A quantitative gibberellin signaling biosensor reveals a role for gibberellins in internode specification at the shoot apical meristem.

Growth at the shoot apical meristem (SAM) is essential for shoot architecture construction. The phytohormones gibberellins (GA) play a pivotal role in coordinating plant growth, but their role in the SAM remains mostly unknown. Here, we developed a ratiometric GA signaling biosensor by engineering one of the DELLA proteins, to suppress its master regulatory function in GA transcriptional responses while preserving its degradation upon GA sensing. We demonstrate that this degradation-based biosensor accurately reports on cellular changes in GA levels and perception during development. We used this biosensor to map GA signaling activity in the SAM. We show that high GA signaling is found primarily in cells located between organ primordia that are the precursors of internodes. By gain- and loss-of-function approaches, we further demonstrate that GAs regulate cell division plane orientation to establish the typical cellular organization of internodes, thus contributing to internode specification in the SAM.

Opposing polarity domains provide direction and play a role in cell division in plant growth.

Polar localization of proteins is important for plant growth and development. Identifying the interactors of polarized proteins provides spatial information and cell-type functions. In this issue of Developmental Cell, Wallner et al. (2024) utilize opposing polarity domain proteins to identify interactors and their functions during cell division in Arabidopsis stomata.

Expressing banana transcription factor MaERFVII3 in Arabidopsis confers enhanced waterlogging tolerance and root growth.

Background: Waterlogging poses a significant threat to plant growth and yield worldwide. Identifying the genes responsible for mitigating waterlogging stress is crucial. Ethylene-responsive factors (ERFs) are transcriptional regulators that respond to various biotic and abiotic stresses in plants. However, their roles and involvement in responding to waterlogging stress remain largely unexplored. Hence, this study aimed to elucidate the role of ERFs in enhancing banana plant resilience to waterlogging. Methods: We hypothesized that introducing a group VII ERF transcription factor in Arabidopsis could enhance waterlogging stress tolerance. To test this hypothesis, we isolated MaERFVII3 from banana roots, where it exhibited a significant induction in response to waterlogging stress. The isolated MaERFVII3 was introduced into Arabidopsis plants for functional gene studies. Results: Compared with wild-type plants, the MaERFVII3-expressing Arabidopsis showed increased survival and biomass under waterlogging stress. Furthermore, the abundance of transcripts related to waterlogging and hypoxia response showed an elevation in transgenic plants but a decrease in wild-type and empty vector plants when exposed to waterlogging stress. Our results demonstrate the significant contribution of MaERFVII3 to waterlogging tolerance in Arabidopsis, providing baseline data for further exploration and potentially contributing to crop improvement programs.

Transcriptome analysis and functional validation reveal the novel role of LhCYCL in axillary bud development in hybrid Liriodendron.

Shoot branching significantly influences yield and timber quality in woody plants, with hybrid Liriodendron being particularly valuable due to its rapid growth. However, understanding of the mechanisms governing shoot branching in hybrid Liriodendron remains limited. In this study, we systematically examined axillary bud development using morphological and anatomical approaches and selected four distinct developmental stages for an extensive transcriptome analysis. A total of 9,449 differentially expressed genes have been identified, many of which are involved in plant hormone signal transduction pathways. Additionally, we identified several transcription factors downregulated during early axillary bud development, including a noteworthy gene annotated as CYC-like from the TCP TF family, which emerged as a strong candidate for modulating axillary bud development. Quantitative real-time polymerase chain reaction results confirmed the highest expression levels of LhCYCL in hybrid Liriodendron axillary buds, while histochemical ß-glucuronidase staining suggested its potential role in Arabidopsis thaliana leaf axil development. Ectopic expression of LhCYCL in A. thaliana led to an increase of branches and a decrease of plant height, accompanied by altered expression of genes involved in the plant hormone signaling pathways. This indicates the involvement of LhCYCL in regulating shoot branching through plant hormone signaling pathways. In summary, our results emphasize the pivotal role played by LhCYCL in shoot branching, offering insights into the function of the CYC-like gene and establishing a robust foundation for further investigations into the molecular mechanisms governing axillary bud development in hybrid Liriodendron.

