• Journal of Microbiology and Biotechnology
• /
• v.14 no.1
• /
• pp.74-80
• /
• 2004

Aspergillus fumigatus is one of the most common fungi in the human environment, both in-doors and out-doors. It is the main causative agent of invasive aspergillosis, a life-threatening mycosis among immunocompromised patients. The genome has been sequenced by an international consortium, including the Wellcome Trust Sanger Institute (U.K.) and The Institute for Genomic Research (TIGR, U.S.A.), and a ten times whole genome shotgun sequence assembly has been made publicly available. In this study, we identified tricarboxylic acid (TCA) cycle enzymes of A. fumigatus by comparative analysis with four other fungal species. The open reading frames showed high amino acid sequence similarity with the other fungal citric acid enzymes and well-conserved functional domains. All genes present in Saccharomyces cerevisiae, Schizosaccharomyces pombe, Candida albicans, and Neurospora crassa were also found in A. fumigatus. In addition, we identified four A. fumigatus genes coding for enzymes in the glyoxylate shunt, which may be required for fungal virulence. The architecture of multi-gene encoded enzymes, such as isocitrate dehydrogenase, 2-ketoglutarate, succinyl-CoA synthetase, and succinate dehydrogenase was well conserved in A. fumigatus. Furthermore, our results show that genes of A. fumigatus can be detected reliably using GlimmerM.

• Applied Biological Chemistry
• /
• v.41 no.3
• /
• pp.219-223
• /
• 1998

We have established a transformation system for entomopathogenic fungus, Metarhizium anisopliae, in order to develop mycoinsecticide by recombinant DNA techniques. Protoplasts of M. anisopliae would be transformed to a benomyl-resistant by introducing pBRG-4 plasmid DNA, which contains a ${\beta}-tubulin$ gene of Aspergillus flavus conferring resistance to benomyl and a pyr4 gene of Neurospora crassa, in the presence of 5% polyethylene glycol and 10 mM calcium chloride. Transformants occuring at a frequency of 10 colonies per $50\;{\mu}g$ pBRG-4 DNA grew on the $5\;{\mu}g/ml$ concentrations of benamyl, while the wild type was inhibited by $2.5\;{\mu}g/ml$. From the Southern analysis using genomic DNAs isolated from M. anisopliae transformants, the positive signals suggested that the ${\beta}-tubulin$ gene had integrated in the M. anisopliae genome by homologous recombination.

• Journal of Microbiology and Biotechnology
• /
• v.15 no.1
• /
• pp.22-28
• /
• 2005

Gibberellins (GAs) play important roles in plant growth and development. Fifty-four fungi were isolated from the roots of 4 kinds of Sesamum indicum plants, and the production of GAs was spectrophotometrically examined. The number of GA-producing fungi was two strains from S. indicum, four strains from Gold S. indicum, and five strains from Brown S. indicum. Eleven fungi with GAs-producing activity were incubated for seven days in 40 ml of Czapek's liquid medium at $25^{\circ}C$ and 120 rpm, and the amount of each GA in the medium was measured by gas chromatographymass spectrometery (GC-MS). Penicillium commune KNU5379 produced more $GA_3$, $GA_4$, and $GA_7$ than Gibberella fujikuroi, Fusarium proliferatum, and Neurospora crassa which are known as GAs-producing fungi. GAs-producing activity of the P. commune KNU5379 was shown to produce 71.69 ng of $GA_1$, 252.42 ng of $GA_3$, 612.00 ng of $GA_4$, 259.00 ng of $GA_7$, and 202.69 ng of $GA_9$ in 25 ml of liquid medium. Bioassay of culture fluid of GA-producing fungi was also performed on rice sprout.

• Proceedings of the Korean Vacuum Society Conference
• /
• 2011.08a
• /
• pp.335-335
• /
• 2011

곰팡이에 의한 감염과 바이오 필름 형성이 우리 인간에게 매우 유해하다는 것은 익히 알려진 바이다. 일례로, 이는 우리가 먹는 음식을 오염시키고 치아를 상하게 하며 수도관을 부식시키고 또 우리 몸속에 집어넣는 의료기기에서 자라 심각한 질병을 유발시키는 등 여러가지 문제를 일으킨다. 곰팡이 감염과 바이오 필름 형성을 막기 위해 항생제 및 화학물질 등을 이용하는 방법들이 있으나 저항성이 증가된 균주의 출현, 환경오염 등의 문제점을 유발시키고 있다. 따라서 좀더 지속적이고 환경친화적인 기술개발이 요구되어지고 있다. 강한 살균력이 있다고 알려져 있는 대기압 플라즈마는 이러한 기술개발에 적용해볼 필요가 있다. 본 연구에서는 상온 대기압 플라즈마를 만들어 내는 여러 타입의 장치를 이용하여 곰팡이 살균과 이스트 바이오필름 형성 저해에 대한 조사를 하였고 플라즈마 처리에 따른 곰팡이 세포들의 세포학적 분자생물학적 영향에 대해 조사해 보았다. 대기압 상온 플라즈마 제트는 대기중에서 방안의 온도로 플라즈마가 생성되며 이를 페트리디쉬위에 접종된 이스트(Saccharomyces cerevisiae) 위 10 mm에서 조사한 결과 2분 이상의 처리를 했을 때 바이오 필름형성을 하지 못함을 보였고 곰팡이(Neurospora crassa)포자에 처리했을 때는 처리시간에 비례하여 발아하는 포자수가 감소하고 포자의 모양도 수축되는 것으로 관찰되었다. 대기압 플라즈마 제트는 대면적이 아닌 국부적 살균으로 살균력은 보이나 그 효율성이 낮아 이를 보완하기위해 DBD 형식의 플라즈마 장치를 만들었으며 이는 페트리디쉬의 모든 면을 동시에 조사할 수 있는 장점이 있다. 다만 제트와는 달리 플라즈마의 생성구간이 넓고 얇아 시료와 2~3 mm 간격을 띄우고 조사하였다. 제트와 마찬가지로 살균의 효과를 보였으며 기존의 단점을 극복하여 동시에 더 많은 면적을 조사 할 수 있게 되었다. 이를 통해 플라즈마의 살균능력을 직접 확인하였고 앞으로 더 많은 실험과 연구를 통해 일련의 과정들에 대한 원인을 규명하고자 한다.

