Constitutive activation of ABA receptors in Arabidopsis reveals unique regulatory circuitries.

Abscisic acid (ABA) is best known for regulating the responses to abiotic stressors. Thus, applications of ABA signaling pathways are considered promising targets for securing yield under stress. ABA levels rise in response to abiotic stress, mounting physiological and metabolic responses that promote plant survival under unfavorable conditions. ABA elicits its effects by binding to a family of soluble receptors found in monomeric and dimeric states, differing in their affinity to ABA and co-receptors. However, the in vivo significance of the biochemical differences between these receptors remains unclear. We took a gain-of-function approach to study receptor-specific functionality. First, we introduced activating mutations that enforce active ABA-bound receptor conformation. We then transformed Arabidopsis ABA-deficient mutants with the constitutive receptors and monitored suppression of the ABA deficiency phenotype. Our findings suggest that PYL4 and PYL5, monomeric ABA receptors, have differential activity in regulating transpiration and transcription of ABA biosynthesis and stress response genes. Through genetic and metabolic data, we demonstrate that PYR1, but not PYL5, is sufficient to activate the ABA positive feedback mechanism. We propose that ABA signaling - from perception to response - flows differently when triggered by different PYLs, due to tissue and transcription barriers, thus resulting in distinct circuitries.

Evolution of Abscisic Acid Signaling Module and Its Perception.

We hereby review the perception and responses to the stress hormone Abscisic acid (ABA), along the trajectory of 500M years of plant evolution, whose understanding may resolve how plants acquired this signaling pathway essential for the colonization of land. ABA levels rise in response to abiotic stresses, coordinating physiological and metabolic responses, helping plants survive stressful environments. In land plants, ABA signaling cascade leads to growth arrest and large-scale changes in transcript levels, required for coping with environmental stressors. This response is regulated by a PYRABACTIN RESISTANCE 1-like (PYL)-PROTEIN PHOSPHATASE 2C (PP2C)-SNF1-RELATED PROTEIN KINASE 2 (SnRK2) module, that initiates phosphor-activation of transcription factors and ion channels. The enzymatic portions of this module (phosphatase and kinase) are functionally conserved from streptophyte algae to angiosperms, whereas the regulatory component -the PYL receptors, putatively evolved in the common ancestor of Zygnematophyceae and embryophyte as a constitutive, ABA-independent protein, further evolving into a ligand-activated receptor at the embryophyta. This evolutionary process peaked with the appearance of the strictly ABA-dependent subfamily III stress-triggered angiosperms' dimeric PYL receptors. The emerging picture is that the ancestor of land plants and its predecessors synthesized ABA, as its biosynthetic pathway is conserved between ancestral and current day algae. Despite this ability, it was only the common ancestor of land plants which acquired the hormonal-modulation of PYL activity by ABA. This raises several questions regarding both ABA's function in ABA-non-responsive organisms, and the evolutionary aspects of the ABA signal transduction pathway.

A ligand-independent origin of abscisic acid perception.

Land plants are considered monophyletic, descending from a single successful colonization of land by an aquatic algal ancestor. The ability to survive dehydration to the point of desiccation is a key adaptive trait enabling terrestrialization. In extant land plants, desiccation tolerance depends on the action of the hormone abscisic acid (ABA) that acts through a receptor-signal transduction pathway comprising a PYRABACTIN RESISTANCE 1-like (PYL)-PROTEIN PHOSPHATASE 2C (PP2C)-SNF1-RELATED PROTEIN KINASE 2 (SnRK2) module. Early-diverging aeroterrestrial algae mount a dehydration response that is similar to that of land plants, but that does not depend on ABA: Although ABA synthesis is widespread among algal species, ABA-dependent responses are not detected, and algae lack an ABA-binding PYL homolog. This raises the key question of how ABA signaling arose in the earliest land plants. Here, we systematically characterized ABA receptor-like proteins from major land plant lineages, including a protein found in the algal sister lineage of land plants. We found that the algal PYL-homolog encoded by Zygnema circumcarinatum has basal, ligand-independent activity of PP2C repression, suggesting this to be an ancestral function. Similarly, a liverwort receptor possesses basal activity, but it is further activated by ABA. We propose that co-option of ABA to control a preexisting PP2C-SnRK2-dependent desiccation-tolerance pathway enabled transition from an all-or-nothing survival strategy to a hormone-modulated, competitive strategy by enabling continued growth of anatomically diversifying vascular plants in dehydrative conditions, enabling them to exploit their new environment more efficiently.

FIE, a nuclear PRC2 protein, forms cytoplasmic complexes in Arabidopsis thaliana.

Polycomb group (PcG) proteins are evolutionarily conserved chromatin modifiers that regulate developmental pathways in plants. PcGs form nuclear multi-subunit Polycomb Repressive Complexes (PRCs). The PRC2 complex mediates gene repression via methylation of lysine 27 on histone H3, which consequently leads to chromatin condensation. In Arabidopsis thaliana, several PRC2 complexes with different compositions were identified, each controlling a particular developmental program.The core subunit FIE is crucial for PRC2 function throughout the plant life cycle, yet accurate information on its spatial and temporal localization was absent. This study focused on identifying FIE accumulation patterns, using microscopy and biochemical approaches. Analysing endogenous FIE and transgenic gFIE-green fluorescent protein fusion protein (gFIE-GFP) showed that FIE accumulates in the nuclei of every cell type examined. Interestingly, gFIE-GFP, as well as the endogenous FIE, also localized to the cytoplasm in all examined tissues. In both vegetative and reproductive organs, FIE formed cytoplasmic high-molecular-mass complexes, in parallel to the nuclear PRC2 complexes. Moreover, size-exclusion chromatography and bimolecular fluorescence complementation assays indicated that in inflorescences FIE formed a cytoplasmic complex with MEA, a PRC2 histone methyltransferase subunit. In contrast, CLF and SWN histone methyltransferases were strictly nuclear. Presence of PRC2 subunits in cytoplasmic complexes has not been previously described in plants. Our findings are in agreement with accumulating evidence demonstrating cytoplasmic localization and function of PcGs in metazoa. The cytosolic accumulation of PRC2 components in plants supports the model that PcGs have alternative non-nuclear functions that go beyond chromatin methylation.

Ultralow calcium requirements of fungi facilitate use of calcium regulating agents to suppress host calcium-dependent defenses, synergizing infection by a mycoherbicide.

Infection by many fungi activates a variety of calcium dependent defenses in the hosts, slowing or suppressing the attacker and limiting the efficacy of mycoherbicides. The calcium requirement for fungal growth is so low that it could only be implied based on fungi containing calcium-dependent signaling enzymes. Analytical grade media contain <2 microM calcium, and the addition of specific chelators does not affect fungal growth. Hydrophobic derivatives of the calcium-specific chelator BAPTA designed to traverse plant cuticles were synthesized in order to chelate calcium internally during fungal attack. Some chelators as well as calcium precipitating oxalate and channel blocker verapamil were applied with a weakly mycoherbicidal Colletotrichum coccodes to cotyledons of compatible Abutilon threophrasti. They suppressed calcium dependent callose biosynthesis in the weed and increased virulence but may have affected other calcium-dependent processes that facilitate virulence. The low calcium requirement of fungi, and their high affinity for calcium, allows the application of calcium-regulating agents as synergists for mycoherbicides where the weed uses calcium-dependent defenses.