Clade A protein phosphatase 2Cs (PP2Cs) are abscisic acid (ABA) co-receptors that block ABA signalling by inhibiting the downstream protein kinases. ABA signalling is activated after PP2Cs are inhibited by ABA-bound PYR/PYL/RCAR ABA receptors (PYLs) in Arabidopsis. However, whether these PP2Cs are regulated by other factors remains unknown. Here, we report that ABI1 (ABA-INSENSITIVE 1) can interact with the U-box E3 ligases PUB12 and PUB13, but is ubiquitinated only when it interacts with ABA receptors in an in vitro assay. A mutant form of ABI1-1 that is unable to interact with PYLs is more stable than the wild-type protein. Both ABI1 degradation and all tested ABA responses are reduced in pub12 pub13 mutants compared with the wild type. Introducing the abi1-3 loss-of-function mutation into pub12 pub13 mutant recovers the ABA-insensitive phenotypes of the pub12 pub13 mutant. We thus uncover an important regulatory mechanism for regulating ABI1 levels by PUB12 and PUB13.

Cold stress is a major environmental factor that adversely affects plant growth and development. The C-repeat binding factor/DRE binding factor 1 (CBF/DREB1) transcriptional regulatory cascade has been shown to play important roles in plant response to cold. Here we demonstrate that two key components of brassinosteroid (BR) signaling modulate freezing tolerance of Arabidopsis plants. The loss-of-function mutant of the GSK3-like kinases involved in BR signaling, bin2-3 bil1 bil2, showed increased freezing tolerance, whereas overexpression of BIN2 resulted in hypersensitivity to freezing stress under both non-acclimated and acclimated conditions. By contrast, gain-of-function mutants of the transcription factors BZR1 and BES1 displayed enhanced freezing tolerance, and consistently cold treatment could induce the accumulation of dephosphorylated BZR1. Biochemical and genetic analyses showed that BZR1 acts upstream of CBF1 and CBF2 to directly regulate their expression. Moreover, we found that BZR1 also regulated other COR genes uncoupled with CBFs, such as WKRY6, PYL6, SOC1, JMT, and SAG21, to modulate plant response to cold stress. Consistently, wrky6 mutants showed decreased freezing tolerance. Taken together, our results indicate that BZR1 positively modulates plant freezing tolerance through CBF-dependent and CBF-independent pathways.

A better understanding of the extent of convergent selection among crops could greatly improve breeding programs. We found that the quantitative trait locus KRN2 in maize and its rice ortholog, OsKRN2 , experienced convergent selection. These orthologs encode WD40 proteins and interact with a gene of unknown function, DUF1644, to negatively regulate grain number in both crops. Knockout of KRN2 in maize or OsKRN2 in rice increased grain yield by ~10% and ~8%, respectively, with no apparent trade-offs in other agronomic traits. Furthermore, genome-wide scans identified 490 pairs of orthologous genes that underwent convergent selection during maize and rice evolution, and these were enriched for two shared molecular pathways. KRN2 , together with other convergently selected genes, provides an excellent target for future crop improvement.

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PHYTOCHROME-INTERACTING FACTORS (PIFs) are a group of basic helix-loop-helix transcription factors that can physically interact with photoreceptors, including phytochromes and cryptochromes. It was previously demonstrated that PIFs accumulated in darkness and repressed seedling photomorphogenesis, and that PIFs linked different photosensory and hormonal pathways to control plant growth and development. In this study, we show that PIFs positively regulate the ABA signaling pathway during the seedling stage specifically in darkness. We found that PIFs positively regulate ABI5 transcript and protein levels in darkness in response to exogenous ABA treatment by binding directly to the G-box motifs in the ABI5 promoter. Consistently, PIFs and the G-box motifs in the ABI5 promoter determine ABI5 expression in darkness, and overexpression of ABI5 could rescue the ABA-insensitive phenotypes of pifq mutants in the dark. Moreover, we discovered that PIFs can physically interact with the ABA receptors PYL8 and PYL9, and that this interaction is not regulated by ABA. Further analyses showed that PYL8 and PYL9 promote PIF4 protein accumulation in the dark and enhance PIF4 binding to the ABI5 promoter, but negatively regulate PIF4mediated ABI5 activation. Taken together, our data demonstrate that PIFs interact with ABA receptors to orchestrate ABA signaling in darkness by controlling ABI5 expression, providing new insights into the pivotal roles of PIFs as signal integrators in regulating plant growth and development.