Journal of Microbiology

Review

Structural Insights into the Lipopolysaccharide Transport (Lpt) System as a Novel Antibiotic Target.: Yurim Yoon, Saemee Song; J. Microbiol. 2024;62(4):261-275. Published online May 31, 2024; DOI: https://doi.org/10.1007/s12275-024-00137-w

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Abstract: Lipopolysaccharide (LPS) is a critical component of the extracellular leaflet within the bacterial outer membrane, forming an effective physical barrier against environmental threats in Gram-negative bacteria. After LPS is synthesized and matured in the bacterial cytoplasm and the inner membrane (IM), LPS is inserted into the outer membrane (OM) through the ATP-driven LPS transport (Lpt) pathway, which is an energy-intensive process. A trans-envelope complex that contains seven Lpt proteins (LptA-LptG) is crucial for extracting LPS from the IM and transporting it across the periplasm to the OM. The last step in LPS transport involves the mediation of the LptDE complex, facilitating the insertion of LPS into the outer leaflet of the OM. As the Lpt system plays an essential role in maintaining the impermeability of the OM via LPS decoration, the interactions between these interconnected subunits, which are meticulously regulated, may be potential targets for the development of new antibiotics to combat multidrug-resistant Gram-negative bacteria. In this review, we aimed to provide an overview of current research concerning the structural interactions within the Lpt system and their implications to clarify the function and regulation of LPS transport in the overall process of OM biogenesis. Additionally, we explored studies on the development of therapeutic inhibitors of LPS transport, the factors that limit success, and future prospects.

Journal Articles

miR-135b Aggravates Fusobacterium nucleatum-Induced Cisplatin Resistance in Colorectal Cancer by Targeting KLF13: Wei Zeng , Jia Pan , Guannan Ye; J. Microbiol. 2024;62(2):63-73. Published online February 24, 2024; DOI: https://doi.org/10.1007/s12275-023-00100-1

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Abstract: Cisplatin resistance is the main cause of colorectal cancer (CRC) treatment failure, and the cause has been reported to be related to Fusobacterium nucleatum (Fn) infection. In this study, we explored the role of Fn in regulating cisplatin resistance of CRC cells and its underlying mechanism involved. The mRNA and protein expressions were examined by qRT-PCR and western blot. Cell proliferation and cell apoptosis were assessed using CCK8 and flow cytometry assays, respectively. Dual-luciferase reporter gene assay was adopted to analyze the molecular interactions. Herein, our results revealed that Fn abundance and miR-135b expression were markedly elevated in CRC tissues, with a favorable association between the two. Moreover, Fn infection could increase miR-135b expression via a concentration-dependent manner, and it also enhanced cell proliferation but reduced apoptosis and cisplatin sensitivity by upregulating miR-135b. Moreover, KLF13 was proved as a downstream target of miR-135b, of which overexpression greatly diminished the promoting effect of miR-135b or Fn-mediated cisplatin resistance in CRC cells. In addition, it was observed that upstream 2.5 kb fragment of miR-135b promoter could be interacted by β-catenin/TCF4 complex, which was proved as an effector signaling of Fn. LF3, a blocker of β-catenin/TCF4 complex, was confirmed to diminish the promoting role of Fn on miR-135b expression. Thus, it could be concluded that Fn activated miR-135b expression through TCF4/β-catenin complex, thereby inhibiting KLF13 expression and promoting cisplatin resistance in CRC.

Comparison of Conjunctival Sac Microbiome between Low and High Myopic Eyes: Kang Xiao , Zhengyu Chen , Qin Long; J. Microbiol. 2023;61(5):571-578. Published online April 21, 2023; DOI: https://doi.org/10.1007/s12275-023-00045-5

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Abstract: Microbial communities played a vital role in maintaining homeostasis of ocular surface. However, no studies explored the myopia-associated conjunctiva microbiota changes until now. In this study, conjunctival sac swab specimens were collected from 12 eyes of low myopia (LM), and 14 eyes of high myopia (HM) patients. The V3–V4 region of the 16S rRNA gene was amplified and then sequenced. Statistical analysis was performed to investigate differences in the taxonomy and diversity between two groups. Compared to LM, higher Ocular Surface Disease Index (OSDI) scores were observed in HM group. The Shannon index of the HM was lower than that of the LM group (P = 0.017). Principle coordinate analysis and Partial Least Squares Discrimination Analysis showed distinct microbiome composition between two groups. At the phylum level, there were higher relative abundances of Proteobacteria (68.27% vs 38.51%) and lower abundances of Actinobacteria (3.71% vs 9.19%) in HM, compared to LM group (P = 0.031, 0.010, respectively). At the genus level, the abundances of Acinetobacter in HM (18.16%) were significantly higher than the LM (6.52%) group (P = 0.011). Actinobacteria levels were negatively correlated with the myopic spherical equivalent and OSDI scores. Moreover, positive correlations were found between Proteobacteria levels and OSDI scores, Acinetobacter levels were positively correlated with myopic spherical equivalent and OSDI scores. In conclusion, HM Patients have bacterial microbiota imbalance in the conjunctival sac, compared with LM patients. Proteobacteria, Actinobacteria, Acinetobacter may play roles in the HM associated ocular surface irritation.

