Journal of Microbiology

Journal Articles

Characterization of the effects of terminators and introns on recombinant gene expression in the basidiomycete Ceriporiopsis subvermispora: Dong Xuan Nguyen , Emi Nishisaka , Moriyuki Kawauchi , Takehito Nakazawa , Masahiro Sakamoto , Yoichi Honda; J. Microbiol. 2020;58(12):1037-1045. Published online September 30, 2020; DOI: https://doi.org/10.1007/s12275-020-0213-2

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Terminators and introns are vital regulators of gene expression in many eukaryotes; however, the functional importance of these elements for controlling gene expression in Agaricomycetes remains unclear. In this study, the effects of Ceriporiopsis subvermispora terminators and introns on the expression of a recombinant hygromycin B phosphotransferase gene (hph) were characterized. Using a transient transformation system, we proved that a highly active terminator (e.g., the gpd terminator) is required for the efficient expression of the hph gene. Mutational analyses of the C. subvermispora gpd terminator revealed that hph expression was dictated by an A-rich region, which included a putative positioning element, and polyadenylation sites. In contrast, our results indicated that introns are not required for the expression of hph directed by the Csβ1-tub and Csgpd promoters in C. subvermispora. This study provides insights into the functions and cis-element requirements of transcriptional terminators in Agaricomycetes, which may be relevant for designing recombinant genes for this important fungal class.

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Development of a 2A peptide-based multigene expression system and its application for enhanced production of ganoderic acids in Ganoderma lucidum
Qiong Wang, Hong-Jun Liu, Yan Xu, Zi-Xu Wang, Bin Sun, Jun-Wei Xu
Journal of Biotechnology.2024; 393: 109. CrossRef
CRISPR/Cas9 using a transient transformation system in Ceriporiopsis subvermispora
Takehito Nakazawa, Chikako Inoue, Dong Xuan Nguyen, Moriyuki Kawauchi, Masahiro Sakamoto, Yoichi Honda
Applied Microbiology and Biotechnology.2022; 106(17): 5575. CrossRef
A promoter assay system using gene targeting in agaricomycetes Pleurotus ostreatus and Coprinopsis cinerea
Dong Xuan Nguyen, Takehito Nakazawa, Genki Myo, Chikako Inoue, Moriyuki Kawauchi, Masahiro Sakamoto, Yoichi Honda
Journal of Microbiological Methods.2020; 179: 106053. CrossRef

Proteome analysis reveals global response to deletion of mrflbA in Monascus ruber: Qingqing Yan , Zhouwei Zhang , Yishan Yang , Fusheng Chen , Yanchun Shao; J. Microbiol. 2018;56(4):255-263. Published online February 28, 2018; DOI: https://doi.org/10.1007/s12275-018-7425-8

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Abstract

Monascus spp. are commonly used for a wide variety of applications in the food and pharmaceutical industries. In previous studies, the knock-out of mrflbA (a putative regulator of the G protein α subunit) in M. ruber led to autolysis of the mycelia, decreased pigmentation and lowered mycotoxin production. Therefore, we aimed to obtain a comprehensive overview of the underlying mechanism of mrflbA deletion at the proteome level. A two-dimensional gel electrophoresis analysis of mycelial proteins indicated that the abundance of 178 proteins was altered in the ΔmrflbA strain, 33 of which were identified with high confidence. The identified proteins are involved in a range of activities, including carbohydrate and amino acid metabolism, hyphal development and the oxidative stress response, protein modification, and the regulation of cell signaling. Consistent with these findings, the activity of antioxidative enzymes and chitinase was elevated in the supernatant of the ΔmrflbA strain. Furthermore, deletion of mrflbA resulted in the transcriptional reduction of secondary metabolites (pigment and mycotoxin). In short, the mutant phenotypes induced by the deletion of mrflbA were consistent with changes in the expression levels of associated proteins, providing direct evidence of the regulatory functions mediated by mrflbA in M. ruber.

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Histone deacetylase MrHos3 negatively regulates the production of citrinin and pigments in Monascus ruber
Qianrui Liu, Yunfan Zheng, Baixue Liu, Fufang Tang, Yanchun Shao
Journal of Basic Microbiology.2023; 63(10): 1128. CrossRef
Histone deacetylase MrRpd3 plays a major regulational role in the mycotoxin production of Monascus ruber
Yunfan Zheng, Yueyan Huang, Zejing Mao, Yanchun Shao
Food Control.2022; 132: 108457. CrossRef
Characterization of key upstream asexual developmental regulators in Monascus ruber M7
Lili Jia, Yuyun Huang, Jae-Hyuk Yu, Marc Stadler, Yanchun Shao, Wanping Chen, Fusheng Chen
Food Bioscience.2022; 50: 102153. CrossRef
Quantitative Proteomics Analysis by Sequential Window Acquisition of All Theoretical Mass Spectra–Mass Spectrometry Reveals Inhibition Mechanism of Pigments and Citrinin Production of Monascus Response to High Ammonium Chloride Concentration
Bo Zhou, Yifan Ma, Yuan Tian, Jingbo Li, Haiyan Zhong
Journal of Agricultural and Food Chemistry.2020; 68(3): 808. CrossRef

Research Support, Non-U.S. Gov't

Screening and Identification of a Novel Esterase EstPE from a Metagenomic DNA Library: So-Youn Park , Hyun-Jae Shin , Geun-Joong Kim; J. Microbiol. 2011;49(1):7-14. Published online March 3, 2011; DOI: https://doi.org/10.1007/s12275-011-0201-7

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Abstract: Esterases represent a large family of hydrolases with broad substrate specificity and functional sequence space. Although many attempts to screen new esterases have been conducted, there have been few reports conducted to discriminate unique enzymes from typical ones based on novel structure and function. In this study, we discovered an esterase and a novel family through a successive assay of whole cells and crude lysates (oxidative open condition). The screened putative esterases from the metagenomic DNA of salted shrimp consisted of 753 bp encoding 27 kDa of polypeptide, namely PE esterase. Sequence analyses revealed that an identical gene was reported from whole genome sequencing of Stenotrophomonas maltophilia K279a. However, its biochemical and phylogenetic characteristics have not yet been evaluated. PE esterase was overexpressed only by the MBP fusion state in E. coli and was easily purified using an affinity column. This enzyme showed a typical spectrum of substrate specificity and possessed the consensus motifs, Ser-Asp-His and GXSXG, which are essential for most esterase/lipase superfamilies. Interestingly, the entire organization of the ORF and consensus sequence around the active site were distinct from the related enzymes, and its structure could be affected by a reducing agent, DTT.

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