Soybean MAPK, GMK1 Is Dually Regulated by Phosphatidic Acid and Hydrogen Peroxide and Translocated to Nucleus during Salt Stress

임종희,안정선,이형석,Jitae Kim,김호방 한국분자세포생물학회 2012 Molecules and cells Vol.34 No.3

Mitogen-activated protein kinase (MAPK) is activated by various biotic and abiotic stresses. Salt stress induces two well-characterized MAPK activating signaling molecules, phosphatidic acid (PA) via phospholipase D and phospholipase C, and reactive oxygen species (ROS) via nicotinamide adenine dinucleotide phosphate (NADPH)-oxidase. In our previous study, the activity of soybean MAPK, GMK1 was strongly induced within 5 min of 300 mM NaCl treatment and this early activity was regulated by PA. In this study, we focused on the regulation of GMK1 at the later stage of the salt stress, because its activity was strongly persistent for up to 30 min. H2O2 activated GMK1 even in the presence of PA generation inhibitors, but GMK1 activity was greatly decreased in the presence of diphenyleneiodonium, an inhibitor of NADPH-oxidase after 5 min of the treatment. On the contrary, the n-butanol and neomycin reduced GMK1 activity within 5 min of the treatment. Thus, GMK1 activity may be sustained by H2O2 10 min after the treatment. Further, GMK1 was translocated into the nucleus 60 min after NaCl treatment. In the relationship between GMK1 and ROS generation, ROS generation was reduced by SB202190, a MAPK inhibitor, but was increa-sed in protoplast overexpressing TESD-GMKK1. However, these effects were occurred at prolonged time of NaCl treatment. These data suggest that GMK1 indirectly regulates ROS generation. Taken together, we propose that soybean GMK1 is dually regulated by PA and H2O2 at a time dependant manner and translocated to the nucleus by the salt stress signal.

어린이 비만예방을 위한 ‘건강 다이어리’ 활용 교육 프로그램 개발과 효과

임종희,이해정,김정원 한국실과교육연구학회 2009 實科敎育硏究 Vol.15 No.4

본 연구는 ‘건강 다이어리‘ 활용 교육 프로그램을 개발하여 이를 학교 교육 현장에 적용함으로써 어린이 스스로 올바른 식습관을 형성하는데 도움이 되도록 하며, 어린이 비만을 예방하는데 도움을 주고자 하는데 있다. 연구의 목적은 ’건강 다이어리‘ 프로그램을 위한 12차시 교육과정을 개발하고 이를 체계적인 교육을 통해 초등학생에게 적용 후 식습관 및 영양지식 향상과 비만도 개선의 효과 여부를 연구하고 분석하는 것이다. 연구결과 실험군 아동들의 비만도는 사전 19.06에서 사후 18.80으로 유의한 수준(p<0.1)의 개선이 이루어진 것으로 나타났으며, 영양지식은 사전 평균 5.13에서 사후 평균 6.84로 유의하게 향상되었다(p<0.01). 연구 결과와 같이 ’건강 다이어리‘를 활용한 교육 프로그램이 영양지식 향상과 비만도 개선에 효과적인 것으로 입증된 바, 초등학교 저학년 때부터 체계적인 교육프로그램을 토대로 지속적인 영양교육을 실시함으로써 사회적으로 문제가 되고 있는 아동비만을 개선하고 예방하는 것이 필요하다. The purpose of this research was to develop an dietary education program for children using 'Health Diary' by which they can keep their body healthy for themselves and eventually acquire adequate dietary habit and prevent obesity. The results of the research were as below. First, a 12-hour curriculum using 'Health Diary' was developed in this research , and after the application of this program to the school, it was found effective in improving the dietary knowledge and prevention of children's obesity. Second, the dietary knowledge of the children was improved significantly by this program(p<0.01). The dietary knowledge of the children rose from 5.13 before to 6.84 after the education, and it shows that this developed curriculum is effective in improving the dietary knowledge. Third, the obesity rate of children was significantly improved from 19.06 to 18.00 in BMI after the application of this curriculum(p<0.1). And it could be regarded as meaningful by considering the short period of application for children. The improvement of children's diet knowledge can be achieved in a short term, but it needs continuous training and practice to improve the obesity rate up to a meaningful level by keeping desirable dietary habits and practicing in every day life. The curriculum developed in this study by using 'Health Diary' was proved to be very useful for the improvement of the children's dietary knowledge, and the no significance in improving children's obesity could be due to the short application period. Therefore, lulder application of the program would be needed to improve and prevent the children's obesity. Also, it would be much effective that they should be educated continuously by systematic education program from the lowest to the highest grades in the school.