Review

Recent Advances in CRISPR‑Cas Technologies for Synthetic Biology: Song Hee Jeong , Ho Joung Lee , Sang Jun Lee; J. Microbiol. 2023;61(1):13-36. Published online February 1, 2023; DOI: https://doi.org/10.1007/s12275-022-00005-5

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5 Citations

Abstract: With developments in synthetic biology, “engineering biology” has emerged through standardization and platformization based on hierarchical, orthogonal, and modularized biological systems. Genome engineering is necessary to manufacture and design synthetic cells with desired functions by using bioparts obtained from sequence databases. Among various tools, the CRISPR-Cas system is modularly composed of guide RNA and Cas nuclease; therefore, it is convenient for editing the genome freely. Recently, various strategies have been developed to accurately edit the genome at a single nucleotide level. Furthermore, CRISPR-Cas technology has been extended to molecular diagnostics for nucleic acids and detection of pathogens, including disease-causing viruses. Moreover, CRISPR technology, which can precisely control the expression of specific genes in cells, is evolving to find the target of metabolic biotechnology. In this review, we summarize the status of various CRISPR technologies that can be applied to synthetic biology and discuss the development of synthetic biology combined with CRISPR technology in microbiology.

Journal Articles

[Protocol] Development of DNA aptamers specific for small therapeutic peptides using a modified SELEX method: Jaemin Lee , Minkyung Ryu , Dayeong Bae , Hong-Man Kim , Seong-il Eyun , Jeehyeon Bae , Kangseok Lee; J. Microbiol. 2022;60(7):659-667. Published online June 22, 2022; DOI: https://doi.org/10.1007/s12275-022-2235-4

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Abstract: Aptamers are short single-stranded DNA or RNA oligonucleotides capable of binding with high affinity and specificity to target molecules. Because of their durability and ease of synthesis, aptamers are used in a wide range of biomedical fields, including the diagnosis of diseases and targeted delivery of therapeutic agents. The aptamers were selected using a process called systematic evolution of ligands by exponential enrichment (SELEX), which has been improved for various research purposes since its development in 1990. In this protocol, we describe a modified SELEX method that rapidly produces high aptamer screening yields using two types of magnetic beads. Using this method, we isolated an aptamer that specifically binds to an antimicrobial peptide. We suggest that by conjugating a small therapeutic-specific aptamer to a gold nanoparticle-based delivery system, which enhances the stability and intracellular delivery of peptides, aptamers selected by our method can be used for the development of therapeutic agents utilizing small therapeutic peptides.

The efficacy of a 2,4-diaminoquinazoline compound as an intranasal vaccine adjuvant to protect against influenza A virus infection in vivo: Kyungseob Noh , Eun Ju Jeong , Timothy An , Jin Soo Shin , Hyejin Kim , Soo Bong Han , Meehyein Kim; J. Microbiol. 2022;60(5):550-559. Published online April 18, 2022; DOI: https://doi.org/10.1007/s12275-022-1661-7

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Abstract: Adjuvants are substances added to vaccines to enhance antigen- specific immune responses or to protect antigens from rapid elimination. As pattern recognition receptors, Toll-like receptors 7 (TLR7) and 8 (TLR8) activate the innate immune system by sensing endosomal single-stranded RNA of RNA viruses. Here, we investigated if a 2,4-diaminoquinazolinebased TLR7/8 agonist, (S)-3-((2-amino-8-fluoroquinazolin- 4-yl)amino)hexan-1-ol (named compound 31), could be used as an adjuvant to enhance the serological and mucosal immunity of an inactivated influenza A virus vaccine. The compound induced the production of proinflammatory cytokines in macrophages. In a dose-response analysis, intranasal administration of 1 μg compound 31 together with an inactivated vaccine (0.5 μg) to mice not only enhanced virus-specific IgG and IgA production but also neutralized influenza A virus with statistical significance. Notably, in a virus-challenge model, the combination of the vaccine and compound 31 alleviated viral infection-mediated loss of body weight and increased survival rates by 40% compared with vaccine only-treated mice. We suggest that compound 31 is a promising lead compound for developing mucosal vaccine adjuvants to protect against respiratory RNA viruses such as influenza viruses and potentially coronaviruses.

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