Crucial role of SWL1 in chloroplast biogenesis and development in Arabidopsis thaliana.

KEY MESSAGE: The disruption of the SWL1 gene leads to a significant down regulation of chloroplast and secondary metabolites gene expression in Arabidopsis thaliana. And finally results in a dysfunction of chloroplast and plant growth. Although the development of the chloroplast has been a consistent focus of research, the corresponding regulatory mechanisms remain unidentified. In this study, the CRISPR/Cas9 system was used to mutate the SWL1 gene, resulting in albino cotyledons and variegated true leaf phenotype. Confocal microscopy and western blot of chloroplast protein fractions revealed that SWL1 localized in the chloroplast stroma. Electron microscopy indicated chloroplasts in the cotyledons of swl1 lack well-defined grana and internal membrane structures, and similar structures have been detected in the albino region of variegated true leaves. Transcriptome analysis revealed that down regulation of chloroplast and nuclear gene expression related to chloroplast, including light harvesting complexes, porphyrin, chlorophyll metabolism and carbon metabolism in the swl1 compared to wild-type plant. In addition, proteomic analysis combined with western blot analysis, showed that a significant decrease in chloroplast proteins of swl1. Furthermore, the expression of genes associated with secondary metabolites and growth hormones was also reduced, which may be attributed to SWL1 associated with absorption and fixation of inorganic carbon during chloroplast development. Together, the above findings provide valuable information to elucidate the exact function of SWL1 in chloroplast biogenesis and development.

The GPAT4/6/8 clade functions in Arabidopsis root suberization nonredundantly with the GPAT5/7 clade required for suberin lamellae.

Lipid polymers such as cutin and suberin strengthen the diffusion barrier properties of the cell wall in specific cell types and are essential for water relations, mineral nutrition, and stress protection in plants. Land plant-specific glycerol-3-phosphate acyltransferases (GPATs) of different clades are central players in cutin and suberin monomer biosynthesis. Here, we show that the GPAT4/6/8 clade in Arabidopsis thaliana, which is known to mediate cutin formation, is also required for developmentally regulated root suberization, in addition to the established roles of GPAT5/7 in suberization. The GPAT5/7 clade is mainly required for abscisic acid-regulated suberization. In addition, the GPAT5/7 clade is crucial for the formation of the typical lamellated suberin ultrastructure observed by transmission electron microscopy, as distinct amorphous globular polyester structures were deposited in the apoplast of the gpat5 gpat7 double mutant, in contrast to the thinner but still lamellated suberin deposition in the gpat4 gpat6 gpat8 triple mutant. Site-directed mutagenesis revealed that the intrinsic phosphatase activity of GPAT4, GPAT6, and GPAT8, which leads to monoacylglycerol biosynthesis, contributes to suberin formation. GPAT5/7 lack an active phosphatase domain and the amorphous globular polyester structure observed in the gpat5 gpat7 double mutant was partially reverted by treatment with a phosphatase inhibitor or the expression of phosphatase-dead variants of GPAT4/6/8. Thus, GPATs that lack an active phosphatase domain synthetize lysophosphatidic acids that might play a role in the formation of the lamellated structure of suberin. GPATs with active and nonactive phosphatase domains appear to have nonredundant functions and must cooperate to achieve the efficient biosynthesis of correctly structured suberin.

PAT mRNA decapping factors are required for proper development in Arabidopsis.

Evolutionarily conserved protein associated with topoisomerase II (PAT1) proteins activate mRNA decay through binding mRNA and recruiting decapping factors to optimize posttranscriptional reprogramming. Here, we generated multiple mutants of pat1, pat1 homolog 1 (path1), and pat1 homolog 2 (path2) and discovered that pat triple mutants exhibit extremely stunted growth and all mutants with pat1 exhibit leaf serration while mutants with pat1 and path1 display short petioles. All three PATs can be found localized to processing bodies and all PATs can target ASYMMETRIC LEAVES 2-LIKE 9 transcripts for decay to finely regulate apical hook and lateral root development. In conclusion, PATs exhibit both specific and redundant functions during different plant growth stages and our observations underpin the selective regulation of the mRNA decay machinery for proper development.

