• BMB Reports
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• 제41권9호
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• pp.645-650
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• 2008

Nucleoside diphosphate kinase (NDPK) is involved in multiple signaling pathways in mammalian systems, including G-protein signaling. Arabidopsis NDPK2, like its mammalian counterparts, is multifunctional despite its initial discovery phytochrome-interacting protein. This similarity raises the possibility that NDPK2 may play a role in G-protein signaling in plants. In the present study, we explore the potential relationship between NDPK2 and the small G proteins, Pra2 and Pra3, as well as the heterotrimeric G protein, GPA1. We report a physical interaction between NDPK2 and these small G proteins, and demonstrate that NDPK2 can stimulate their GTPase activities. Our results suggest that NDPK2 acts as a GTPase-activating protein for small G proteins in plants. We propose that NDPK2 might be a missing link between the phytochrome-mediated light signaling and G protein-mediated signaling.

• KSBB Journal
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• 제21권1호
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• pp.16-19
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• 2006

최근의 연구에 의하면 열이나, 산화적 스트레스에 대해서 NDPK는 구조적인 변화를 유발하며, 효소 활성과 구조가 oxidant에 의해 변화된다는 보고를 근거로 하여 정상적인 효모균주와 효모의 NDPK 유전자가 파괴된 mutant에서, 산화적 스트레스에 관련된 역할을 규명하고자 2-D 전기영동 방법을 통해서, $H_2O_2$의 처리전과 처리 후에 전사패턴이 변화된 유전자들, 즉, 산화적조절 신호체제에 연관되어졌을 것이라고 생각되어지는 몇 개의 단백질 리스트를 얻었다. 이 결과는 NDPK의 redox state의 조절에 관련된 효소의 성질을 규명함에 있어 유용한 유전자 신호 체제정보를 제공할 것으로 생각되어진다.

• 한국생명과학회:학술대회논문집
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• 한국생명과학회 2005년도 국제학술심포지움 The 44th Annual Meeting of Korean Society for Life Science
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• pp.23-36
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• 2005

Bax, a mammalian pro-apoptotic member of the Bcl-2 family, induces cell death when expressed In yeast. To investigate whether .Bax expression can induce cell death in plant, we produced transgenic Arabidopsis plants that contained murine Bax cDNA under control of a glucocorticoid-inducible promoter. Transgenic plants treated with dexamethasone, a strong synthetic glucocorticoid, induced Bax accumulation and cell death, suggesting that some elements of cell death mechanism by Bax may be conserved among various orgarusms. Therefore, we developed novel yeast genetic system, and cloned several Plant Bax Inhibitors (PBIs). Here, we report the function of two PBIs In detail. PBIl is ascorbate peroxidase (sAPX). Fluorescence method of dihydrorhodamine123 oxidation revealed that expression of Bax in yeast cells generated reactive oxygen species (ROS), and which was greatly reduced by co-expression with sAPX. These results suggest that sAPX inhibits the generation of ROS by Bax, which in turn suppresses Bax-induced cell death in yeast. PBI2 encodes nucleoside diphosphate kinase (NDPK). ROS stress strongly induces the expression of the NDPK2 gene in Arabidopsis thaliana (AtNDPK2). Transgenic plants overexpressing AtNDPK2 have lower lovels of ROS than wildtype plants. Mutants lacking AtNDPK2 had higher levels of ROS than wildtype. H$_{2O2}$ treatment induced the phosphorylation of two endogenous proteins whose molecular weights suggested they are AtMPK3 and AtMPK6. In the absence of H2O2 treatment, phosphorylation of these proteins was slightly elevated in plants overexpressing AtNDPK2 but markedly decreased In the AtNDPK2 deletion mutant. Yeast two-hybrid and in vitro protein pull-down assays revealed that AtNDPK2 specifically interacts with AtMPK3 and AtMPK6. Furthermore, AtNDPK2 also enhances the MBP phosphorylation activity of AtMPK3 i'n vitro. Finally, constitutive overexpression of AtNDPK2 in Arabidopsis plants conferred an enhanced tolerance to multiple environmental stresses that elicit ROS accumulation In situ. Thus, AtNDPK2 appears to play a novel regulatory role in H2O2-mediated MAPK signaling in plants.

