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Screening of ABA Transporters by a Yeast Two-Hybrid System-Based Screening Using the Receptor Complex as a Sensor

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Abscisic Acid

Part of the book series: Methods in Molecular Biology ((MIMB,volume 2462))

Abstract

Abscisic acid (ABA) is a plant hormone that is involved in many physiological events and is present in most plant tissues. How ABA moves within plants has not been well understood. To characterize the physiological consequences as well as the underlying molecular events responsible for ABA movement, it is crucial to identify the transporter proteins. Several approaches have been successful in identifying ABA transporters. In this chapter, we outline a methodology to directly identify proteins capable of transporting ABA in a heterologous yeast system. In our assay, the ABA receptor [PYRABACTIN RESISTANCE1 (PYR1) and PYR1-LIKE (PYL), also known as REGULATORY COMPONENTS OF ABA RECEPTOR (RCAR)] and the protein phosphatases of type 2C (PP2C) coreceptor interact in an ABA-dependent manner. A yeast two-hybrid (Y2H) system is used to monitor interactions between the receptor and PP2C and, hence, the ABA concentration in the yeast cells. Screening cDNAs that induce or inhibit the receptor–PP2C interaction in the presence of ABA allows us to identify candidate transporters. ABA transport activities of the putative transporter proteins can be validated by quantifying hormone levels in the yeast cells using liquid chromatography tandem-mass spectrometry (LC-MS/MS).

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Authors and Affiliations

  1. RIKEN Center for Sustainable Resource Science, Yokohama, Kanagawa, Japan

    Shunsuke Watanabe, Yuri Kanno & Mitsunori Seo

Authors
  1. Shunsuke Watanabe
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  2. Yuri Kanno
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  3. Mitsunori Seo
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Corresponding author

Correspondence to Mitsunori Seo .

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Editors and Affiliations

  1. Max-Planck-Institut für Molekulare Pflanzenphysiologie, Potsdam-Golm, Germany

    Takuya Yoshida

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Watanabe, S., Kanno, Y., Seo, M. (2022). Screening of ABA Transporters by a Yeast Two-Hybrid System-Based Screening Using the Receptor Complex as a Sensor. In: Yoshida, T. (eds) Abscisic Acid. Methods in Molecular Biology, vol 2462. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-2156-1_7

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  • DOI: https://doi.org/10.1007/978-1-0716-2156-1_7

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  • Publisher Name: Humana, New York, NY

  • Print ISBN: 978-1-0716-2155-4

  • Online ISBN: 978-1-0716-2156-1

  • eBook Packages: Springer Protocols

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