• The korean journal of orthodontics
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• v.33 no.3 s.98
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• pp.169-183
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• 2003

Tooth movement by orthodontic force effects great tissue changes within the periodontium, especially by shifting the blood flow in the pressure side and resulting in a hypoxic state of low oxygen tension. The aim of this study is to elucidate the possible mechanism of apoptosis in response to hypoxia in MC3T3El osteoblasts, the main cells in bone remodeling during orthodontic tooth movement. MC3T3El osteoblasts under hypoxic conditions ($2\%$ orygen) resulted in apoptosis in a time-dependent manner as estimated by DNA fragmentation assay and nuclear morphology stained with fluorescent dye, Hoechst 33258. Pretreatment with Z-VAD-FMK, a pancaspase inhibitor, or Z-DEVD-CHO, a specific caspase-3 inhibitor, completely suppressed the DNA ladder in response to hypoxia. An increase in caspase-3-like protease (DEVDase) activity was observed during apoptosis, but no caspase-1 activity (YVADase) was detected. To confirm what caspases are involved in apoptosis, Western blot analysis was performed using anti-caspase-3 or -6 antibodies. The 10-kDa protein, corresponding to the active products of caspase-3, and the 10-kDa protein of the active protein of caspase-6 were generated in hypoxia-challenged cells in which the processing of the full length form of caspase-3 and -6 was evident. While a time course similar to this caspase-3 and -6 activation was evident, hypoxic stress caused the cleavage of lamin A, which was typical of caspase-6 activity. In addition, the stress elicited the release of cytochrome c into the cytosol during apoptosis. Furthermore, we observed that pre-treatment with SB203580, a selective p38 mitogen activated protein kinase inhibitor, attenuated the hypoxia-induced apoptosis. The addition of SB203S80 suppressed caspase-3 and -6-like protease activity by hypoxia up to $50\%$. In contrast, PD98059 had no effect on the hypoxia-induced apoptosis. To confirm the involvement of MAP kinase, JNK/SAPK, ERK, or p38 kinase assay was performed. Although p38 MAPK was activated in response to hypoxic treatment, the other MAPK -JNK/SAPK or ERK- was either only modestly activated or not at all. These results suggest that p38 MAPK is involved in hypoxia-induced apoptosis in MC3T3El osteoblasts.

• Research in Plant Disease
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• v.10 no.4
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• pp.241-247
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• 2004

Recently, we reported a satellite RNA (Paf-satRNA) which is encapsidated in a pepper isolate of Cucumber mosaic virus (CMV-Paf) regulated symptom attenuation of the helper virus. To characterize mild symptom domain of Paf-satRNA, a series of chimeric cDNAs of satRNAs were created by using full-length cDNA clones of Paf-satRNA and a Pep-satRNA, chlorosis-inducing satRNA in pepper plants, and analyzed for determinants of symptom attenuation. When compared the nucleotide sequences, the 3' and 5' terminal sequences of the two wild-type (wt) satRNAs contained relatively conserved sequences which are the typical to CMV satRNA. Ten bases insertions were found in PepY-satRNA, and two variable regions, 81st to 113th and 183rd to 265th from the 5'-end, were located in the middle parts of the satRNAs. To delineate the attenuated symptom-related domain for the Paf-satRNA, in vitro transcripts RNAs transcribed from the wt cDNAs and constructed chimeric cDNAs were combined with genomic RNAs, RNA1, RNA2 and RNA3, of CMV-Fny and inoculated onto Nicotiana benthamiana plants. These transcripts were fully infectious onto the N. benthamiana and infectivity was confirmed by the RT-PCR. Chimeric Paf(H/N)-satRNA and PepY(N/A)-satRNA as well as Paf-satRNA induced very mild mosaic or symptomless infection on N. benthamiana. By contrast, typical mosaic symptom and stunting of infected plants were induced when PepY-satRNA, PepY(H/N)-satRNA and Paf(N/A)-satRNA were infected to N. benthamiana. Paf-satRNA coinfected with CMV-Fny RNAs induced very mild to sympomless on pepper plants whereas PepY-satRNA-infected pepper expressed typical chlorosis mosaic symptom. Two kinds of chimeric mutants, Paf(H/N)-satRNA and PepY(N/A)-satRNA, induced mild mosaic or symptomless infection onto pepper plants, while PepY(H/N)-satRNA and Paf(N/A)-satRNA showed typical chlorosis and mosaic symptom with stunting. This results suggest that mild symptom-related domain for the Paf-satRNA was located on HpaI-NarI region.

