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Bennett, M. J.

Publications and source records attributed to Bennett, M. J..

14 recordsLinked to original sources

Mathematical modeling of SUMOylation reveals tissue-specific and dynamic responses to abiotic stress

SUMOylation is an essential and evolutionarily conserved post-translational modification that enables rapid cellular adaptation to environmental stress. Despite its importance, how the SUMOylation machinery controls its dynamics remains poorly understood. Here, we integrated a mathematical model of the plant SUMO cycle with quantitative imaging and biochemical data to decode the regulatory logic of SUMOylation dynamics. The model reveals a fundamental principle: steady-state SUMOylation levels are determined not by the full complement of SUMO-cycle components, but by the abundance of SUMO modifier proteins and E1 enzymes, together with SUMO protease abundance and activity. Strikingly, when we integrated fluorescence measurements from reporters for SUMO-cycle components in Arabidopsis roots after three hours of abiotic stress, the model predicted decreased SUMOylation levels across several tissues, contrary to prevailing evidence. Motivated by this unexpected prediction, we performed time-resolved imaging to better characterize the dynamics of key SUMO-cycle components after salt and osmotic stress. These data revealed highly dynamic component levels within the first three hours of stress that were not captured in the previous measurements. Incorporating this temporal data into the model uncovered a twin-peaked SUMOylation response to salt stress: an early surge underpinned by protease dynamics, followed by a decline and a second peak driven by elevated SUMO modifier and E2 levels. Western blot analysis revealed a corresponding twin-peaked pattern of SUMOylation. Together, our findings demonstrate that the plant SUMO cycle is a highly dynamic system that achieves tissue- and stress-specific control of SUMOylation through the coordinated interplay of multiple SUMO-cycle components.

plant biology↗

Water stress adaptive responses in plants require movement of ABA and AB-aldehyde from vascular to target tissues

Vascular plants rapidly coordinate root and shoot responses to water stress. Abscisic acid (ABA) mediates these adaptations; however, it remains unclear which cells produce ABA, whether ABA synthesis shifts during stress, and whether ABA movement is required for its adaptive functions. Here, we map ABA biosynthesis at cellular resolution in Arabidopsis and report that water-stress adaptive responses in roots and shoots require movement of ABA and its precursor AB-aldehyde from vascular tissues to target cells. We suggest that ABA accumulation arises from two parallel routes: (i) ABA synthesized in the vasculature via ABA2 and AAO3, then moving to guard cells, and (ii) phloem-derived AB-aldehyde being converted to ABA in the epidermis or bundle sheath by AAO1 and AAO2. Finally, we predict that tightly packed cells beneath leaf veins facilitate efficient ABA delivery to guard cells, an anatomical arrangement that has enabled angiosperms to evolve the use of ABA to rapidly close stomata.

plant biology↗

Comprehensive characterisation of IAA inactivation pathways reveals the impact of glycosylation on auxin metabolism and plant development

Together with biosynthesis and transport, inactivation regulates the concentration of indole-3-acetic acid (IAA), a key auxinic compound with a myriad of functions in plant development. Main inactive IAA metabolites are categorised into oxidised forms and ester- or amide-linked conjugates. DIOXYGENASE FOR AUXIN OXIDATION1 (DAO1) and DAO2, 2-oxoglutarate and iron-dependent dioxygenases, contribute to IAA oxidative inactivation in collaboration with group II GRETCHEN HAGEN3 (GH3) IAA-amido synthetases, while a group of UDP-glycosyltransferases (UGTs) conjugate IAA to sugars. To study the IAA inactivation routes, we generated combinatorial mutants between all group II GH3s (gh3oct) and DAO1 or DAO2, as well as between the DAOs and main UGTs. In vivo [13C6]IAA feeding experiments traced the metabolic fate of the exogenously applied IAA, supporting the main IAA inactivation pathway, in which DAO acts downstream of GH3s. They also indicated that UGT-mediated IAA glycosylation is more important than previously assumed for modulating IAA levels and plant development. Our metabolic and transcriptomic data further revealed that gh3oct may still produce some GH3 activity, explaining previous reported phenotypic inconsistencies. Our data additionally suggest that other not yet identified metabolic activities might play a role in IAA overproducing plants, and that the premature downregulation of flowering time integrators like FLOWERING LOCUS C (FLC) likely underlies the early flowering of gh3oct and gh3oct dao1 plants.

plant biology↗

Elucidating tissue and sub-cellular specificity of the entire SUMO network reveals how stress responses are fine-tuned in a eukaryote