Induction of Drought Stress Resistance by Multi-Functional PGPR Bacillus licheniformis K11 in Pepper

임종희,김상달 한국식물병리학회 2013 Plant Pathology Journal Vol.29 No.2

Drought stress is one of the major yield affecting factor for pepper plant. The effects of PGPRs were analyzed in relation with drought resistance. The PGPRs inoculated pepper plants tolerate the drought stress and survived as compared to non-inoculated pepper plants that died after 15 days of drought stress. Variations in protein and RNA accumulation patterns of inoculated and noninoculated pepper plants subjected to drought conditions for 10 days were confirmed by two dimensional polyacrylamide gel electrophoresis (2D-PAGE) and differential display PCR (DD-PCR), respectively. A total of six differentially expressed stress proteins were identified in the treated pepper plants by 2D-PAGE. Among the stress proteins, specific genes of Cadhn, VA, sHSP and CaPR-10 showed more than a 1.5-fold expressed in amount in B. licheniformis K11-treated drought pepper compared to untreated drought pepper. The changes in proteins and gene expression patterns were attributed to the B. licheniformis K11. Accordingly, auxin and ACC deaminase producing PGPR B. licheniformis K11could reduce drought stress in drought affected regions without the need for overusing agrochemicals and chemical fertilizer. These results will contribute to the development of a microbial agent for organic farming by PGPR.

Biocontrol of Phytophthora Blight of Red Pepper Caused by Phytophthora capsici Using Bacillus subtilis AH18 and B. licheniformis K11 Formulations

임종희,김상달 한국응용생명화학회 2010 Applied Biological Chemistry (Appl Biol Chem) Vol.53 No.6

Bacillus subtilis AH18 and B. licheniformis K11 are multi-functional plant growth-promoting rhizobacteria (PGPR) that promote growth of red peppers and suppress phytophtora blight caused by Phytophtora capsici. A formulation of the two Bacillus strains was developed by producing heat-shocked endospores that had increased heat resistance. For large-scale cultivation in an 8-L jar fermentor, optimal culture conditions were pH 6.8, fermentation temperature of 30, and agitation speed of 180 rpm, with antifoaming agents added at a ventilation condition of 0.8 vvm. Induction of vegetative growth was achieved using 5.0×10−6 M dipicolinic acid and 7.0×10−4 M CaCl2. The subsequent large-scale cultivation produced viable cell numbers of 109 colony forming units/mL. High and low temperature stability tests of pilot products subjected to extreme storage conditions in accordance with thermal stability test standards revealed that storage for up to three years was possible without adverse effects. The combined use of two Bacillus microbial agents in vivo synergistically suppressed phytophthora blight, even following storage of the bacterial formulation at 54oC for 6 weeks. In field tests involving a phytophthora blight outbreak in red pepper, 40% of untreated peppers died, whereas 80% of peppers treated with the bacterial formulation survived for 65 days. The results indicate the potential of the B. subtilis AH18 and B. licheniformis K11 co-treatment for organic pepper farming.

랫드에서 Alginase의 급성 및 4주간 정맥 내 반복투여 독성시험에 관한 연구

임종희,남정석,제정환,이광훈,이학모,이원우,이병희,정지윤,박재학,이영순,Ihm, Jong-Hee,Nam, Jeong-Seok,Che, Jeong-Hwan,Li, Guang-Xun,Lee, Hak-Mo,Lee, Won-Woo,Yi, Beoung-Hi,Jung, Ji-Youn,Park, Jae-Hak,Lee, Yong-Soon 한국독성학회 1998 Toxicological Research Vol.14 No.3

