Molecular characterization of maize bHLH transcription factor (ZmKS), a new ZmOST1 kinase substrate.

In order to identify potential substrates of the maize kinase in the ABA signalling network, ZmOST1 was used as bait against a library of cDNAs from dehydrated young leaves. A ZmOST1-interactive polypeptide ZmKS (gene locus tag: GRMZM2G114873), showing homology with the Arabidopsis thaliana basic helix-loop-helix (bHLH) DNA-binding transcription factor was identified. Using a comparative genomic approach, the ZmKS corresponding protein was identified as conceptual translated bHLH transcription factor ABA-responsive kinase substrate. ZmKS is localized in the nucleus, shows a potential binding specificity preferentially detectable on cis-acting E-box like heptameric motifs CCACTTG and CAAGTTG, and is phosphorylated by maize protein kinase ZmOST1. ZmKS is expressed in embryo, leaf and root, expression being affected by ABA and osmotic stress. Transgenic Arabidopsis plants, with gain of ZmKS function, show a delay in germination and a transcriptional stomatal opening-facilitator activity, switchover upon ZmKS phosphorylation, suggesting that ZmKS is an ABA-repressed trans-acting activator.

ABI1 and PP2CA phosphatases are negative regulators of Snf1-related protein kinase1 signaling in Arabidopsis.

Plant survival under environmental stress requires the integration of multiple signaling pathways into a coordinated response, but the molecular mechanisms underlying this integration are poorly understood. Stress-derived energy deprivation activates the Snf1-related protein kinases1 (SnRK1s), triggering a vast transcriptional and metabolic reprogramming that restores homeostasis and promotes tolerance to adverse conditions. Here, we show that two clade A type 2C protein phosphatases (PP2Cs), established repressors of the abscisic acid (ABA) hormonal pathway, interact with the SnRK1 catalytic subunit causing its dephosphorylation and inactivation. Accordingly, SnRK1 repression is abrogated in double and quadruple pp2c knockout mutants, provoking, similarly to SnRK1 overexpression, sugar hypersensitivity during early seedling development. Reporter gene assays and SnRK1 target gene expression analyses further demonstrate that PP2C inhibition by ABA results in SnRK1 activation, promoting SnRK1 signaling during stress and once the energy deficit subsides. Consistent with this, SnRK1 and ABA induce largely overlapping transcriptional responses. Hence, the PP2C hub allows the coordinated activation of ABA and energy signaling, strengthening the stress response through the cooperation of two key and complementary pathways.

The maize OST1 kinase homolog phosphorylates and regulates the maize SNAC1-type transcription factor.

The Arabidopsis kinase OPEN STOMATA 1 (OST1) plays a key role in regulating drought stress signalling, particularly stomatal closure. We have identified and investigated the functions of the OST1 ortholog in Z. mays (ZmOST1). Ectopic expression of ZmOST1 in the Arabidopsis ost1 mutant restores the stomatal closure phenotype in response to drought. Furthermore, we have identified the transcription factor, ZmSNAC1, which is directly phosphorylated by ZmOST1 with implications on its localization and protein stability. Interestingly, ZmSNAC1 binds to the ABA-box of ZmOST1, which is conserved in SnRK2s activated by ABA and is part of the contact site for the negative-regulating clade A PP2C phosphatases. Taken together, our results indicate that ZmSNAC1 is a substrate of ZmOST1 and delineate a novel osmotic stress transcriptional pathway in maize.