• 한국초지조사료학회지
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• 제26권2호
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• pp.83-90
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• 2006

환경스트레스에 내성을 갖는 버즈풋 트레포일(Lotus corniculatus L.) 형질전환체를 개발하기 위하여, AtNDPK 유전자가 E35S 프로모터에 의해 조절되도록 재조합한 발현벡터 pEN-K를 Agrobacterium 형질전환 방법으로 버즈풋 트레포일에 도입하였다. Agrobacterium과 공동배양한 버즈풋 트레포일 캘러스를 $100{\mu}g/m1$의 kanamycin 및 $500{\mu}g/m1$의 cefotaxim을 첨가한 SH-3-kc 배지에서 배양하며 식물체로 재분화시켰다. 재분화된 버즈풋 트레포일의 genomic DNA를 분리, PCR 및 Southern blot 분석을 실시한 결과, AtNDPK 유전자가 도입된 형질전환체의 경우 agarose gel 전기영동 및 X-ray 필름상에서 DNA band 및 hybridization signal을 확인할 수 있었으나, 형질전환되지 않은 대조구의 버즈풋 트레포일에서는 DNA band 및 hybridization signal이 관찰되지 않았다.

• 원예과학기술지
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• 제34권1호
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• pp.154-162
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• 2016

유전자 집적에 의해 아황산가스 저항성 증진 페튜니아를 개발하고자 비생물적스트레스 저항성 유전자로 널리 알려진 SOD2와 NDPK2유전자가 각각 도입된 SOD2 형질전환 계통 SOD2-2-1-1-35($T_4$)[S($T_4$)]와 NDPK2 형질전환 계통 NDPK2-7-1($T_2$)[N7-1($T_2$)]간 상호교잡을 실시하여 교잡 제1세대를 획득하였다. 교잡 제1세대 중 SOD2와 NDPK2유전자가 모두 집적된 개체 획득율은 32.1-73.0%이었고, SOD2와 NDPK2유전자가 모두 집적된 개체는 SOD2 또는 NDPK2유전자가 단독 도입된 개체에 비해 아황산가스 30ppm처리 피해율이 2.6-5.1배 낮아 아황산가스 저항성이 증진되었음을 확인하였다. 또한 아황산가스 저항성이 증진된 교잡 제1세대를 자가수분에 의해 세대진전 시켜 획득한 교잡 제2세대의 아황산가스 저항성 검정을 통하여 교잡 제1세대에서의 아황산가스 저항성이 후대에서도 안정적으로 발현함을 확인하였다. 또한, 광합성 효율 증진 조사를 통해 아황산가스 저항성이 증진된 SOD2와 NDPK2유전자 집적 후대가 아황산 가스에 대한 스트레스를 덜 받고 있음을 확인하였고, RT-qPCR분석을 통해 SOD2와 NDPK2유전자 집적 후대의 아황산가스 저항성 증진이 SOD2와 NDPK2 유전자 집적에 의한 효과임을 확인하였다.

• Journal of Plant Biotechnology
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• 제5권3호
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• pp.143-148
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• 2003