Heterotic growth of hybrids of Arabidopsis thaliana is enhanced by elevated atmospheric CO2.

PREMISE: With the global atmospheric CO2 concentration on the rise, developing crops that can thrive in elevated CO2 has become paramount. We investigated the potential of hybridization as a strategy for creating crops with improved growth in predicted elevated atmospheric CO2. METHODS: We grew parent accessions and their F1 hybrids of Arabidopsis thaliana in ambient and elevated atmospheric CO2 and analyzed numerous growth traits to assess their productivity and underlying mechanisms. RESULTS: The heterotic increase in total dry mass, relative growth rate and leaf net assimilation rate was significantly greater in elevated CO2 than in ambient CO2. The CO2 response of net assimilation rate was positively correlated with the CO2 response of leaf nitrogen productivity and with that of leaf traits such as leaf size and thickness, suggesting that hybridization-induced changes in leaf traits greatly affected the improved performance in elevated CO2. CONCLUSIONS: Vegetative growth of hybrids seems to be enhanced in elevated CO2 due to improved photosynthetic nitrogen-use efficiency compared with parents. The results suggest that hybrid crops should be well-suited for future conditions, but hybrid weeds may also be more competitive.

Azelaic acid can efficiently compete for the auxin binding site TIR1, altering auxin polar transport, gravitropic response, and root growth and architecture in Arabidopsisthaliana roots.

The present study investigates the phytotoxic potential of azelaic acid (AZA) on Arabidopsis thaliana roots. Effects on root morphology, anatomy, auxin content and transport, gravitropic response and molecular docking were analysed. AZA inhibited root growth, stimulated lateral and adventitious roots, and altered the root apical meristem by reducing meristem cell number, length and width. The treatment also slowed down the roots' gravitropic response, likely due to a reduction in statoliths, starch-rich organelles involved in gravity perception. In addition, auxin content, transport and distribution, together with PIN proteins' expression and localisation were altered after AZA treatment, inducing a reduction in auxin transport and its distribution into the meristematic zone. Computational simulations showed that AZA has a high affinity for the auxin receptor TIR1, competing with auxin for the binding site. The AZA binding with TIR1 could interfere with the normal functioning of the TIR1/AFB complex, disrupting the ubiquitin E3 ligase complex and leading to alterations in the response of the plant, which could perceive AZA as an exogenous auxin. Our results suggest that AZA mode of action could involve the modulation of auxin-related processes in Arabidopsis roots. Understanding such mechanisms could lead to find environmentally friendly alternatives to synthetic herbicides.

The phosphorylation of carboxyl-terminal eIF2α by SPA kinases contributes to enhanced translation efficiency during photomorphogenesis.

Light triggers an enhancement of global translation during photomorphogenesis in Arabidopsis, but little is known about the underlying mechanisms. The phosphorylation of the α-subunit of eukaryotic initiation factor 2 (eIF2α) at a conserved serine residue in the N-terminus has been shown as an important mechanism for the regulation of protein synthesis in mammalian and yeast cells. However, whether the phosphorylation of this residue in plant eIF2α plays a role in regulation of translation remains elusive. Here, we show that the quadruple mutant of SUPPRESSOR OF PHYA-105 family members (SPA1-SPA4) display repressed translation efficiency after light illumination. Moreover, SPA1 directly phosphorylates the eIF2α C-terminus under light conditions. The C-term-phosphorylated eIF2α promotes translation efficiency and photomorphogenesis, whereas the C-term-unphosphorylated eIF2α results in a decreased translation efficiency. We also demonstrate that the phosphorylated eIF2α enhances ternary complex assembly by promoting its affinity to eIF2ß and eIF2γ. This study reveals a unique mechanism by which light promotes translation via SPA1-mediated phosphorylation of the C-terminus of eIF2α in plants.

