G protein signaling and metabolic pathways as evolutionarily conserved mechanisms to combat calcium deficiency.

Calcium (Ca) deficiency causes necrotic symptoms of foliar edges known as tipburn; however, the underlying cellular mechanisms have been poorly understood due to the lack of an ideal plant model and research platform. Using a phenotyping system that quantitates growth and tipburn traits in the model bryophyte Marchantia polymorpha, we evaluated metabolic compounds and the Gß-null mutant (gpb1) that modulate the occurrence and expansion of the tipburn. Transcriptomic comparisons between wild-type and gpb1 plants revealed the phenylalanine/phenylpropanoid biosynthesis pathway and reactive oxygen species (ROS) important for Ca deficiency responses. gpb1 plants reduced ROS production possibly through transcriptomic regulations of class III peroxidases and induced the expression of phenylpropanoid pathway enzymes without changing downstream lignin contents. Supplementation of intermediate metabolites and chemical inhibitors further confirmed the cell-protective mechanisms of the phenylpropanoid and ROS pathways. Marchantia polymorpha, Arabidopsis thaliana, and Lactuca sativa showed comparable transcriptomic changes where genes related to phenylpropanoid and ROS pathways were enriched in response to Ca deficiency. In conclusion, our study demonstrated unresolved signaling and metabolic pathways of Ca deficiency response. The phenotyping platform can speed up the discovery of chemical and genetic pathways, which could be widely conserved between M. polymorpha and angiosperms.

Oil Body Formation in Marchantia polymorpha Is Controlled by MpC1HDZ and Serves as a Defense against Arthropod Herbivores.

The origin of a terrestrial flora in the Ordovician required adaptation to novel biotic and abiotic stressors. Oil bodies, a synapomorphy of liverworts, accumulate secondary metabolites, but their function and development are poorly understood. Oil bodies of Marchantia polymorpha develop within specialized cells as one single large organelle. Here, we show that a class I homeodomain leucine-zipper (C1HDZ) transcription factor controls the differentiation of oil body cells in two different ecotypes of the liverwort M. polymorpha, a model genetic system for early divergent land plants. In flowering plants, these transcription factors primarily modulate responses to abiotic stress, including drought. However, loss-of-function alleles of the single ortholog gene, MpC1HDZ, in M. polymorpha did not exhibit phenotypes associated with abiotic stress. Rather, Mpc1hdz mutant plants were more susceptible to herbivory, and total plant extracts of the mutant exhibited reduced antibacterial activity. Transcriptomic analysis of the mutant revealed a reduction in expression of genes related to secondary metabolism that was accompanied by a specific depletion of oil body terpenoid compounds. Through time-lapse imaging, we observed that MpC1HDZ expression maxima precede oil body formation, indicating that MpC1HDZ mediates differentiation of oil body cells. Our results indicate that M. polymorpha oil bodies, and MpC1HDZ, are critical for defense against herbivory, but not for abiotic stress tolerance. Thus, C1HDZ genes were co-opted to regulate separate responses to biotic and abiotic stressors in two distinct land plant lineages.

Evolutionarily conserved regulatory mechanisms of abscisic acid signaling in land plants: characterization of ABSCISIC ACID INSENSITIVE1-like type 2C protein phosphatase in the liverwort Marchantia polymorpha.

Abscisic acid (ABA) is postulated to be a ubiquitous hormone that plays a central role in seed development and responses to environmental stresses of vascular plants. However, in liverworts (Marchantiophyta), which represent the oldest extant lineage of land plants, the role of ABA has been least emphasized; thus, very little information is available on the molecular mechanisms underlying ABA responses. In this study, we isolated and characterized MpABI1, an ortholog of ABSCISIC ACID INSENSITIVE1 (ABI1), from the liverwort Marchantia polymorpha. The MpABI1 cDNA encoded a 568-amino acid protein consisting of the carboxy-terminal protein phosphatase 2C (PP2C) domain and a novel amino-terminal regulatory domain. The MpABI1 transcript was detected in the gametophyte, and its expression level was increased by exogenous ABA treatment in the gemma, whose growth was strongly inhibited by ABA. Experiments using green fluorescent protein fusion constructs indicated that MpABI1 was mainly localized in the nucleus and that its nuclear localization was directed by the amino-terminal domain. Transient overexpression of MpABI1 in M. polymorpha and Physcomitrella patens cells resulted in suppression of ABA-induced expression of the wheat Em promoter fused to the beta -glucuronidase gene. Transgenic P. patens expressing MpABI1 and its mutant construct, MpABI1-d2, lacking the amino-terminal domain, had reduced freezing and osmotic stress tolerance, and associated with reduced accumulation of ABA-induced late embryogenesis abundant-like boiling-soluble proteins. Furthermore, ABA-induced morphological changes leading to brood cells were not prominent in these transgenic plants. These results suggest that MpABI1 is a negative regulator of ABA signaling, providing unequivocal molecular evidence of PP2C-mediated ABA response mechanisms functioning in liverworts.