimental analyses demonstrated that BPP-turmeric (i) does not alter phagocytic and killing activity of 3D4/31 against either Salmonella serotypes, but that it (ii) represses mRNA transcription of interleukin (IL)-6, IL-8, and tumor necrosis factor α in response to Salmonella infection. Collectively, these results imply that BPP-turmeric has an immunomodulatory effect that represses pro-inflammatory cytokine expression in porcine macrophages, suggesting that it may protect swine from salmonellosis via controlling Salmonella-induced hyperinflammation.
Late-Exponential Gene Expression in codY-Deficient Bacillus anthracis in a Host-Like Environment
Kim, Se Kye,Jung, Kyoung Hwa,Yoon, Sung Nyo,Kim, Yun Ki,Chai, Young Gyu Springer-Verlag 2016 Current microbiology Vol.73 No.5
<P>CodY is a pleiotropic regulator commonly found in Gram-positive bacteria and regulates various biological processes during the stringent response in a nutrient-limiting environment. CodY also participates in virulence factor expression in many low G+C Gram-positive pathogens, as observed in Bacillus anthracis. However, the mechanism by which B. anthracis CodY regulates metabolism and virulence factors in response to environmental changes is unclear. Here, we attempted to identify the link between CodY and B. anthracis regulation with codY-deficient and codY-overexpressing mutants using high-throughput transcriptional analysis. Growth pattern analyses of codY mutants in both rich and minimal media showed defects in early cell proliferation, with opposite patterns in the early stationary phase: CodY overexpression prolonged bacterial growth, whereas deletion inhibited growth. RNA sequencing of codY-deficient B. anthracis showed both positive and negative changes in the gene expression of proteases and virulence factors as well as genes related to stringent response-related metabolism and biosynthetic processing. We also found that changes in codY expression could alter virulence gene expression of B. anthracis, suggesting modes of regulation in its virulence in a CodY concentration-dependent manner. Collectively, we conclude from these results that CodY can both positively and negatively regulate its regulon via direct and/or indirect approaches, and that its mode of regulation may be concentration dependent.</P>
Kim, Se Kye,Park, Moon Kyoo,Kim, Sang Hoon,Oh, Kwang Gun,Jung, Kyoung Hwa,Hong, Chong-Hae,Yoon, Jang W.,Chai, Young Gyu Springer-Verlag 2014 Current microbiology Vol.69 No.4
<P>RelA and SpoT synthesize ppGpp, a key effector molecule that facilitates the adaptation of bacteria to nutrient starvation and other stresses, known as the stringent response. To investigate the role of Rsh Bant , a putative RelA/SpoT homolog (encoded by BAS4302) in Bacillus anthracis, we examined the alteration of the secretome profiles after the overexpression of a functional His-Rsh Bant protein in the B. anthracis strain Sterne at the stationary growth phase. In the ppGpp-deficient E. coli mutant strain CF1693, overexpression of Rsh Bant restored a ppGpp-dependent growth defect on minimal glucose media. The secretome profiles obtained using a two-dimensional electrophoresis (2-DE) analysis were altered by overexpression of Rsh Bant in B. anthracis. Among the 66 protein spots differentially expressed >1.5-fold, the 29 proteins were abundant for further identification using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS). Functional categorization of those proteins implicated their involvement in various biological activities. Taken together, our results imply that overexpression of a functional His-Rsh Bant can lead to the increased levels of intracellular ppGpp in B. anthracis, resulting in the significant changes in its secretome profiling. The stringent response-controlled proteins identified are likely useful as potential targets for serodiagnostic applications.</P>
Kim Se Kye,Lee Jun Bong,Yoon Jang Won 한국미생물학회 2022 The journal of microbiology Vol.60 No.9
The type VI secretion system (T6SS) is a novel secretion system found in many Gram-negative bacteria that plays a role in bacterial competition, virulence, and host immune evasion. The enterohemorrhagic Escherichia coli (EHEC) O157:H7 strain EDL933 has a single functional T6SS gene cluster. In this study, we attempted to characterize the transcriptional pattern of the T6SS effector gene Z0264 in EDL933. Transcriptional analyses showed that Z0264 and other T6SS genes were transcribed in vitro in a growth-phase-dependent manner, but Z0264 was not secreted in the rich medium. Using adapter- and radioactivity-free transcription start site analysis, we identified an overlapping divergent promoter between Z0264 and Z0265. A β-galactosidase assay with truncated promoter regions showed that the divergent promoter is functional. In addition, we demonstrated the role of H-NS as a repressor in the transcription of Z0264. Notably, the cDNA PCR assay showed that the mRNA transcript from the Z0264 promoter did not include the entire main T6SS cluster, suggesting segmented gene expression by multiple promoters in the T6SS cluster. In conclusion, we identified a divergent promoter for Z0264 located in the T6SS cluster of EDL933 and characterized its in vitro transcriptional activity during growth. Our findings provide insights and a preliminary understanding of the regulatory mechanisms underlying T6SS transcription.
Se Kye Kim,Jun Bong Lee,Hyung Tae Lee,Jang Won Yoon 대한수의학회 2024 Journal of Veterinary Science Vol.25 No.1
Background: Staphylococcus aureus and S. pseudintermedius are the major etiological agents of staphylococcal infections in humans, livestock, and companion animals. The misuse of antimicrobial drugs has led to the emergence of antimicrobial-resistant Staphylococcus spp., including methicillin-resistant S. aureus (MRSA) and methicillin-resistant S. pseudintermedius (MRSP). One novel therapeutic approach against MRSA and MRSP is a peptide nucleic acid (PNA) that can bind to the target nucleotide strands and block expression. Previously, two PNAs conjugated with cell-penetrating peptides (P-PNAs), antisense PNA (ASP)-cmk and ASP-deoD, targeting two essential genes in S. aureus, were constructed, and their antibacterial activities were analyzed. Objectives: This study analyzed the combined antibacterial effects of P-PNAs on S. aureus and S. pseudintermedius clinical isolates. Methods: S. aureus ATCC 29740 cells were treated simultaneously with serially diluted ASP-cmk and ASP-deoD, and the minimal inhibitory concentrations (MICs) were measured. The combined P-PNA mixture was then treated with S. aureus and S. pseudintermedius veterinary isolates at the determined MIC, and the antibacterial effect was examined. Results: The combined treatment of two P-PNAs showed higher antibacterial activity than the individual treatments. The MICs of two individual P-PNAs were 20 and 25 μM, whereas that of the combined treatment was 10 μM. The application of a combined treatment to clinical Staphylococcus spp. revealed S. aureus isolates to be resistant to P-PNAs and S. pseudintermedius isolates to be susceptible. Conclusions: These observations highlight the complexity of designing ASPs with high efficacy for potential applications in treating staphylococcal infections in humans and animals.