Structural determinants for pyrabactin recognition in ABA receptors in Oryza sativa
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We determined the structure of OsPYL/RCAR3:OsPP2C50 complex with pyrabactin. Our results suggest that a less-conserved phenylalanine of OsPYL/RCAR subfamily I is one of considerations of ABA agonist development for Oryza sativa.
Pyrabactin is a synthetic chemical mimicking abscisic acid (ABA), a naturally occurring phytohormone orchestrating abiotic stress responses. ABA and pyrabactin share the same pocket in the ABA receptors but pyrabactin modulates ABA signaling differently, exhibiting both agonistic and antagonistic effects. To explore structural determinants of differential functionality of pyrabactin, we determined the crystal structure of OsPYL/RCAR3:pyrabactin:OsPP2C50, the first rice ABA receptor:co-receptor complex structure with a synthetic ABA mimicry. The water-mediated interaction between the wedging Trp-259 of OsPP2C50 and pyrabactin is lost, undermining the structural integrity of the ABA receptor:co-receptor. The loss of the interaction of the wedging tryptophan of OsPP2C with pyrabactin appears to contribute to the weaker functionality of pyrabactin. Pyrabactin in the OsPYL/RCAR3:OsPP2C50 complex adopts a conformation different from that in ABA receptors from Arabidopsis. Phe125, specific to the subfamily I of OsPYL/RCARs in the ABA binding pocket, appears to be the culprit for the differential conformation of pyrabactin. Although the gate closure essential for the integrity of ABA receptor:co-receptor is preserved in the presence of pyrabactin, Phe125 apparently restricts accessibility of pyrabactin, leading to decreased affinity for OsPYL/RCAR3 evidenced by phosphatase assay. However, Phe125 does not affect conformation and accessibility of ABA. Yeast two-hybrid, germination and gene transcription analyses in rice also support that pyrabactin imposes a weak effect on the control of ABA signaling. Taken together, our results suggest that phenylalanine substitution of OsPYL/RCARs subfamily I may be one of considerations for ABA synthetic agonist development.
KeywordsABA Pyrabactin ABA receptor Type 2C protein phosphatase Oryza sativa
We thank the staff members at beamline 5C, Pohang Accelerator Laboratory for technical assistance in crystallographic data collection. This work was supported by the Next-Generation BioGreen 21 Program (PJ01335001 and PJ01367602) through the Rural Development Agency and the National Research Foundation of Korea grants awarded by the Basic Science Research Program (NRF-2018R1A2B6004367) and the Science Research Center Program (SRC-2017R1A5A1014560).
SL conceived research plans and supervised experiments. SH and SL designed experiments. SH and YL performed most of the experiments and analyzed the data. EJP, MKM, YL, T-HK, and B-GK performed some experiments and analyzed the data for supporting main idea. SH, YL, and SL wrote the article with contributions of all the authors.
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