• Proceedings of the Korean Society of Plant Pathology Conference
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• 1995.06b
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• pp.77-96
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• 1995

To obtain basic informations for the improvement of seed potato production in Korea, some etiological properties of potato virus Y(PVY) distributed in the major seed potato production area(Daekwanryeong) were characterized, and the nucleotide and amino acid sequences of the coat protein gene of the PVY strains isolated were analyzed. PVY strains in Daekwonryeong, an alpine area, were identified to be two strains, PVYo and PVYN by symptoms of indicator plants, and their distribution in potato fields was similar. Major symptom on potato varieties by PVY was grouped as either mosaic alone or mosaic accompanied with veinal necrosis in the lower leaves. The symptom occurrence of the two symptoms was similar with Irish Cobbler, but Superior showed a higher rate of mosaic symptom than the other. The PVY strain which was isolated from potato cv. Superior showing typical mosaic symptoms produced symptoms of PVY-O on the indicator plants of Chenopodium amaranticolor, Nicotiana tabacum cv. Xanthi nc and Physalis floridana, but no symptom o Capsicum annum cv. Ace. Moreover, results from the enzyme-linked immunosorbent assay with monoclonal and polyclonal antibodies showed that the isolated PVY reacts strongly with PYV-O antibodies but does not react specifically with PVY-T antibodies. The purified virus particles were flexious with a size of 730$\times$11nm. On the basis of the above characteristics, the strain was identified to be a PVY-O and named as of PVY-K strain. The flight of vector aphids was observed in late May, however, the first occurrence of infected plants was in mid June with the bait plants surrounded with PVY-infected potato plants and early July with the bait plants surrounded with PVY-free potato plants. PVY infection rates by counting symptoms on bait plants (White Burley) were 1.1% with the field surrounded with PVY-free potato plants and 13.7% the fields surrounded with PVY-infected potato plants, showing the effect of infection pressure. The propagated PVY-K strain on tobacco(N. sylvestris) was purified, and the RNA of the virus was extracted by the method of phenol extraction. The size of PVY-K RNA was measured to be 9, 500 nucleotides on agarose gel electrophoresis. The double-stranded cDNAs of PVY-K coat protein(CP) gene derived by the method of polymerase chain reaction were transformed into the competent cells of E. coli JM 109, and 2 clones(pYK6 and pYK17) among 11 clones were confirmed to contain the full-length cDNA. Purified plasmids from pYK17 were cut with Sph I and Xba I were deleted with exonuclease III and were used for sequencing analysis. The PVY-K CP gene was comprised of 801 nucleotides when counted from the clevage site of CAG(Gln)-GCA(Ala) to the stop codon of TGA and encoded 267 amino acids. The molecular weight of the encoded polypeptides was calculated to be 34, 630 daltons. The base composition of the CP gene was 33.3% of adenine, 25.2% of guanine, 20.1% of cytosine and 21.4% of uracil. The polypeptide encoded by PVY-K CP gene was comprised of 22 alanines, 20 threonines, 19 glutamic acids and 18 glycines in order. The homology of nucleotide sequence of PVY-K CP gene with those of PVY-O(Japan), PVY-T(Japan), PVY-TH(Japan), PVYN(the Netherlands), and PVYN(France) was represented as 97.3%, 88.9%, 89.3%, 89.6% and 98.5%, respectively. The amino acid sequence homology of the polypeptide encoded by PVY-K CP gene with those encoded by viruses was represented as 97.4%, 92.5%, 92.9%, 92.9%, and 98.5%, respectively.

