• Journal of Microbiology and Biotechnology
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• v.20 no.11
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• pp.1484-1490
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• 2010

In the Streptomyces hygroscopicus JCM4427 geldanamycin biosynthetic gene cluster, five putative regulatory genes were identified by protein homology searching. Among those genes, gel14, gel17, and gel19 are located downstream of polyketide synthase genes. Gel14 and Gel17 are members of the LAL family of transcriptional regulators, including an ATP/GTP-binding domain at the N-terminus and a DNA-binding helix-turn-helix domain at the C-terminus. Gel19 is a member of the TetR family of transcriptional regulators, which generally act to repress transcription. To verify the biological significance of the putative regulators in geldanamycin production, they were individually characterized by gene disruption, genetic complementation, and transcriptional analyses. All three genes were confirmed as positive regulators of geldanamycin production. Specifically, Gel17 and Gel19 are required for gel14 as well as gelA gene expression.

• Journal of Microbiology
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• v.44 no.6
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• pp.649-654
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• 2006

A standard type II polyketide synthase (PKS) gene cluster was isolated while attempting to clone the biosynthetic gene for lipstatin from Streptomyces toxytricini NRRL 15,443. This result was observed using a Southern blot of a PstI-digested S. toxytricini chromosomal DNA library with a 444 bp amplified probe of a ketosynthase (KS) gene fragment. Four open reading frames [thioesterase (TE), $\beta$-ketoacyl systhase (KAS), chain length factor (CLF), and acyl carrier protein (ACP)], were identified through the nucleotide sequence determination and analysis of a 4.5 kb cloned DNA fragment. In order to confirm the involvement of a cloned gene in lipstatin biosynthesis, a gene disruption experiment for the KS gene was performed. However, the resulting gene disruptant did not show any significant difference in lipstatin production when compared to wild-type S. toxytricini. This result suggests that lipstatin may not be synthesized by a type II PKS.

• The Korean Journal of Mycology
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• v.38 no.1
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• pp.29-33
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• 2010

KGD1 gene was cloned by functional complementation of defects in $\beta$-1,3-glucan synthase activity of the previously isolated Saccharomyces cerevisiae mutant LP0353, which displays a number of cell wall defects at restrictive temperature. We performed the gene disruption experiment to characterize the function of KGD1 gene, which encodes $\beta$-ketoglutarate dehydrogenase, in cell wall biosynthesis. The disruption of KGD1 showed the decreased growth rate, the increase of chitin synthases activity, alterations in cell wall composition, and increase of susceptibility to cell wall inhibitors such as Calcofluor white and Nikkomycin Z. These results suggested that KGD1 might be involved in cell wall biogenesis, especially the biosynthesis of $\beta$-1,6-glucan and chitin in S. cerevisaie.

• The Plant Pathology Journal
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• v.35 no.5
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• pp.445-458
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• 2019

The lipopolysaccharide (LPS) composed of lipid A, core, and O-antigen is the fundamental constituent of the outer membrane in gram-negative bacteria. This study was conducted to investigate the roles of LPS in Burkholderia glumae, the phytopathogen causing bacterial panicle blight and seedling rot in rice. To study the roles of the core oligosaccharide (OS) and the O-antigen region, mutant strains targeting the waaC and the wbiFGHI genes were generated. The LPS profile was greatly affected by disruption of the waaC gene and slight reductions were observed in the O-antigen region following wbiFGHI deletions. The results indicated that disruption in the core OS biosynthesis-related gene, waaC, was associated with increased sensitivity to environmental stress conditions including acidic, osmotic, saline, and detergent stress, and to polymyxin B. Moreover, significant impairment in the swimming and swarming motility and attenuation of bacterial virulence to rice were also observed in the waaC-defective mutant. The motility and virulence of O-antigen mutants defective in any gene of the wbiFGHI operon, were not significantly different from the wild-type except in slight decrease in swimming and swarming motility with wbiH deletion. Altogether, the results of present study indicated that the LPS, particularly the core OS region, is required for tolerance to environmental stress and full virulence in B. glumae. To our knowledge, this is the first functional study of LPS in a plant pathogenic Burkholderia sp. and presents a step forward toward full understanding of B. glumae pathogenesis.

• Journal of Microbiology and Biotechnology
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• v.24 no.9
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• pp.1226-1231
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• 2014

The rare actinomycete Pseudonocardia autotrophica was previously shown to produce a solubility-improved toxicity-reduced novel polyene compound named $\underline{N}ystatin$-like $\underline{P}seudonocardia$ $\underline{P}olyene$ (NPP). The low productivity of NPP in P. autotrophica implies that its biosynthetic pathway is tightly regulated. In this study, $wblA_{pau}$ was isolated and identified as a novel negative regulatory gene for NPP production in P. autotrophica, which showed approximately 49% amino acid identity with a global antibiotic down-regulatory gene, wblA, identified from various Streptomycetes species. Although no significant difference in NPP production was observed between P. autotrophica harboring empty vector and the S. coelicolor wblA under its native promoter, approximately 12% less NPP was produced in P. autotrophica expressing the wblA gene under the strong constitutive $ermE^*$ promoter. Furthermore, disruption of the $wblA_{pau}$ gene from P. autotrophica resulted in an approximately 80% increase in NPP productivity. These results strongly suggest that identification and inactivation of the global antibiotic down-regulatory gene wblA ortholog are a critical strategy for improving secondary metabolite overproduction in not only Streptomyces but also non-Streptomyces rare actinomycete species.

