• Journal of Life Science
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• v.20 no.2
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• pp.176-182
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• 2010

Magnaporthe oryzae is destructive plant-pathogenic fungus and causes rice blast. The pathogen uses several mechanisms to circumvent the inhibitory actions of fungicides. ATP-binding cassette (ABC) transporters are known to provide protection against toxic compounds in the environment. PC facilitated bioinformatic analysis, particularly with respect to accessing and extracting database information and domain identification. We predicted ABC transporter genes from the M. oryzae genome sequence with computation and bioinformatics tools. A total of thirty three genes were predicted to encode ABC transporters. Three of thirty three putative genes corresponded to three known ABC transporter genes (ABC1, ABC2 and ABC3). Copy numbers of the ABC transporter genes were proven by Southern blot analysis, which revealed that twenty genes tested exist as a single copy. We amplified the DNA complementary to RNA corresponding to eleven of these by reverse transcriptase polymerase chain reaction.

• Journal of Microbiology and Biotechnology
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• v.16 no.9
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• pp.1422-1428
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• 2006

An ABC transporter apparatus of the Gram-negative bacterial type I secretion pathway can be used as a secretory protein expression system in Escherichia coli. Four types of coexpression systems for the Pseudomonas fluorescens lipase gene, tliA, and its cognate ABC transporter gene cluster, tliDEF, were constructed. When the relative expression levels were changed by adding different concentrations of IPTG, the secretion (16.9 U/ml of culture) of TliA in E. coli [pTliDEFA-223+pACYC184] was significantly higher than E. coli [pKK223-3+pTliDEFA-184] secreting the lowest level of TliA (5.2 U/ml of culture). Maximal accumulation of the lipase secreted occurred in the mid-exponential phase, implying that the efficient protein secretion via an ABC transporter was restricted only to actively growing cells. Finally, the secretion level of TliA in E. coli [pTliDEFA-223+pACYC184] was increased to 26.4 U/ml by inducing gene expression at the culture initiation time. These results indicate that a significant increase in the ABC transporter-dependent protein secretion can be achieved by simply controlling the relative expression levels between the ABC transporter and its passenger protein, even in the recombinant E. coli cells.

• The Plant Pathology Journal
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• v.30 no.4
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• pp.375-383
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• 2014

Fungi tolerate exposure to various abiotic stresses, including cytotoxic compounds and fungicides, via their ATP-driven efflux pumps belonging to ATP-binding cassette (ABC) transporters. To clarify the molecular basis of interaction between the fungus and various abiotic stresses including fungicides, we constructed a cDNA library from germinated conidia of Colletotrichum acutatum, a major anthracnose pathogen of pepper (Capsicum annum L.). Over 1,000 cDNA clones were sequenced, of which single clone exhibited significant nucleotide sequence homology to ABC transporter genes. We isolated three fosmid clones containing the C. acutatum ABC1 (CaABC1) gene in full-length from genomic DNA library screening. The CaABC1 gene consists of 4,059 bp transcript, predicting a 1,353-aa protein. The gene contains the typical ABC signature and Walker A and B motifs. The 5'-flanking region contains a CAAT motif, a TATA box, and a Kozak region. Phylogenetic and structural analysis suggested that the CaABC1 is a typical ABC transporter gene highly conserved in various fungal species, as well as in Chromista, Metazoans, and Viridiplantae. We also found that CaABC1 was up-regulated during conidiation and a minimal medium condition. Moreover, CaABC1 was induced in iprobenfos, kresoxim-methyl, thiophanate-methyl, and hygromycin B. These results demonstrate that CaABC1 is necessary for conidiation, abiotic stress, and various fungicide resistances. These results will provide the basis for further study on the function of ABC transporter genes in C. acutatum.