• Journal of Microbiology
• /
• v.43 no.6
• /
• pp.475-486
• /
• 2005

Fungal secondary metabolites constitute a wide variety of compounds which either playa vital role in agricultural, pharmaceutical and industrial contexts, or have devastating effects on agriculture, animal and human affairs by virtue of their toxigenicity. Owing to their beneficial and deleterious characteristics, these complex compounds and the genes responsible for their synthesis have been the subjects of extensive investigation by microbiologists and pharmacologists. A majority of the fungal secondary metabolic genes are classified as type I polyketide synthases (PKS) which are often clustered with other secondary metabolism related genes. In this review we discuss on the significance of our recent discovery of chalcone synthase (CHS) genes belonging to the type III PKS superfamily in an industrially important fungus, Aspergillus oryzae. CHS genes are known to playa vital role in the biosynthesis of flavonoids in plants. A comparative genome analyses revealed the unique character of A. oryzae with four CHS-like genes (csyA, csyB, csyC and csyD) amongst other Aspergilli (Aspergillus nidulans and Aspergillus fumigatus) which contained none of the CHS-like genes. Some other fungi such as Neurospora crassa, Fusarium graminearum, Magnaporthe grisea, Podospora anserina and Phanerochaete chrysosporium also contained putative type III PKSs, with a phylogenic distinction from bacteria and plants. The enzymatically active nature of these newly discovered homologues is expected owing to the conservation in the catalytic residues across the different species of plants and fungi, and also by the fact that a majority of these genes (csyA, csyB and csyD) were expressed in A. oryzae. While this finding brings filamentous fungi closer to plants and bacteria which until recently were the only ones considered to possess the type III PKSs, the presence of putative genes encoding other principal enzymes involved in the phenylpropanoid and flavonoid biosynthesis (viz., phenylalanine ammonia-lyase, cinnamic acid hydroxylase and p-coumarate CoA ligase) in the A. oryzae genome undoubtedly prove the extent of its metabolic diversity. Since many of these genes have not been identified earlier, knowledge on their corresponding products or activities remain undeciphered. In future, it is anticipated that these enzymes may be reasonable targets for metabolic engineering in fungi to produce agriculturally and nutritionally important metabolites.

• Proceedings of the Microbiological Society of Korea Conference
• /
• 2000.10a
• /
• pp.39-45
• /
• 2000

It hab been proposed that CHS3-mediated chitin synthesis during the vegitative cell cycle is regulated by CHS4. To investigate direct protein-protein interaction between their coding products, we used yeast two hybrid system and found that a domain of Chs3p was responsible for interaction with Chs4p. This domain, termed MIRC3-4 (maximum interacting region of chs3p with chs4p), spans from 647 to 700 residues. It is well conserved among CHS3 homologs of various fungi such as Candida albicans, Emericella nidulans, Neurospora crassa, Magnaporthe grisea, Ustilago maydis, Glomus versiforme, Exophiala dermatitidis, Rhizopus microsporus. A series of mutaion in the MIRC3-4 resulted in no appearance of chitin ring at the early G 1 phase but did not affect chitin synthesis in the cell wall after cytokinesis. Absence of chitin ring could be caused either by delocalization of Chs3p to the septum or by improper interaction with Chs4p. To discriminate those two, not mutually exclusive, alternatives, mutants cells were immunostained with Chs3p-specific antibody. Some exhibited localization of chs3p to the septum, while others failed. These results indicate that simultaneous localization and activation Chs3p by Chs4p is required for chitin ring synthesis.

