Jae-Seoun Hur

Anti-proliferative effects of Lethariella zahlbruckneri extracts in human HT-29 human colon cancer cells.

This study was performed to elucidate the anti-proliferative effects and the apoptotic mechanisms of extracts from Lethariella zahlbruckneri in HT-29 human colon cancer cells. Both the acetone extract (AEL) and methanolic extract (MEL) of L. zahlbruckneri decreased viable cell numbers in a dose- and time-dependent manner in HT-29 cells. The AEL showed stronger cytotoxicity than MEL. Cell death induced by AEL increased cell populations in the sub-G1 phase, as well as the formation of apoptotic bodies and nuclear condensation, whereas MEL did not. Therefore, the potential of AEL to induce apoptosis was examined. Apoptosis induced by AEL was associated with the activation of initiator caspases-8 and -9, as well as the effector caspase-3. AEL stimulated Bid cleavage. This indicated that the apoptotic action of caspase-8-mediated Bid cleavage leads to the activation of caspase-9. AEL increased the expression of the pro-apoptotic protein, Bax, and decreased the expression of the anti-apoptotic protein, Bcl-2. AEL also increased the expression of the caspase-independent mitochondrial apoptosis factor, AIF, in HT-29 cells. These results indicate that AEL inhibited HT-29 cell proliferation by inducing apoptosis, which might be mediated via both caspase-dependent and -independent pathways.

Evolution of complex symbiotic relationships in a morphologically derived family of lichen-forming fungi.

We studied the evolutionary history of the Parmeliaceae (Lecanoromycetes, Ascomycota), one of the largest families of lichen-forming fungi with complex and variable morphologies, also including several lichenicolous fungi. We assembled a six-locus data set including nuclear, mitochondrial and low-copy protein-coding genes from 293 operational taxonomic units (OTUs). The lichenicolous lifestyle originated independently three times in lichenized ancestors within Parmeliaceae, and a new generic name is introduced for one of these fungi. In all cases, the independent origins occurred c. 24 million yr ago. Further, we show that the Paleocene, Eocene and Oligocene were key periods when diversification of major lineages within Parmeliaceae occurred, with subsequent radiations occurring primarily during the Oligocene and Miocene. Our phylogenetic hypothesis supports the independent origin of lichenicolous fungi associated with climatic shifts at the Oligocene-Miocene boundary. Moreover, diversification bursts at different times may be crucial factors driving the diversification of Parmeliaceae. Additionally, our study provides novel insight into evolutionary relationships in this large and diverse family of lichen-forming ascomycetes.

Bactericidal activity and water purification of immobilized TiO2 photocatalyst in bean sprout cultivation

Immobilized TiO2 photocatalysts were used to sterilize and reclaim the wastewater of bean sprout cultivation from a continuous hydro-circulation system. The photocatalysts effectively killed bacteria and degraded organic pollutants in the wastewater. Stimulation of bean sprout growth and suppression of decaying pathogens were also induced by the TiO2 photocatalytic activity.

Outbreak of bacterial canker on Hort16A (Actinidia chinensis Planchon) caused by Pseudomonas syringae pv. actinidiae in Korea

Abstract Bacterial canker was first observed on the kiwifruit cv. ‘Hort16A (Actinidia chinensis)’ in Jeju Province, Korea, in spring 2006. Die back or blight on young canes, often with red-rusty exudation on canes or trunks, and dark brown irregular spots surrounded with yellowish halos on leaves are the typical symptoms. These symptoms closely resemble those caused by Pseudomonas syringae pv. actinidiae on the kiwifruit cv. ‘Hayward (Actinidia deliciosa)’. A sudden outbreak and rapid spread of the bacterial canker resulted in the death of severely infected vines and eradication of completely devastated orchards of the kiwifruit cv. ‘Hort16A’. Contaminated pruning shears and climatic conditions appear to have been responsible for the sudden outbreak and rapid spread of the epidemics on the kiwifruit cv. ‘Hort16A’ vines. The causal bacterium was isolated from diseased vines of the kiwifruit cv. ‘Hort16A’ and identified as P. syringae pv. actinidiae, which is the same bacterial pathogen responsible for cankers on the kiwifruit cv. ‘Hayward’ by morphological, cultural, physiological and biochemical, molecular and pathogenicity analyses.