Alginase$Alginase^{ⓡ}$ (Arginine esterase) is one of the snake venoms which is mainly consisted of arginine esterase and acts as a thrombus -forming inhibitor/thrombus-lysin. These present studies were performed to investigate of the acute and subacute toxicity of the Alginase$Alginase^{ⓡ}$ in rats. In acute toxicity study, rats were single administered intravenously with dosages of 0.001, 0.01. 0.1, 1 and 10U/kg B.W. and examined the number of death, clinical sign, body weight and pathological change for 7days after administration of Alginase$Alginase^{ⓡ}$. At maximum dose level (10U/kg B.W.), Alginase$Alginase^{ⓡ}$ induced symptoms of shock with cyanosis and dyspnea. But these symptoms dissappeared after 30~50 minutes and we could not find any other toxic effect in rats. Therefore, $LD_{50}$ Value of Alginase was over 10U/kg B.W. in rats. In four-week intravenous toxicity study of Alginase$Alginase^{ⓡ}$, rats were administered intravenously seven days per week for 28 days, with dosages of 0, 0.0125, 0.125 and 1.25U/kg B.W./day, respectively. Alginase$Alginase^{ⓡ}$ did not caused any death and showed any clinical signs in rats. No significant Alginase$Alginase^{ⓡ}$ -related changes were found in feed uptake, water consumption, hematology, serum biochemistry, urinalysis, ocular examination, organ weight and histopathological examination. From the results, Alginase$Alginase^{ⓡ}$ seems not to have any toxic effect in rats when it were given daily intravenous injections below the dosage 1.25U/kg B.W./day for four weeks.

Selection and Characterization of the Bacteriocin-Producing Bacterium, Bacillus subtilis BP6 Isolated from Chicken Gut against Salmonella gallinarum Causing Fowl-typhus

임종희,Sang-Dal Kim 한국응용생명화학회 2009 Journal of Applied Biological Chemistry (J. Appl. Vol.52 No.1

To select the antagonistic microorganisms against Salmonella gallinarum causing fowl typhoid, 31 strains were isolated from the gut of Korean native chicken. Among these, six strains could inhibit the growth of S. gallinarum. Strain BP6, which had strong inhibition activity against S. gallinarum, was identified as Bacillus subtilis BP6 by Biolog system and 16s rDNA sequencing. B. subtilis BP6 had antimicrobial activity toward various food-born pathogen such as Micrococcus luteus, B.cereus, Vibrio parahaemoliticus, Staphylococcus aureus, Listeria monocytogenes, and Escherichia coli O-157. The purified bacteriocin BP6 showed a single band of 20.7 kDa on sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Furthermore, low (<10 kDa) and high molecular weight (>10 kDa) substances from culture supernatant also inhibited the growth of S. gallinarum simultaneously. When the purified bacteriocin BP6 and n-butanol extract of non-peptide antibiotic-like substances were treated with protease, proteinase K, and trypsin, the antimicrobial activity of purified bacteriocin BP6 was lost, whereas that of n-butanol extract was not. The antimicrobial activity of purified bacteriocin BP6, however, was not affected by the α-amylase treatment. The bacteriocin BP6 was heat-stable and had a bacteriolytic mode of action resulting in the degradation of the S. gallinarum cell wall, suggesting the bacteriocin-producing bacterium B.subtilis BP6 could be a non-antibiotic probiotic for prevention of chicken or duck typhus.

OsbZIP75 positively regulates plant defense against the bacterial leaf blight pathogen Xanthomonas oryzae pv. oryzae

임종희,이상구,이은혜,박상렬,안일평,황덕주 한국식물생명공학회 2019 Plant biotechnology reports Vol.13 No.6

Basic leucine zipper (bZIP) proteins are involved in various biological processes, including biotic and abiotic stress responses. In response to biotic stress in rice (Oryza sativa L.), the roles of the TGA class of OsbZIP proteins are well-character-ized, whereas those of the non-TGA class of OsbZIP proteins remain largely unknown. Here, we analyze the function of OsbZIP75/RF2a in defense against Xanthomonas oryzae pv. oryzae (Xoo). The expression of OsbZIP75/RF2a was increased in response to infection with an incompatible Xoo race at 6 h post-inoculation, and rice plants overexpressing OsbZIP75/RF2a showed reduced symptom than wild-type plants upon infection with a compatible Xoo race. Defense related genes, especially OsPR10a, were up-regulated in OsbZIP75OX plants, and OsbZIP75/RF2a activated the OsPR10a promoter in a transient expression assay. Overall, these results indicate that OsbZIP75/RF2a acts as a positive regulator of defense against Xoo.