A Genome-Wide Analysis of the CEP Gene Family in Cotton and a Functional Study of GhCEP46-D05 in Plant Development.

C-TERMINALLY ENCODED PEPTIDEs (CEPs) are a class of peptide hormones that have been shown in previous studies to play an important role in regulating the development and response to abiotic stress in model plants. However, their role in cotton is not well understood. In this study, we identified 54, 59, 34, and 35 CEP genes from Gossypium hirsutum (2n = 4x = 52, AD1), G. barbadense (AD2), G. arboreum (2n = 2X = 26, A2), and G. raimondii (2n = 2X = 26, D5), respectively. Sequence alignment and phylogenetic analyses indicate that cotton CEP proteins can be categorized into two subgroups based on the differentiation of their CEP domain. Chromosomal distribution and collinearity analyses show that most of the cotton CEP genes are situated in gene clusters, suggesting that segmental duplication may be a critical factor in CEP gene expansion. Expression pattern analyses showed that cotton CEP genes are widely expressed throughout the plant, with some genes exhibiting specific expression patterns. Ectopic expression of GhCEP46-D05 in Arabidopsis led to a significant reduction in both root length and seed size, resulting in a dwarf phenotype. Similarly, overexpression of GhCEP46-D05 in cotton resulted in reduced internode length and plant height. These findings provide a foundation for further investigation into the function of cotton CEP genes and their potential role in cotton breeding.

CEPs suppress auxin signaling but promote cytokinin signaling to inhibit root growth in Arabidopsis.

C-terminally encoded peptides (CEPs) are peptide hormones that function as mobile signals coordinating crucial developmental programs in plants. Previous studies have revealed that CEPs exert negative regulation on root development through interaction with CEP receptors (CEPRs), CEP DOWNSTREAMs (CEPDs), the cytokinin receptor ARABIDOPSIS HISTIDINE KINASE (AHKs) and the transcriptional repressor Auxin/Indole-3-Acetic Acid (AUX/IAA). However, the precise molecular mechanisms underlying CEPs-mediated regulation of root development via auxin and cytokinin signaling pathways still necessitate further detailed investigation. In this study, we examined prior research and elucidated the underlying molecular mechanisms. The results showed that both synthetic AtCEPs and overexpression of AtCEP5 markedly supressed primary root elongation and lateral root (LR) formation in Arabidopsis. Molecular biology and genetics elucidated how CEPs inhibit root growth by suppressing auxin signaling while promoting cytokinin signaling. In summary, this study elucidated the inhibitory effects of AtCEPs on Arabidopsis root growth and provided insights into their potential molecular mechanisms, thus enhancing our comprehension of CEP-mediated regulation of plant growth and development.

Role of sugars in the apical hook development of Arabidopsis etiolated seedlings.

KEY MESSAGE: The sugar supply in the medium affects the apical hook development of Arabidopsis etiolated seedlings. In addition, we provided the mechanism insights of this process. Dicotyledonous plants form an apical hook structure to shield their young cotyledons from mechanical damage as they emerge from the rough soil. Our findings indicate that sugar molecules, such as sucrose and glucose, are crucial for apical hook development. The presence of sucrose and glucose allows the apical hooks to be maintained for a longer period compared to those grown in sugar-free conditions, and this effect is dose-dependent. Key roles in apical hook development are played by several sugar metabolism pathways, including oxidative phosphorylation and glycolysis. RNA-seq data revealed an up-regulation of genes involved in starch and sucrose metabolism in plants grown in sugar-free conditions, while genes associated with phenylpropanoid metabolism were down-regulated. This study underscores the significant role of sugar metabolism in the apical hook development of etiolated Arabidopsis seedlings.

The significance of the two cytosolic glutamine synthetase enzymes, GLN1;3 and GLN1;5, in the context of seed development and germination in Arabidopsis thaliana.