Bax, a mammalian pro-apoptotic member of the Bcl-2 family induces cell death when expressed in yeast. To investigate whether Bax expression can induce cell death in plant, we produced transgenic Arabidopsis plants that contained murine Bax cDNA under control of a glucocorticoid-inducible promoter. Transgenic plants treated with dexamethasone, a strong synthetic glucocorticoid, induced Bax accumulation and cell death, suggesting that some elements of cell death mechanism by Bax may be conserved among various organisms. Therefore, we developed novel yeast genetic system, and cloned several Plant Bax Inhibitors (PBIs). Here, we report the function of two PBIs in detail. PBI1 is ascorbate peroxidase (sAPX). Fluorescence method of dihydrorhodamine123 oxidation revealed that expression of Bax in yeast cells generated reactive oxygen species (ROS), and which was greatly reduced by co-expression with sAPX. These results suggest that sAPX inhibits the generation of ROS by Bax, which in turn suppresses Baxinduced cell death in yeast. PBI2 encodes nucleoside diphosphate kinase (NDPK). ROS stress strongly induces the expression of the NDPK2 gene in Arabidopsis thaliana (AtNDPK2). Transgenic plants overexpressing AtNDPK2 have lower levels of ROS than wildtype plants. Mutants lacking AtNDPK2 had higher levels of ROS than wildtype. $H_2O_2$ treatment induced the phosphorylation of two endogenous proteins whose molecular weights suggested they are AtMPK3 and AtMPK6. In the absence of $H_2O_2$ treatment, phosphorylation of these proteins was slightly elevated in plants overexpressing AtNDPK2 but markedly decreased in the AtNDPK2 deletion mutant. Yeast two-hybrid and in vitro protein pull-down assays revealed that AtNDPK2 specifically interacts with AtMPK3 and AtMPK6. Furthermore, AtNDPK2 also enhances the MSP phosphorylation activity of AtMPK3 in vitro. Finally, constitutive overexpression of AtNDPK2 in Arabidopsis plants conferred an enhanced tolerance to multiple environmental stresses that elicit ROS accumulation in situ. Thus, AtNDPK2 appears to playa novel regulatory role in $H_2O_2$-mediated MAPK signaling in plants.

• 한국식물생명공학회:학술대회논문집
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• 한국식물생명공학회 2003년도 식물바이오벤처 페스티발
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• pp.65-71
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• 2003

Bax, a mammalian pro-apoptotic member of the Bcl-2 family, induces cell death when expressed in yeast. To investigate whether Bax expression can induce cell death in plant, we produced transgenic Arabidopsis plants that contained murine Bax cDNA under control of a glucocorticoid-inducible promoter. Transgenic plants treated with dexamethasone, a strong synthetic glucocorticoid, induced Bax accumulation and cell death, suggesting that some elements of cell death mechanism by Bax may be conserved among various organisms. Therefore, we developed novel yeast genetic system, and cloned several Plant Bax Inhibitors (PBIs). Here, we report the function of two PBIs in detail. PBI1 is ascorbate peroxidase (sAPX). Fluorescence method of dihydrorho-damine 123 oxidation revealed that expression of Bax in yeast cells generated reactive oxygen species (ROS), and which was greatly reduced by co-expression with sAPX. These results suggest that sAPX inhibits the generation of ROS by Bax, which in turn suppresses Baxinduced cell death in yeast. PBI2 encodes nucleoside diphosphate kinase (NDPK). ROS stress strongly induces the expression of the NDPK2 gene in Arabidopsis thaliana (AtNDPK2). Transgenic plants overexpressing AtNDPK2 have lower levels of ROS than wildtype plants. Mutants lacking AtNDPK2 had higher levels of ROS than wildtype. $H_2O_2$ treatment induced the phosphorylation of two endogenous proteins whose molecular weights suggested they are AtMPK3 and AtMPK6. In the absence of $H_2O_2$ treatment, phosphorylation of these proteins was slightly elevated in plants overexpressing AtNDPK2 but markedly decreased in the AtNDPK2 deletion mutant. Yeast two-hybrid and in vitro protein pull-down assays revealed that AtNDPK2 specifically interacts with AtMPK3 and AtMPK6. Furthermore, AtNDPK2 also enhances the MBP phosphorylation activity of AtMPK3 in vitro. Finally, constitutive overexpression of AtNDPK2 in Arabidopsis plants conferred an enhanced tolerance to multiple environmental stresses that elicit ROS accumulation in situ. Thus, AtNDPK2 appears to play a novel regulatory role in $H_2O_2$-mediated MAPK signaling in plants.