• The Plant Pathology Journal
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• v.32 no.1
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• pp.8-15
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• 2016

To identify a novel biopesticide controlling rice blast disease caused by Magnaporthe oryzae, 700 plant extracts were evaluated for their inhibitory effects on mycelial growth of M. oryzae. The L. foenum-graecum Herba extract showed the lowest inhibition concentration ($IC_{50}$) of $39.28{\mu}g/ml$, which is lower than the $IC_{50}$ of blasticidin S ($63.06{\mu}g/ml$), a conventional fungicide for rice blast disease. When treatments were combined, the $IC_{50}$ of blasticidin S was dramatically reduced to $10.67{\mu}g/ml$. Since ABC transporter genes are involved in fungicide resistance of many organisms, we performed RT-PCR to investigate the transcriptional changes of 40 ABC transporter family genes of M. oryzae treated with the plant extract, blasticidin S, and tetrandrine, a recognized ABC transporter inhibitor. Four ABC transporter genes were prominently activated by blasticidin S treatment, but were suppressed by combinational treatment of blasticidin S with the plant extract, or with tetrandrine that didn't show cellular toxicity by itself in this study. Mycelial death was detected via confocal microscopy at 24 h after plant extract treatment. Finally, subsequent rice field study revealed that the plant extract had high control efficacy of 63.3% and should be considered a biopesticide for rice blast disease. These results showed that extract of L. foenum graecum Herba suppresses M. oryzae ABC transporter genes inducing mycelial death and therefore may be a potent novel biopesticide.

• Korean journal of applied entomology
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• v.59 no.4
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• pp.341-348
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• 2020

The ATP-binding cassette (ABC) transporter superfamily represents the largest transmembrane protein that transports a variety of substrates across extra- and intra-cellular membranes. In insects, the ABC transporter proteins play crucial roles in insecticide resistance. To date, no studies have investigated the involvement of ABC transporter gene for cross-resistance to insecticide chemistries. Here, we studied such possible mechanisms against six conventional insecticides using transgenic Drosophila melanogaster strains carrying Mdr49 transcript variant A. For the 91-R and 91-C strains of Drosophila melanogaster, although they have a common origin, 91-R has been intensely selected with DDT for over 60 years, while 91-C has received no insecticide selection. Our transgenic analyses showed that overexpression of 91-R-MDR49 transcript variant A along with three amino acid variations can yield a relatively low degree of cross-resistance to carbofuran (2.0~6.7-fold) and permethrin (2.5~10.5-fold) but did not show cross-resistance to abamectin, imidacloprid, methoxychlor, and prothiofos as compared to the Gal4-driver control strain without transgene expression. These results indicate that the overexpression of Mdr49A in itself leads to a cross-resistance and three amino acid changes have additional effects on positive cross-resistance to carbofuran and permethrin.

• Journal of Marine Life Science
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• v.1 no.2
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• pp.79-87
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• 2016

Heavy metals such as cadmium (Cd) are highly toxic to aquatic organisms and human, even at trace concentration. Herein we investigated the effect of Cd on the gene expression of ATP-binding cassette (ABC) transporters and glutathione S-transferase (GST) in marine ciliate Euplotes crassus. Seven ABC transporters and one GST genes were partially cloned and sequences, and thereafter, transcriptional modulation of these genes after exposure to Cd for 8 h was investigated using quantitative real time RT- PCR (qRT-PCR). As results, sequence analysis and phylogenetic study revealed that E. crassus ABCs are likely typical ABC transports, in particular, B/C family, and GST gene may be similar to GST theta isoform. A significant increase in the expression of ABCs, except for ABCB21 was observed in a concentration dependent manner after exposure to Cd (0.1 and 0.5 mg/l) for 8 h. The GST mRNA level was the highest at 0.5 mg/l Cd and then reduced until control level. These findings suggest that ABCs and GST may be involved in a protective mechanism against Cd-mediated toxicity in E. crassus.

• Journal of Microbiology and Biotechnology
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• v.14 no.3
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• pp.635-638
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• 2004

An ATP-binding-cassette (ABC) transporter gene in the cephabacin biosynthetic gene cluster of Lysobacter lactamgenus was characterized. The amplified orf10 (cpbJ) gene was subcloned into pET-28a(+) vector and expressed in E. coli BL21(DE3) strain by 0.5 mM IPTG at $30^{\circ}C$. The membrane fraction of recombinant E. coli cells was separated by ultracentrifugation, and solubilized using 2.5% octyl-$\beta$-D-glucoside. Using the solubilized membrane fraction, the artificial proteoliposomes were reconstituted and analyzed for the biological activity of CpbJ protein. Upon measuring ATPase activity, the proteoliposome made from recombinant E. coli membrane proteins showed slightly higher activity than that from host E. coli membrane proteins. In the measurement of membrane transport activity, the reconstituted proteoliposome of recombinant E. coli membrane proteins exhibited higher activity when both substrates of cephalosporin C and L-Ala-L-Ser were applied, compared to the case of cephalosporin C or L-Ala-L-Ser only. It implies that the CpbJ protein is an ABC transporter secreting cephabacin antibiotics synthesized in cytoplasm.