Glutamine synthetase (GS), an initial enzyme in nitrogen (N) plant metabolism, exists as a group of isoenzymes found in both cytosolic (GS1) and plastids (GS2) and has gathered significant attention for enhancing N use efficiency and crop yield. This work focuses on the A. thaliana GLN1;3 and GLN1;5 genes, the two predicted most expressed genes in seeds, among the five isogenes encoding GS1 in this species. The expression patterns were studied using transgenic marker line plants and qPCR during seed development and germination. The observed patterns highlight distinct functions for the two genes and confirm GLN1;5 as the most highly expressed GS1 gene in seeds. The GLN1;5, expression, oriented towards hypocotyl and cotyledons, suggests a role in protein turnover during germination, while the radicle-oriented expression of GLN1;3 supports a function in early external N uptake. While the single mutants exhibited a normal phenotype, except for a decrease in seed parameters, the double gln1;3/gln1;5 mutant displayed a germination delay, substantial impairment in growth, nitrogen metabolism, and number and quality of the seeds, as well as a diminishing in flowering. Although seed and pollen-specific, GLN1;5 expression is upregulated in the meristems of the gln1;3 mutants, filling the lack of GLN1;3 and ensuring the normal functioning of the gln1;3 mutants. These findings validate earlier in silico data on the expression patterns of GLN1;3 and GL1;5 genes in seeds, explore their different functions, and underscore their essential role in plant growth, seed production, germination, and early stages of plant development.

An AGO10:miR165/6 module regulates meristem activity and xylem development in the Arabidopsis root.

The RNA-silencing effector ARGONAUTE10 influences cell fate in plant shoot and floral meristems. ARGONAUTE10 also accumulates in the root apical meristem (RAM), yet its function(s) therein remain elusive. Here, we show that ARGONAUTE10 is expressed in the root cell initials where it controls overall RAM activity and length. ARGONAUTE10 is also expressed in the stele, where post-transcriptional regulation confines it to the root tip's pro-vascular region. There, variations in ARGONAUTE10 levels modulate metaxylem-vs-protoxylem specification. Both ARGONAUTE10 functions entail its selective, high-affinity binding to mobile miR165/166 transcribed in the neighboring endodermis. ARGONAUTE10-bound miR165/166 is degraded, likely via SMALL-RNA-DEGRADING-NUCLEASES1/2, thus reducing miR165/166 ability to silence, via ARGONAUTE1, the transcripts of cell fate-influencing transcription factors. These include PHABULOSA (PHB), which controls meristem activity in the initials and xylem differentiation in the pro-vasculature. During early germination, PHB transcription increases while dynamic, spatially-restricted transcriptional and post-transcriptional mechanisms reduce and confine ARGONAUTE10 accumulation to the provascular cells surrounding the newly-forming xylem axis. Adequate miR165/166 concentrations are thereby channeled along the ARGONAUTE10-deficient yet ARGONAUTE1-proficient axis. Consequently, inversely-correlated miR165/166 and PHB gradients form preferentially along the axis despite ubiquitous PHB transcription and widespread miR165/166 delivery inside the whole vascular cylinder.

ARGONAUTE10 controls cell fate specification and formative cell divisions in the Arabidopsis root.

A key question in plant biology is how oriented cell divisions are integrated with patterning mechanisms to generate organs with adequate cell type allocation. In the root vasculature, a gradient of miRNA165/6 controls the abundance of HD-ZIP III transcription factors, which in turn control cell fate and spatially restrict vascular cell proliferation to specific cells. Here, we show that vascular development requires the presence of ARGONAUTE10, which is thought to sequester miRNA165/6 and protect HD-ZIP III transcripts from degradation. Our results suggest that the miR165/6-AGO10-HDZIP III module acts by buffering cytokinin responses and restricting xylem differentiation. Mutants of AGO10 show faster growth rates and strongly enhanced survival under severe drought conditions. However, this superior performance is offset by markedly increased variation and phenotypic plasticity in sub-optimal carbon supply conditions. Thus, AGO10 is required for the control of formative cell division and coordination of robust cell fate specification of the vasculature, while altering its expression provides a means to adjust phenotypic plasticity.

Golgi-localized APYRASE 1 is critical for Arabidopsis growth by affecting cell wall integrity under boron deficiency.

