Journal of Microbiology

Journal Articles

Heterologous Production and Structure Determination of a New Lanthipeptide Sinosporapeptin Using a Cryptic Gene Cluster in an Actinobacterium Sinosporangium siamense: Keita Saito , Keiichiro Mukai , Issara Kaweewan , Hiroyuki Nakagawa , Takeshi Hosaka , Shinya Kodani; J. Microbiol. 2023;61(6):641-648. Published online June 12, 2023; DOI: https://doi.org/10.1007/s12275-023-00059-z

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Abstract: Lipolanthine is a subclass of lanthipeptide that has the modification of lipid moiety at the N-terminus. A cryptic biosynthetic gene cluster comprising four genes (sinA, sinKC, sinD, and sinE) involved in the biosynthesis of lipolanthine was identified in the genome of an actinobacterium Sinosporangium siamense. Heterologous coexpression of a precursor peptide coding gene sinA and lanthipeptide synthetase coding gene sinKC in the host Escherichia coli strain BL21(DE3) resulted in the synthesis of a new lanthipeptide, sinosporapeptin. It contained unusual amino acids, including one labionin and two dehydrobutyrine residues, as determined using NMR and MS analyses. Another coexpression experiment with two additional genes of decarboxylase (sinD) and N-acetyl transferase (sinE) resulted in the production of a lipolanthine-like modified sinosporapeptin.

Transcriptome‑based Mining of the Constitutive Promoters for Tuning Gene Expression in Aspergillus oryzae: Kobkul Laoteng , Jutamas Anantayanon , Chanikul Chutrakul , Sarocha Panchanawaporn , Sukanya Jeennor; J. Microbiol. 2023;61(2):199-210. Published online February 6, 2023; DOI: https://doi.org/10.1007/s12275-023-00020-0

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Abstract: Transcriptional regulation has been adopted for developing metabolic engineering tools. The regulatory promoter is a crucial genetic element for strain optimization. In this study, a gene set of Aspergillus oryzae with highly constitutive expression across different growth stages was identified through transcriptome data analysis. The candidate promoters were functionally characterized in A. oryzae by transcriptional control of β-glucuronidase (GUS) as a reporter. The results showed that the glyceraldehyde triphosphate dehydrogenase promoter (PgpdA1) of A. oryzae with a unique structure displayed the most robust strength in constitutively controlling the expression compared to the PgpdA2 and other putative promoters tested. In addition, the ubiquitin promoter (Pubi) of A. oryzae exhibited a moderate expression strength. The deletion analysis revealed that the 5' untranslated regions of gpdA1 and ubi with the length of 1028 and 811 nucleotides, counted from the putative translation start site (ATG), respectively, could efficiently drive the GUS expression. Interestingly, both promoters could function on various carbon sources for cell growth. Glucose was the best fermentable carbon source for allocating high constitutive expressions during cell growth, and the high concentrations (6–8% glucose, w/v) did not repress their functions. It was also demonstrated that the secondary metabolite gene coding for indigoidine could express under the control of PgpdA1 or Pubi promoter. These strong and moderate promoters of A. oryzae provided beneficial options in tuning the transcriptional expression for leveraging the metabolic control towards the targeted products.

Zinc-binding domain mediates pleiotropic functions of Yvh1 in Cryptococcus neoformans: Jae-Hyung Jin , Myung Kyung Choi , Hyun-Soo Cho , Yong-Sun Bahn; J. Microbiol. 2021;59(7):658-665. Published online July 1, 2021; DOI: https://doi.org/10.1007/s12275-021-1287-1

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Abstract: Yvh1 is a dual-specificity phosphatase (DUSP) that is evolutionarily conserved in eukaryotes, including yeasts and humans. Yvh1 is involved in the vegetative growth, differentiation, and virulence of animal and plant fungal pathogens. All Yvh1 orthologs have a conserved DUSP catalytic domain at the N-terminus and a zinc-binding (ZB) domain with two zinc fingers (ZFs) at the C-terminus. Although the DUSP domain is implicated in the regulation of MAPK signaling in humans, only the ZB domain is essential for most cellular functions of Yvh1 in fungi. This study aimed to analyze the functions of the DUSP and ZB domains of Yvh1 in the human fungal pathogen Cryptococcus neoformans, whose Yvh1 (CnYvh1) contains a DUSP domain at the C-terminus and a ZB domain at the N-terminus. Notably, CnYvh1 has an extended internal domain between the two ZF motifs in the ZB domain. To elucidate the function of each domain, we constructed individual domain deletions and swapping strains by complementing the yvh1Δ mutant with wild-type (WT) or mutated YVH1 alleles and examined their Yvh1-dependent phenotypes, including growth under varying stress conditions, mating, and virulence factor production. Here, we found that the complementation of the yvh1Δ mutant with the mutated YVH1 alleles having two ZFs of the ZB domain, but not the DUSP and extended internal domains, restored the WT phenotypic traits in the yvh1Δ mutant. In conclusion, the ZB domain, but not the N-terminal DUSP domain, plays a pivotal role in the pathobiological functions of cryptococcal Yvh1.

Extracellular products-mediated interspecific interaction between Pseudomonas aeruginosa and Escherichia coli: Yang Yuan , Jing Li , Jiafu Lin , Wenjuan Pan , Yiwen Chu , Balakrishnan Prithiviraj , Yidong Guo , Xinrong Wang , Kelei Zhao; J. Microbiol. 2021;59(1):29-40. Published online December 23, 2020; DOI: https://doi.org/10.1007/s12275-021-0478-0

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Abstract: The Gram-negative pathogen Pseudomonas aeruginosa adopts several elaborate strategies to colonize a wide range of natural or clinical niches and to overcome the neighboring bacterial competitors in polymicrobial communities. However, the relationship and interaction mechanism of P. aeruginosa with other bacterial pathogens remains largely unexplored. Here we explore the interaction dynamics of P. aeruginosa and Escherichia coli, which frequently coinfect the lungs of immunocompromised hosts, by using a series of on-plate proximity assays and RNA-sequencing. We show that the extracellular products of P. aeruginosa can inhibit the growth of neighboring E. coli and induce a large-scale of transcriptional reprogramming of E. coli, especially in terms of cellular respiration- related primary metabolisms and membrane components. In contrast, the presence of E. coli has no significant effect on the growth of P. aeruginosa in short-term culture, but causes a dysregulated expression of genes positively controlled by the quorum-sensing (QS) system of P. aeruginosa during subsequent pairwise culture. We further demonstrate that the divergent QS-regulation of P. aeruginosa may be related to the function of the transcriptional regulator PqsR, which can be enhanced by E. coli culture supernatant to increase the pyocyanin production by P. aeruginosa in the absence of the central las-QS system. Moreover, the extracellular products of E. coli promote the proliferation and lethality of P. aeruginosa in infecting the Caenorhabditis elegans model. The current study provides a general characterization of the extracellular products-mediated interactions between P. aeruginosa and E. coli, and may facilitate the understanding of polymicrobial infections.

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