• Research in Plant Disease
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• v.7 no.3
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• pp.155-163
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• 2001

Two attenuated Cucumber mosaic virus (CMV) isolates, Paf-CMV and Rs2-CMV that had been selected from CMV isolates associated with satellite RNA (satRNA) were tested for cross-protection effect in pepper plants. The viruses selected as attenuated strains appeared to be identical serologically and physically to the challenge virus (Mf-CMV), but they were lower in the dilution end-point of infectivity of crude sap than Mf-CMV When symptoms were observed in several indicator plants after inoculation, Paf-CMV and Rs2-CMV were symptomless or showed mild mosaic symptoms while another satRNA isolate Ap-CMV developed severe mosaic symptoms on the leaves as Mf-CMV. The nucleotide sequences of the satRNAs were determined by sequencing full-length cDNA clones. Paf-, Rs2- and Ap-satRNAs were 386, 335, and 347 nucleotides long, respectively, The sequences were then compared with the other known Y-satRNA, revealing that nucleotide sequences of the satRNAs consisted of 5'- and 3'-terminal conserved regions. However variations occurred on the middle regions of the sequences, especially those related to symptom interference, showing significant differences between Paf-satRNA and other isolates. Infectious transcripts of Paf-satRNA and Rs2-satRNA induced mild mosaic symptoms in pepper plants when supported by genomic RNAs of Mf-CMV. Under greenhouse conditions, Paf-CMV and Rs2-CMV were tested for cross-protection effect in pepper and tobacco (Nicotiana tabacum cv, Xanthi nc) plants against Mf-CMV. No symptoms were developed on the plants vaccinated with Paf-CMV until 3 weeks after inoculation with the virulent strain; however another attenuated isolate, Rs2-CMV, showed less effectiveness in cross-protection. Depending on the concentration of the challenged virus, symptoms sometimes appeared later in the upper leaves. However, in plants challenged with low concentrations (below 0.2 mg/ml) of the challenge inoculum, symptoms caused by the virulent strain did not develop on the plants vaccinated with Paf-CMV. In the field experiments, the number of pepper plants with severe mosaic symptoms in the control plots was progressively increased after transplanting and reached approximately 50% after 50 days. On the other hand, the incidence of mosaic disease appeared very low on the plants that had received the protective inoculation with Paf-CMV.

• Journal of Plant Biotechnology
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• v.42 no.1
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• pp.25-33
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• 2015

A PyrcpPR-10 gene with differentially expressed was isolated by using the suppression subtractive hybridization assay between '93-3-98' (highly resistant against scab caused by Venturia nashicola) and 'Sweat Skin'(highly susceptible) and analyzed the expression pattern according to organs and cultivars. The full length of PyrcpPR-10 was cloned as 743bp with 480bp's ORP, and was determined to encode a protein of 159 amino acid residues. On analyzing PyrcpPR-10 gene sequence compared with resistant and susceptible cultivars, 'Hwangsilri' (resistant), 'Gamcheonbae' (moderately resistant), 'Wonhwang' (moderately susceptible), 'Niitaka' (highly susceptible), and 'Sweat Skin' (highly susceptible) had identical gene sequence but 'Bartlett' (highly resistant) showed partly different sequences. The deduced amino acid sequence showed 64 ~ 98% homology and had the GXGGXG motif to known amino acid of other plants PR-10 by the BLAST X analysis. Among several organs or tissues, petal was showed highest expression level of PyrcpPR-10 gene followed by leaf, floral axis, bud, and bark. The expression level of PyrcpPR-10 gene was dramatically increased at 24 hr after inoculation in all cultivars and also up-regulated in accordance with resistant degree of cultivars. While resistant cultivars ('Bartlett', '93-3-98', and 'Hwangsilri') induced relatively high expression level of PyrcpPR-10 gene, susceptible cultivars ('Niitaka', and 'Sweat Skin') showed low expression level. PyrcpPR-10 gene is assumed that it is directly connected with defense mechanisms to pear scab.