Bacillus subtilis S1-0210 as a Biocontrol Agent against Botrytis cinerea in Strawberries

Bacillus subtilis S1-0210 was selected as a biological agent against Botrytis cinerea in strawberry. The isolate inhibited mycelial growth of B. cinerea in vitro tests. A wettable powder formulation of B. subtilis S1-0210 significantly reduced infection rates with lower than 5%, compared with higher than 70% of infection rates in untreated control. The formulation showed 85 to 89% control efficacies of gray mold incidences on fruits of strawberry in pots. Pre-treatment of the agent was more effective in controlling gray mold on fruits and leaves than post-treatment at the early stage of disease development. The formulation also showed 70% control efficacy of gray mold incidence on fruits of strawberry in a field trial. The results indicate that B. subtilis S1-0210 in the wettable powder formulation may be a potential biocontrol agent to control gray mold on strawberry.

Integrating coalescent and phylogenetic approaches to delimit species in the lichen photobiont Trebouxia

The accurate assessment of species boundaries in symbiotic systems is a prerequisite for the study of speciation, co-evolution and selectivity. Many studies have shown the high genetic diversity of green algae from the genus Trebouxia, the most common photobiont of lichen-forming fungi. However, the phylogenetic relationships, and the amount of cryptic diversity of these algae are still poorly understood, and an adequate species concept for trebouxiophycean algae is still missing. In this study we used a multifaceted approach based on coalescence (GMYC, STEM) and phylogenetic relationships to assess species boundaries in the trebouxioid photobionts of the lichen-forming fungus Lasallia pustulata. We further investigated whether putative species of Trebouxia found in L. pustulata are shared with other lichen-forming fungi. We found that L. pustulata is associated with at least five species of Trebouxia and most of them are shared with other lichen-forming fungi, showing different patterns of species-to-species and species-to-community interactions. We also show that one of the putative Trebouxia species is found exclusively in association with L. pustulata and is restricted to thalli from localities with Mediterranean microclimate. We suggest that the species delimitation method presented in this study is a promising tool to address species boundaries within the heterogeneous genus Trebouxia.

Agrobacterium tumefaciens -Mediated Transformation of the Lichen Fungus, Umbilicaria muehlenbergii

Transformation-mediated mutagenesis in both targeted and random manners has been widely applied to decipher gene function in diverse fungi. However, a transformation system has not yet been established for lichen fungi, severely limiting our ability to study their biology and mechanism underpinning symbiosis via gene manipulation. Here, we report the first successful transformation of the lichen fungus, Umbilicaria muehlenbergii, via the use of Agrobacterium tumefaciens. We generated a total of 918 transformants employing a binary vector that carries the hygromycin B phosphotransferase gene as a selection marker and the enhanced green fluorescent protein gene for labeling transformants. Randomly selected transformants appeared mitotically stable, based on their maintenance of hygromycin B resistance after five generations of growth without selection. Genomic Southern blot showed that 88% of 784 transformants contained a single T-DNA insert in their genome. A number of putative mutants affected in colony color, size, and/or morphology were found among these transformants, supporting the utility of Agrobacterium tumefaciens-mediated transformation (ATMT) for random insertional mutagenesis of U. muehlenbergii. This ATMT approach potentially offers a systematic gene functional study with genome sequences of U. muehlenbergii that is currently underway.