• Korean Journal of Plant Resources
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• v.22 no.3
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• pp.273-280
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• 2009

A cDNA clone encoding a MDR-like ABC transporter protein was isolated from Brassica rapa seedlings, through rapid amplification of cDNA ends (RACE). This gene (named as Brmdr 1; GenBank accession no.: DQ296184 ) had a total length of 4222 bp with an open reading frame of 3900 bp, and encoded a predicted polypeptide of 1300 amino acids with a molecular weight of 143.1 kDa. The BrMDR1 protein shared 71.0, 62.5, 60.0 and 58.2% identity with other MDR proteins isolated from Arabidopsis thaliana (AAN28720), Coptis japonica (CjMDR), Gossypium hirsutum (GhMDR) and Triticum aestivum (TaMDR) at amino acid level, respectively. Southern blot analysis showed that Brmdr1 was a low-copy gene. Expression pattern analysis revealed that Brmdr1 constitutively expressed in the root, stem petals and stamens, but with lower expression in leaves and open flowers. The domains analysis showed that BrMDR1 protein possessed two transmembrane domains (TMDs) and two nucleotide binding domains (NBDs) arranging in "TMD1-NBD1-TMD2-NBD2" direction, which is consistent with other MDR transporters. Within NBDs three characteristic motifs common to all ABC transporters, "Walker A", "Walker B" and C motif, were found. These results indicate that BrMDR1 is a MDR-like ABC transporter protein that may be involved in the transport and accumulation of secondary metabolites.

• Korean Journal of Environmental Agriculture
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• v.31 no.1
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• pp.30-36
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• 2012

BACKGROUND: The resistance of rice bakanae disease pathogens against the fungicide prochloraz has been reported. Understanding the resistance mechanisms is an important for better control of the pathogens. In the present study, we investigated the resistance mechanisms of Fusarium fujikuroi CF337 (CF337) against prochloraz. METHODS AND RESULTS: Morphological changes in the cell wall of CF337 grown in potato dextrose broth (PDB) with or without prochloraz was investigated by transmission electron microscopy. Growth inhibition of CF337 was examined in PDB containing prochloraz or an ABC transporter inhibitor or both of them. Cell wall thickness of CF337 grown in PDB with prochloraz was significantly increased from $80.73{\pm}1.99nm$ to $193.11{\pm}7.07nm$. Significant inhibition in the growth of CF337 was observed in the presence of both prochloraz and the inhibitor, but no growth inhibition was observed in the presence of the inhibitor or prochloraz. Sequence analysis of ATP-binding cassette transporter (ABC) gene of CF337 showed 70 to 80% similarities to the genes of the pathogens resistant to other fungicides. CONCLUSION: Efflux transporter system and changes in cell wall thickness were suggested as resistance mechanisms of CF337 against prochloraz.

• Parasites, Hosts and Diseases
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• v.53 no.3
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• pp.335-339
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• 2015

Cryptosporidium andersoni ATP-binding cassette (CaABC) is an important membrane protein involved in substrate transport across the membrane. In this research, the nucleotide binding domain (NBD) of CaABC gene was amplified by PCR, and the eukaryotic expression vector of pEGFP-C1-CaNBD was reconstructed. Then, the recombinant plasmid of pEGFP-C1-CaNBD was transformed into the mouse intestinal epithelial cells (IECs) to study the iron transportation function of CaABC. The results indicated that NBD region of CaABC gene can significantly elevate the transport efficiency of $Ca^{2+}$, $Mg^{2+}$, $K^+$, and $HCO_3{^-}$ in IECs (P<0.05). The significance of this study is to find the ATPase inhibitors for NBD region of CaABC gene and to inhibit ATP binding and nutrient transport of CaABC transporter. Thus, C. andersoni will be killed by inhibition of nutrient uptake. This will open up a new way for treatment of cryptosporidiosis.