• The Plant Pathology Journal
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• v.24 no.1
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• pp.1-7
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• 2008

Fusarium verticillioides (teleomorph Gibberella moniliformis) is associated with maize worldwide causing ear rot and stalk rot, and produces fumonisins, a group of mycotoxins detrimental to humans and animals. While research tools are available, our understanding of the molecular mechanisms associated with fungal virulence and fumonisin biosynthesis in F. verticillioides is still limited. One of the restraints that hampers F. verticillioides gene characterization is the fact that homologous recombination (HR) frequency is very low (<2%). Screening for a true gene knock-out mutant is a laborious process due to a high number of ectopic integrations. In this study, we generated a F. verticillioides mutant (SF41) deleted for FvKU70, a gene directly responsible for non-homologous end-joining mechanism, with the aim of improving HR frequency. Here, we demonstrate that FvKU70 deletion does not impact key Fverticillioides phenotypes, e.g., development, secondary metabolism, and virulence, while dramatically improving HR frequency. Significantly, we also confirmed that a high percentage (>85%) of the HR mutant strains harbor a desired mutation with no additional copy of the mutant allele inserted in the genome. We conclude that SF41 is suitable for use as a type strain when performing high-throughput gene function studies in F. verticillioides.

• Bulletin of the Korean Chemical Society
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• v.26 no.11
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• pp.1823-1825
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• 2005

Angiogenesis is tightly regulated by a variety of angiogenic activators and inhibitors. Disruption of the balanced angiogenesis leads to the progress of diseases such as cancer, rheumatoid arthritis, and diabetic blindness. Even though a number of proteins involved in angiogenesis have been identified so far, more protein factors remain to be identified due to complexity of the process. Here I report that pituitary tumor-transforming gene (PTTG) induces migration and tube formation of human umbilical vein endothelial cells (HUVECs). High levels of both vascular endothelial growth factor (VEGF) and basic fibroblast growth factor (bFGF) are detected in conditioned medium obtained from cells transfected with PTTG expression plasmid. Taken together, these results suggest that PTTG is an angiogenic factor that induces production of both VEGF and bFGF.

• Journal of Microbiology and Biotechnology
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• v.16 no.6
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• pp.979-982
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• 2006

Two knockout vectors, in which the truncated Kluyveromyces lactis URAS gene is flanked by a direct repeat, were developed for Saccharomyces cerevisiae. Each vector was designed to harbor 5'- and 3'-end homology regions for integration. Two knockout fragments were devised to integrate into the correct locus in a complementary manner to disrupt a gene of interest and. concomitantly to make functional Kl URA3 for transfomant selection. The use of dual complementary knockout cassettes was expected to dramatically reduce integration into unwanted loci in the genome. The knockout system developed in this study was successfully used for disruption of the GAL1 gene in S. cerevisiae.

• Journal of Microbiology and Biotechnology
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• v.28 no.7
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• pp.1068-1077
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• 2018

DKxanthenes are a class of yellow secondary metabolites produced by myxobacterial genera Myxococcus and Stigmatella. We identified a putative 49.5 kb DKxanthene biosynthetic gene cluster from Myxococcus stipitatus DSM 14675 by genomic sequence and mutational analyses. The cluster consisted of 15 genes (MYSTI_06004-MYSTI_06018) encoding polyketide synthases, non-ribosomal peptide synthases, and proteins with unknown functions. Disruption of the genes by plasmid insertion resulted in defects in the production of yellow pigments. High-performance liquid chromatography and liquid chromatography-tandem mass spectrometry analyses indicated that the yellow pigments produced by M. stipitatus DSM 14675 might be novel DKxanthene derivatives. M. stipitatus did not require DKxanthenes for the formation of heat-resistant viable spores, unlike Myxococcus xanthus. Furthermore, DKxanthenes showed growth inhibitory activity against the fungi Aspergillus niger, Candida albicans, and Rhizopus stolonifer.

• Proceedings of the Korean Biophysical Society Conference
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• 1999.06a
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• pp.65-65
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• 1999

Na$^{＋}$-Ca$^{2＋}$ Exchanger is known to playa critical role in the regulation of intracellular $Ca^{2＋}$ in many tissues and cells. In heart, the Na$^{＋}$-Ca$^{2＋}$ exchange is the principal $Ca^{2＋}$ extrusion mechanism and affects cardiac excitation-contraction coupling. To understand the functional role of cardiac Na$^{＋}$ -Ca$^{2＋}$ exchanger (NCXl) in vivo, we tried to ablate the cardiac Na$^{2+}$-Ca$^{2+}$ exchanger gene locus by the use of the gene targeting technologies.(omitted)ted)