A Salt Stress-activated Mitogen-activated Protein Kinase in Soybean Is Regulated by Phosphatidic Acid in Early Stages of the Stress Response

임종희,이형석,Jitae Kim,Ho Bang Kim,Kim Seyoung,김병문,안정선 한국식물학회 2012 Journal of Plant Biology Vol.55 No.4

Salt stress inhibits plant growth and development and plants activate kinase-dependent survival pathways in response to salt stress. However, the role of soybean mitogenactivated protein kinases (MAPKs) in salt stress response has yet to be characterized. In this study, we found that salt stress activates Glycine max MAP kinase 1 (GMK1), a soybean MAPK. The activity of GMK1 induced with increasing salt concentrations, up to 300 mM NaCl, after 5 min of the treatment and was regulated by post-translational modification. We found that mastoparan, a heteromeric G-protein activator,also activated GMK1, and that n-butanol, a phospholipase D inhibitor, and neomycin, a phospholipase C inhibitor, inhibited its activity. Moreover, GMK1 activity was reduced by suramin,a heteromeric G-protein inhibitor, and by two inhibitors of phosphatidic acid (PA) generation after 5 min of 300 mM NaCl treatment. Endogenous PA levels were highest 5 min after induction of salt stress, and exogenous PA directly activated GMK1. From these data, we propose that salt stress signaling is transduced from heteromeric G-protein to GMK1 via phospholipases in the early stages of the response to salt stress in soybean.

Genetic Monitoring of Multi-Functional Plant Growth Promoting Rhizobacteria Bacillus subtilis AH18 and Bacillus licheniformis K11 by Multiplex and Real-Time Polymerase Chain Reaction in a Pepper Farming Field

임종희,Chang-Hwan Ahn,Hee-Young Jeong,Yo-Hwan Kim,Sang-Dal Kim 한국응용생명화학회 2011 Applied Biological Chemistry (Appl Biol Chem) Vol.54 No.2

The plant growth promoting rhizobacteria (PGPR) strains Bacillus subtilis AH18 and Bacillus licheniformis K11 were selected as biocontrol agents for the suppression of phytophthora blight caused by Phytophthora capsici. A genetic monitoring method was developed utilizing multiplex and real-time polymerase chain reaction (PCR) in a pepper farming soil. 2,3-Dihydro-2,3-dihydroxy benzoate dehydrogenase of a key siderophore synthesis enzyme (sid), auxin efflux carrier gene (aec), and cellulase gene (cel) of B. subtilis AH18 were used as genetic methods to monitor the presence and concentration of the inoculated microbial agents. Monitoring of B.licheniformis K11 was carried out by amplification of a cellulase gene (celK), siderophore synthase gene (dhbF), and iturin synthase A gene (ituA). B. subtilis AH18 and B. lichenifomis K11 could be detected upto 20 days by multiplex PCR in pepper farming soil. B. subtilis AH18 sid and three Bacillus lichenifomis K11 genes could be detected upto week 5 by real-time PCR in the natural unsterilized soil. P. capsici pathogen became nearly undetectable after 20 days biocontrol treatment in the field soil. These results could lead to the development of select PGRPs as useful microbial agents for the organic farming of peppers.

Soybean mitogen-activated protein kinase GMK2 is activated with GMK1 in Bradyrhizobium-Soybean interactions

임종희,안정선,손승민,이형석,김호방 한국유전학회 2014 Genes & Genomics Vol.36 No.6

Biological nitrogen fixation in root nodules isthe primary nitrogen source for plants. In leguminousplants, nodulation is initiated by the recognition of thenodulation (Nod) factor, the signaling molecule secreted byrhizobia. Our previous study showed that 47-kDa and44-kDa MAPKs were activated in soybean treated with agenistein-induced culture filtrate (GCF) of Bradyrhizobiumjaponicum. We investigated the activity and regulation ofthe 47-kDa MAPK, GMK1; however, the 44-kDa MAPKwas not extensively studied. Herein, we identified the44-kDa MAPK as Glycine max MAP kinase 2 (GMK2) andshowed that its transcription was activated by GCF. GMK1and 44-kDa MAPK were activated by phosphatidic acid(PA) and GCF-mediated induction of GMK1 and GMK2activities were reduced by treatment with PA generationinhibitors. The activity of GCF-activated GMK2, but notGMK1, was decreased by calcium signaling inhibitors. Ourdata indicate that GMK2 expression is regulated at thetranscriptional level and that GMK2 activity is regulated byphosphatidic acid and calcium during soybean-Bradyrhizobiuminteractions.