Versatile biocatalysis of fungal cytochrome P450 monooxygenases

Cytochrome P450 (CYP) monooxygenases, the nature’s most versatile biological catalysts have unique ability to catalyse regio-, chemo-, and stereospecific oxidation of a wide range of substrates under mild reaction conditions, thereby addressing a significant challenge in chemocatalysis. Though CYP enzymes are ubiquitous in all biological kingdoms, the divergence of CYPs in fungal kingdom is manifold. The CYP enzymes play pivotal roles in various fungal metabolisms starting from housekeeping biochemical reactions, detoxification of chemicals, and adaptation to hostile surroundings. Considering the versatile catalytic potentials, fungal CYPs has gained wide range of attraction among researchers and various remarkable strategies have been accomplished to enhance their biocatalytic properties. Numerous fungal CYPs with multispecialty features have been identified and the number of characterized fungal CYPs is constantly increasing. Literature reveals ample reviews on mammalian, plant and bacterial CYPs, however, modest reports on fungal CYPs urges a comprehensive review highlighting their novel catalytic potentials and functional significances. In this review, we focus on the diversification and functional diversity of fungal CYPs and recapitulate their unique and versatile biocatalytic properties. As such, this review emphasizes the crucial issues of fungal CYP systems, and the factors influencing efficient biocatalysis.

Antifungal Activity of Lichen-forming Fungi against Colletotrichum acutatum on Hot Pepper

Antifungal activity of Korean and Chinese lichen-forming fungi (LFF) was evaluated against plant pathogenic fungus of Colletotrichum acutatum, causal agent of anthracnose on hot pepper. This is the first attempt to evaluate antifungal activity of LFF, instead of lichen thalli, against C. acutatum. Total 100 LFF were isolated from the lichens with discharged spore method or tissue culture method. Among the 100 isolates, 8 LFF showed more than 50% of inhibition rates of mycelial growth of the target pathogen. Especially, Lecanora argentata was highly effective in inhibition of mycelial growth of C. accutatum at the rate of 68%. Antifungal activity of other LFF was in the order of Cetrelia japonica (61.4%), Ramalina conduplicans (59.5%), Umbilicaria esculenta (59.5%), Ramalina litoralis (56.7%), Cetrelia braunsiana (56.5%), Nephromopsis pallescensn (56.1%), and Parmelia simplicior (53.8%). Among the tested LFF, 61 isolates of LFF exhibited moderate antifungal activity against the target pathogen at the inhibition rates from 30 to 50%. Antifungal activity of the LFF against C. acutatum was variable at the species level rather than genus level of LFF. This study suggests that LFF can be served as a promising bioresource to develop novel biofungicides.

Lichen Secondary Metabolites in Flavocetraria cucullata Exhibit Anti-Cancer Effects on Human Cancer Cells through the Induction of Apoptosis and Suppression of Tumorigenic Potentials

Lichens are symbiotic organisms which produce distinct secondary metabolic products. In the present study, we tested the cytotoxic activity of 17 lichen species against several human cancer cells and further investigated the molecular mechanisms underlying their anti-cancer activity. We found that among 17 lichens species, F. cucullata exhibited the most potent cytotoxicity in several human cancer cells. High performance liquid chromatography analysis revealed that the acetone extract of F. cucullata contains usnic acid, salazinic acid, Squamatic acid, Baeomycesic acid, d-protolichesterinic acid, and lichesterinic acid as subcomponents. MTT assay showed that cancer cell lines were more vulnerable to the cytotoxic effects of the extract than non-cancer cell lines. Furthermore, among the identified subcomponents, usnic acid treatment had a similar cytotoxic effect on cancer cell lines but with lower potency than the extract. At a lethal dose, treatment with the extract or with usnic acid greatly increased the apoptotic cell population and specifically activated the apoptotic signaling pathway; however, using sub-lethal doses, extract and usnic acid treatment decreased cancer cell motility and inhibited in vitro and in vivo tumorigenic potentials. In these cells, we observed significantly reduced levels of epithelial-mesenchymal transition (EMT) markers and phosphor-Akt, while phosphor-c-Jun and phosphor-ERK1/2 levels were only marginally affected. Overall, the anti-cancer activity of the extract is more potent than that of usnic acid alone. Taken together, F. cucullata and its subcomponent, usnic acid together with additional component, exert anti-cancer effects on human cancer cells through the induction of apoptosis and the inhibition of EMT.