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Protein interactome analysis of 12 mitogen-activated protein kinase kinase kinase in rice using a yeast two-hybrid system

Authors

  • Raksha Singh,

    1. Department of Molecular Biology, College of Life Sciences, Sejong University, Gunja-dong, Gwangjin-gu, Seoul, Republic of Korea
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    • These authors contributed equally to this work.

  • Jae-Eun Lee,

    1. Department of Molecular Biology, College of Life Sciences, Sejong University, Gunja-dong, Gwangjin-gu, Seoul, Republic of Korea
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    • These authors contributed equally to this work.

  • Sarmina Dangol,

    1. Department of Molecular Biology, College of Life Sciences, Sejong University, Gunja-dong, Gwangjin-gu, Seoul, Republic of Korea
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  • Jihyun Choi,

    1. Department of Molecular Biology, College of Life Sciences, Sejong University, Gunja-dong, Gwangjin-gu, Seoul, Republic of Korea
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  • Ran Hee Yoo,

    1. Plant Systems Engineering Center, Korea Research Institute of Bioscience and Biotechnology, Daejeon, Republic of Korea
    2. Biosystems and Bioengineering Program, University of Science and Technology (UST), Yuseong-gu, Daejeon, Republic of Korea
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  • Jae Sun Moon,

    1. Plant Systems Engineering Center, Korea Research Institute of Bioscience and Biotechnology, Daejeon, Republic of Korea
    2. Biosystems and Bioengineering Program, University of Science and Technology (UST), Yuseong-gu, Daejeon, Republic of Korea
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  • Jae-Kyung Shim,

    1. Department of Molecular Biology, College of Life Sciences, Sejong University, Gunja-dong, Gwangjin-gu, Seoul, Republic of Korea
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  • Randeep Rakwal,

    1. Graduate School of Life and Environmental Sciences, University of Tsukuba, Tsukuba, Ibaraki, Japan
    2. Department of Anatomy I, School of Medicine, Showa University, Shinagawa, Tokyo, Japan
    3. Research Laboratory for Biotechnology and Biochemistry (RLABB), Kathmandu, Nepal
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  • Ganesh Kumar Agrawal,

    1. Research Laboratory for Biotechnology and Biochemistry (RLABB), Kathmandu, Nepal
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  • Nam-Soo Jwa

    Corresponding author
    1. Department of Molecular Biology, College of Life Sciences, Sejong University, Gunja-dong, Gwangjin-gu, Seoul, Republic of Korea
    • Correspondence: Professor Nam-Soo Jwa, Department of Molecular Biology, College of Life Sciences, Sejong University, Gunja-dong, Gwangjin-gu, Seoul, Republic of Korea

      E-mail: nsjwa@sejong.ac.kr

      Fax: +82-2-3408-4336

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  • Colour Online: See the article online to view Figs. 1–5 in colour.

Abstract

The mitogen-activated protein kinase (MAPK) cascade is composed at least of MAP3K (for MAPK kinase kinase), MAP2K, and MAPK family modules. These components together play a central role in mediating extracellular signals to the cell and vice versa by interacting with their partner proteins. However, the MAP3K-interacting proteins remain poorly investigated in plants. Here, we utilized a yeast two-hybrid system and bimolecular fluorescence complementation in the model crop rice (Oryza sativa) to map MAP3K-interacting proteins. We identified 12 novel nonredundant interacting protein pairs (IPPs) representing 11 nonredundant interactors using 12 rice MAP3Ks (available as full-length cDNA in the rice KOME (http://cdna01.dna.affrc.go.jp/cDNA/) at the time of experimental design and execution) as bait and a rice seedling cDNA library as prey. Of the 12 MAP3Ks, only six had interacting protein partners. The established MAP3K interactome consisted of two kinases, three proteases, two forkhead-associated domain-containing proteins, two expressed proteins, one E3 ligase, one regulatory protein, and one retrotransposon protein. Notably, no MAP3K showed physical interaction with either MAP2K or MAPK. Seven IPPs (58.3%) were confirmed in vivo by bimolecular fluorescence complementation. Subcellular localization of 14 interactors, together involved in nine IPPs (75%) further provide prerequisite for biological significance of the IPPs. Furthermore, GO of identified interactors predicted their involvement in diverse physiological responses, which were supported by a literature survey. These findings increase our knowledge of the MAP3K-interacting proteins, help in proposing a model of MAPK modules, provide a valuable resource for developing a complete map of the rice MAPK interactome, and allow discussion for translating the interactome knowledge to rice crop improvement against environmental factors.

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