• 화훼연구
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• 제19권4호
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• pp.269-273
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• 2011

본 연구는 환경스트레스 저항성이 증진된 페튜니아를 개발하기 위하여 NDPK2유전자 도입 형질전환 계통 NDPK2-7-1와 SOD2 유전자 도입 형질전환 계통 SOD2-2-1-1-35간의 교잡에 의해 획득된 후대들의 비생물적 스트레스 저항성을 조사하기 위해 수행되었다. 비생물적 스트레스 유발원인 메틸바이올로젠(methyl viologen, MV) $100{\mu}M$과 $200{\mu}M$ 처리에서 교잡후대들은 그들의 교배모본 SOD2 유전자나 NDPK2 유전자가 단독으로 도입된 형질전환 계통이나 비형질전환체 보다 메틸바이올로젠에 의한 피해를 적게 받았다. 이는 SOD2 유전자나 NDPK2 유전자가 단독으로 도입된 형질전환 계통간 교잡에 의해 획득된 후대들이 그들의 교배모본 (SOD2 유전자나 NDPK2 유전자가 단독으로 도입된 형질전환 계통)이나 비형질전환체 보다 산화적 스트레스에 대한 저항성이 증진되었음을 증명해 준다고 할 수 있다. 이들 교잡후대들은 초장 등 11종류의 양적형질의 특성이 비형질전환체에 비해 약간 길거나 짧긴 하였지만 비형질전환체와 거의 유사하였으며, 꽃 색갈이나 모양 또한 그들의 교배모본 (SOD2 유전자나 NDPK2 유전자가 단독으로 도입된 형질전환 계통)이나 비형질전환체와 차이가 없었다.

• 한국초지조사료학회지
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• 제25권4호
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• pp.281-286
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• 2005

환경스트레스에 내성을 갖는 버즈풋 트레포일(Lotus crniculatus L.) 형질전환체를 개발하기 위하여, AtNDPK 유전자가 SWPA2 프로모터에 의해 조절되도록 재조합한 발현벡터 pCAMBIA 2300 /SWPA2::AtNDPK2를 Agrobacterium 형질전환 방법으로 버즈풋 트레포일에 도입하였다. Apgrobacterium과 버즈풋 트레포일 캘러스의 공동배양한 캘러스를 $100{\mu}g/m1$의 kanamycin 및 $500{\mu}g/ml$의 cefotaxim을 첨가한 SH-3-kc 배지에서 배양하며 형질전환된 캘러스를 선발한 다음, BOi2Y 배지에서 2개월 이상 배양하며 식물체로 재분화시켰다. 재분화된 버즈풋 트레포일의 genomic DNA를 분리, PCR 및 Southern blot 분석을 실시한 결과, AtNDPK유전자를 도입한 형질전환체의 경우 agarose gel 전기영동 및 X-ray 필름상에서 DNA band 및 hybridization signal을 확인할 수 있었으나, 형질전환 되지 않은 대조구의 버즈풋 트레포일에서는 DNA band 및 hybridization signal이 관찰되지 않았다.

• Journal of Plant Biotechnology
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• 제38권3호
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• pp.215-220
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• 2011

This study was conducted to investigate how to enhance resistance to oxidative stress in petunia progeny obtained by a crossing between transgenic plants, MnSOD (SOD2) ($T_4$) and NDPK2 ($T_2$), to develop transgenic petunia much more resistant to environmental stress. At the treatment of MV 200 ${\mu}M$, the progeny was significantly less damaged than its parental plants (SOD2- or NDPK2-transgenic lines) as well as wild type plants, implying its resistance to oxidative stress was enhanced compare to that of SOD2- or NDPK2- transgenic plants. In an expression of 11 quantitative traits, the progeny remained similar to control plants, although it infrequently displayed slightly longer or wider than either parental or wild type plants. In the expression of 6 qualitative traits, there was no significant difference between parental or non-transgenic control plants.