• The Korean Journal of Mycology
• /
• v.32 no.1
• /
• pp.16-22
• /
• 2004

The gibberellins (GAs) play important roles in plant growth and development. Twenty three fungi were isolated from the roots of Lindera obtusiloba and Vaccinium koreanum. The numbers of GA-producing fungi were six strains from Lindera obtusiloba and four strains from Vaccinium koreanum. The fungi with GAs-producing activity were incubated for seven days in 40 ml of Czapek's liquid medium at $25^{\circ}C$, 120 rpm, and the amount of each GA in the medium was measured by gas chromatography-mass spectrometer (GC-MS) to determine the productivity of GAs. Penicillium griseofulvum KNU5379 produced more GA in case of $GA_{3}$ than Neurospora crassa known as a GAs-producing fungus. P. griseofulvum KNU5379 was shown to produce $GA_1\;9.79\;ng,\;GA_3\;133.58\;ng,\;GA_4\;2.64\;ng,\;GA_7\;7.80\;ng\;and\;GA_{53}\;0.73\;ng$ in 25 ml of liquid medium. Bioassay using culture fluid of GAs-production fungi was performed on rice sprout.

• The Korean Journal of Mycology
• /
• v.41 no.1
• /
• pp.1-8
• /
• 2013

COP9 signalosome (CSN), which is originally identified as the regulator of the photomorphogenic development in plant, is highly conserved protein complex in diverse eukaryotic organisms. Most eukaryotic CSN complex is composed of 8 subunits, which is structurally and functionally similar to the lid subunit of 26S proteasome and eIF3 translation initiation complex. CSN play important functions in the regulation of cell cycle and checkpoint response by controlling Cullin-Ring E3 ubiquitin ligases (CRL) activities. CSN exhibits an isopeptidase activity which cleaves the neddylated moiety of cullin components. In fission yeast, S-phase cell cycle progression was delayed and the sensitivity to g-ray or UV was increased in CSN1 and CSN2 deletion mutants, indicating that yeast CSN is also involved in the checkpoint regulation. CSN in fungal system more closely resembles that of the higher organisms in the structure and assembly of their components. Functionally, CSN is associated with the regulation of conidiation rhythms in Neurospora crassa and the sexual development in Aspsergillus nidulans. Recent studies also revealed that CSN functions as an essential cell cycle regulator, playing key roles in the regulation of DNA replication and DNA damage response in Aspergillus. Overall, CSN of microorganisms, such as fission yeast and fungi, share functionally common aspects with higher organisms, implying that they can be useful tools to study the role of CSN in the CRL-mediated diverse cellular activities.

• Journal of Microbiology and Biotechnology
• /
• v.30 no.2
• /
• pp.206-215
• /
• 2020

Trichoderma reesei is the major filamentous fungus used to produce cellulase and there is huge interest in promoting its ability to produce higher titers of cellulase. Among the many factors affecting cellulase production in T. reesei, the mycelial phenotype is important but seldom studied. Herein, a close homolog of the Neurospora crassa COT1 kinase was discovered in T. reesei and designated TrCOT1, which is of 83.3% amino acid sequence identity. Functional disruption of Trcot1 in T. reesei by RNAi-mediated gene silencing resulted in retarded sporulation on potato dextrose agar and dwarfed colonies on minimal medium agar plates containing glucose, xylan, lactose, xylose, or glycerol as the sole carbon source. The representative mutant strain, SUS2/Trcot1i, also displayed reduced mycelia accumulation but hyperbranching in the MM glucose liquid medium, with hyphal growth unit length values decreased to 73.0 ㎛/tip compared to 239.8 ㎛/tip for the parent strain SUS2. The hyperbranching phenotype led to slightly but significantly increased cellulase secretion from 24 to 72 h in a batch culture. However, the cellulase production per unit of mycelial biomass was much more profoundly improved from 24 to 96 h.

• Journal of Microbiology and Biotechnology
• /
• v.18 no.1
• /
• pp.15-22
• /
• 2008

Four series (S, M, R, and W) of Alternaria longipes isolates were obtained based on consecutive selection with Dimethachlon (Dim) and ultraviolet irradiation. These isolates were then characterized according to their tolerance to Dim, sensitivity to osmotic stress, and phenotypic properties. All the selected Dim-resistant isolates showed a higher osmosensitivity than the parental strains, and the last generation was more resistant than the first generation in the M, R, and W series. In addition, the changes in the Dim resistance and osmotic sensitivity were not found to be directly correlated, and no distinct morphologic characteristics were found among the resistant and sensitive isolates, with the exception of the resistant isolate K-11. Thus, to investigate the molecular basis of the fungicide resistance, a group III two-component histidine kinase (HK) gene, AlHK1, was cloned from nineteen A. longipes isolates. AlHK1p was found to be comprised of a six 92-amino-acid repeat domain (AARD), HK domain, and response regulator domain, similar to the Os-1p from Neurospora crassa. A comparison of the nucleotide sequences of the AlHK1 gene from the Dim-sensitive and -resistant isolates revealed that all the resistant isolates contained a single-point mutation in the AARD of AlHK1p, with the exception of isolate K-11, where the AlHK1p contained a deletion of 107 amino acids. Moreover, the AlHK1p mutations in the isolates of each respective series involved the same amino acid substitution at the same site, although the resistance levels differed significantly in each series. Therefore, these findings suggested that a mutation in the AARD of AlHK1p was not the sole factor responsible for A. longipes resistance to dicarboximide fungicides.