Many nucleoside triphosphate-diphosphohydrolases (NTPDases/APYRASEs, APYs) play a key role in modulating extracellular nucleotide levels. However, the Golgi-localized APYs, which help control glycosylation, have rarely been studied. Here, we identified AtAPY1, a gene encoding an NTPDase in the Golgi apparatus, which is required for cell wall integrity and plant growth under boron (B) limited availability. Loss of function in AtAPY1 hindered cell elongation and division in root tips while increasing the number of cortical cell layers, leading to swelling of the root tip and abundant root hairs under low B stress. Further, expression pattern analysis revealed that B deficiency significantly induced AtAPY1, especially in the root meristem and stele. Fluorescent-labeled AtAPY1-GFP localized to the Golgi stack. Biochemical analysis showed that AtAPY1 exhibited a preference of UDP and GDP hydrolysis activities. Consequently, the loss of function in AtAPY1 might disturb the homoeostasis of NMP-driven NDP-sugar transport, which was closely related to the synthesis of cell wall polysaccharides. Further, cell wall-composition analysis showed that pectin content increased and borate-dimerized RG-II decreased in apy1 mutants, along with a decrease in cellulose content. Eventually, altered polysaccharide characteristics presumably cause growth defects in apy1 mutants under B deficiency. Altogether, these data strongly support a novel role for AtAPY1 in mediating responses to low B availability by regulating cell wall integrity.

Complex control of seed germination timing by ERF50 involves RGL2 antagonism and negative feedback regulation of DOG1.

Seed dormancy governs germination timing, with both evolutionary and applied consequences. Despite extensive studies on the hormonal and genetic control of these processes, molecular mechanisms directly linking dormancy and germination remain poorly understood. By screening a collection of lines overexpressing Arabidopsis transcription factors, we identified ERF50 as a key gene to control dormancy and germination. To study its regulation, we measured seed-related physiological parameters in loss-of-function mutants and carried out transactivation, protein interaction and ChIP-PCR analyses. We found direct ERF50-mediated repression of DOG1 and activation of EXPA2 transcription, which results in enhanced seed germination. Although ERF50 expression is increased by DOG1 in dormant seeds, ERF50 germination-promoting activity is blocked by RGL2. The physiological, genetic and molecular evidence gathered here supports that ERF50 controls germination timing by regulating DOG1 levels to leverage its role as enhancer of seed germination, via RGL2 antagonism on EXPA2 expression. Our results highlight the central role of ERF50 as a feedback regulator to couple and fine-tune seed dormancy and germination.

The mechanism behind tenuazonic acid-mediated inhibition of plant plasma membrane H+-ATPase and plant growth.

The increasing prevalence of herbicide-resistant weeds has led to a search for new herbicides that target plant growth processes differing from those targeted by current herbicides. In recent years, some studies have explored the use of natural compounds from microorganisms as potential new herbicides. We previously demonstrated that tenuazonic acid (TeA) from the phytopathogenic fungus Stemphylium loti inhibits the plant plasma membrane (PM) H+-ATPase, representing a new target for herbicides. In this study, we further investigated the mechanism by which TeA inhibits PM H+-ATPase and the effect of the toxin on plant growth using Arabidopsis thaliana. We also studied the biochemical effects of TeA on the PM H+-ATPases from spinach (Spinacia oleracea) and A. thaliana (AHA2) by examining PM H+-ATPase activity under different conditions and in different mutants. Treatment with 200 µM TeA-induced cell necrosis in larger plants and treatment with 10 µM TeA almost completely inhibited cell elongation and root growth in seedlings. We show that the isoleucine backbone of TeA is essential for inhibiting the ATPase activity of the PM H+-ATPase. Additionally, this inhibition depends on the C-terminal domain of AHA2, and TeA binding to PM H+-ATPase requires the Regulatory Region I of the C-terminal domain in AHA2. TeA likely has a higher binding affinity toward PM H+-ATPase than the phytotoxin fusicoccin. Finally, our findings show that TeA retains the H+-ATPase in an inhibited state, suggesting that it could act as a lead compound for creating new herbicides targeting the PM H+-ATPase.