Diversification of heat shock transcription factors expanded thermal stress responses during early plant evolution.

The copy numbers of many plant transcription factor (TF) genes substantially increased during terrestrialization. This allowed TFs to acquire new specificities and thus create gene regulatory networks (GRNs) with new biological functions to help plants adapt to terrestrial environments. Through characterizing heat shock factor (HSF) genes MpHSFA1 and MpHSFB1 in the liverwort Marchantia polymorpha, we explored how heat-responsive GRNs widened their functions in M. polymorpha and Arabidopsis thaliana. An interspecies comparison of heat-induced transcriptomes and the evolutionary rates of HSFs demonstrated the emergence and subsequent rapid evolution of HSFB prior to terrestrialization. Transcriptome and metabolome analyses of M. polymorpha HSF-null mutants revealed that MpHSFA1 controls canonical heat responses such as thermotolerance and metabolic changes. MpHSFB1 also plays essential roles in heat responses, as well as regulating developmental processes including meristem branching and antheridiophore formation. Analysis of cis-regulatory elements revealed development- and stress-related TFs that function directly or indirectly downstream of HSFB. Male gametophytes of M. polymorpha showed higher levels of thermotolerance than female gametophytes, which could be explained by different expression levels of MpHSFA1U and MpHSFA1V on sex chromosome. We propose that the diversification of HSFs is linked to the expansion of HS responses, which enabled coordinated multicellular reactions in land plants.

Crosstalk between heterotrimeric G protein-coupled signaling pathways and WRKY transcription factors modulating plant responses to suboptimal micronutrient conditions.

Nutrient stresses induce foliar chlorosis and growth defects. Here we propose heterotrimeric G proteins as signaling mediators of various nutrient stresses, through meta-analyses of >20 transcriptomic data sets associated with nutrient stresses or G protein mutations. Systematic comparison of transcriptomic data yielded 104 genes regulated by G protein subunits under common nutrient stresses: 69 genes under Gß subunit (AGB1) control and 35 genes under Gα subunit (GPA1) control. Quantitative real-time PCR experiments validate that several transcription factors and metal transporters changed in expression level under suboptimal iron, zinc, and/or copper concentrations, while being misregulated in the Arabidopsis Gß-null (agb1) mutant. The agb1 mutant had altered metal ion profiles and exhibited severe growth arrest under zinc stress, and aberrant root waving under iron and zinc stresses, while the Gα-null mutation attenuated leaf chlorosis under iron deficiency in both Arabidopsis and rice. Our transcriptional network analysis inferred computationally that WRKY-family transcription factors mediate the AGB1-dependent nutrient responses. As corroborating evidence of our inference, ectopic expression of WRKY25 or WRKY33 rescued the zinc stress phenotypes and the expression of zinc transporters in the agb1-2 background. These results, together with Gene Ontology analyses, suggest two contrasting roles for G protein-coupled signaling pathways in micronutrient stress responses: one enhancing general stress tolerance and the other modulating ion homeostasis through WRKY transcriptional regulatory networks. In addition, tolerance to iron stress in the rice Gα mutant provides an inroad to improve nutrient stress tolerance of agricultural crops by manipulating G protein signaling.

Hyperspectral imaging of liverwort Marchantia polymorpha identifies MpWRKY10 as a key regulator defining Foliar pigmentation patterns.

Foliar pigmentation patterns vary among plant species and growth conditions. In this study, we utilize hyperspectral imaging to assess foliar pigmentation in the bryophyte Marchantia polymorpha under nutrient stress and identify associated genetic factors. Using singular value decomposition (SVD) for feature selection, we quantitate color variations induced by deficiencies in phosphate, nitrate, magnesium, calcium, and iron. Pseudo-colored thallus images show that disrupting MpWRKY10 causes irregular pigmentation with auronidin accumulation. Transcriptomic profiling shows that MpWRKY10 regulates phenylpropanoid pathway enzymes and R2R3-MYB transcription factors during phosphate deficiency, with MpMYB14 upregulation preceding pigment accumulation. MpWRKY10 is downregulated in older, pigmented thalli under phosphate deficiency but maintained in young thalli, where it suppresses pigmentation genes. This downregulation is absent in pigmented thalli due to aging. Comparative transcriptome analysis suggests similar WRKY and MYB roles in nutrient response and pigmentation in red-leaf lettuce, alluding to conserved genetic factors controlling foliar pigmentation patterns under nutrient deficiency.

G protein controls stress readiness by modulating transcriptional and metabolic homeostasis in Arabidopsis thaliana and Marchantia polymorpha.

The core G protein signaling module, which consists of Gα and extra-large Gα (XLG) subunits coupled with the Gßγ dimer, is a master regulator of various stress responses. In this study, we compared the basal and salt stress-induced transcriptomic, metabolomic and phenotypic profiles in Gα, Gß, and XLG-null mutants of two plant species, Arabidopsis thaliana and Marchantia polymorpha, and showed that G protein mediates the shift of transcriptional and metabolic homeostasis to stress readiness status. We demonstrated that such stress readiness serves as an intrinsic protection mechanism against further stressors through enhancing the phenylpropanoid pathway and abscisic acid responses. Furthermore, WRKY transcription factors were identified as key intermediates of G protein-mediated homeostatic shifts. Statistical and mathematical model comparisons between A. thaliana and M. polymorpha revealed evolutionary conservation of transcriptional and metabolic networks over land plant evolution, whereas divergence has occurred in the function of plant-specific atypical XLG subunit. Taken together, our results indicate that the shifts in transcriptional and metabolic homeostasis at least partially act as the mechanisms of G protein-coupled stress responses that are conserved between two distantly related plants.

Evolutionarily conserved hierarchical gene regulatory networks for plant salt stress response.

Plant cells constantly alter their gene expression profiles to respond to environmental fluctuations. These continuous adjustments are regulated by multi-hierarchical networks of transcription factors. To understand how such gene regulatory networks (GRNs) have stabilized evolutionarily while allowing for species-specific responses, we compare the GRNs underlying salt response in the early-diverging and late-diverging plants Marchantia polymorpha and Arabidopsis thaliana. Salt-responsive GRNs, constructed on the basis of the temporal transcriptional patterns in the two species, share common trans-regulators but exhibit an evolutionary divergence in cis-regulatory sequences and in the overall network sizes. In both species, WRKY-family transcription factors and their feedback loops serve as central nodes in salt-responsive GRNs. The divergent cis-regulatory sequences of WRKY-target genes are probably associated with the expansion in network size, linking salt stress to tissue-specific developmental and physiological responses. The WRKY modules and highly linked WRKY feedback loops have been preserved widely in other plants, including rice, while keeping their binding-motif sequences mutable. Together, the conserved trans-regulators and the quickly evolving cis-regulatory sequences allow salt-responsive GRNs to adapt over a long evolutionary timescale while maintaining some consistent regulatory structure. This strategy may benefit plants as they adapt to changing environments.