SUMOylation is essential in plant and animal cells, but it remains unknown how SUMO components act in concert to modify specific targets in response to environmental stresses. In this study we characterise every SUMO component in the Arabidopsis root to create the first complete SUMO Cell Atlas in eukaryotes. This unique resource reveals wide spatial variation, where SUMO proteins and proteases have sub-functionalized in both their expression and subcellular localisation. During stress SUMO conjugation is mainly driven by tissue specific regulation of the SUMO E2 ligase. Stress-specific modulation of the SUMO pathway reveals unique combinations of Proteases being targeted for regulation in distinct root tissues by salt, osmotic and biotic signals. Our SUMO Cell Atlas resources reveal how this PTM influences cellular and tissue scale adaptations during root development and stress responses. We provide the first comprehensive study elucidating how multiple stress inputs can regulate an entire PTM system.

plant biology↗

Root metaxylem area influences drought tolerance and transpiration in pearl millet in a soil texture dependent manner

O_LIPearl millet is a key cereal for food security in drylands but its yield is strongly impacted by drought. We investigated how root anatomical traits contribute to mitigating the effects of vegetative drought stress in pearl millet. C_LIO_LIWe examined associations between root anatomical traits and agronomical performance in a pearl millet diversity panel under irrigated and vegetative drought stress treatments in field trials. The impact of associated anatomical traits on transpiration was assessed using subpanels grown in different soil within a greenhouse. C_LIO_LIIn the field, total metaxylem area was positively correlated with grain weight and its maintenance under drought. In the greenhouse, genotypes with larger metaxylem area grown in sandy soil exhibited a consumerist water use strategy under irrigation, which shifted to a conservative strategy under drought. Water savings was mediated by transpiration restriction under high evaporative demand. This mechanism was dependent on soil hydraulics as it was not observed in peat soil with higher hydraulic conductivity upon soil drying. C_LIO_LIWe propose that water savings under drought, mediated by large metaxylem area and its interaction with soil hydraulics, help mitigate vegetative drought stress. Our findings highlight the role of soil hydraulic properties in shaping plant hydraulics and drought tolerance. C_LI

physiology↗

Maize genetic diversity identifies moisture-dependent root-branch signaling pathways

Plants grow complex root systems to extract unevenly distributed resources from soils. Spatial differences in soil moisture are perceived by root tips leading to the patterning of new root branches towards available water, a process called hydropatterning. Little is known about hydropatterning behavior and its genetic basis in crops plants. Here, we develop an assay to measure hydropatterning in maize and reveal substantial differences between tropical/subtropical and temperate maize breeding germplasm that likely resulted from divergent selection. Genetic dissection of hydropatterning confirmed the regulatory role of auxin and revealed that the gaseous hormone ethylene acts to locally inhibit root branching from air-exposed tissues. These findings demonstrate the crop relevance of hydropatterning and establish its genetic basis.

plant biology↗

Harnessing Landrace Diversity Empowers Wheat Breeding for Climate Resilience

Breeding crops resilient to climate change is urgently needed to help ensure food security. A key challenge is to harness genetic diversity to optimise adaptation, yield, stress resilience and nutrition. We examined the genetic and phenotypic diversity of the A.E. Watkins landrace collection of bread wheat (Triticum aestivum), a major global cereal, through whole-genome re-sequencing (827 Watkins landraces and 208 modern cultivars) and in-depth field evaluation spanning a decade. We discovered that modern cultivars are derived from just two of the seven ancestral groups of wheat, leaving five groups as previously untapped sources for breeding. This provides access to landrace-specific functional variations using structured germplasm, genotyping and informatics resources. Employing complementary genetic populations and approaches, we identified thousands of high-resolution quantitative trait loci (QTL) and significant marker-trait associations for major traits, revealing many Watkins-unique loci that can confer superior traits in modern wheat. Furthermore, we identified and functionally verified causative genes for climate-change adaptation, nutritional enhancement and resistance to wheat blast. Finally, we assessed the phenotypic effects of 44,338 Watkins-unique haplotypes, introgressed from 143 prioritised QTL in the context of modern cultivars, bridging the gap between landrace diversity and current breeding. This study establishes a framework for systematically utilising genetic diversity in crop improvement to achieve sustainable food security.

plant biology↗

Glutaredoxin regulation of primary root growth confers early drought stress tolerance in pearl millet

Seedling root traits impact plant establishment under challenging environments. Pearl millet is one of the most heat and drought tolerant cereal crops that provides a vital food source across the sub-Saharan Sahel region. Pearl millets early root system features a single fast-growing primary root which we hypothesize is an adaptation to the Sahelian climate. Using crop modelling, we demonstrate that early drought stress is an important constraint in agrosystems in the Sahel where pearl millet was domesticated. Furthermore, we show that increased pearl millet primary root growth is correlated with increased early water stress tolerance in field conditions. Genetics including GWAS and QTL approaches identify genomic regions controlling this key root trait. Combining gene expression data, re-sequencing and re-annotation of one of these genomic regions identified a glutaredoxin-encoding gene PgGRXC9 as the candidate stress resilience root growth regulator. Functional characterization of its closest Arabidopsis homolog AtROXY19 revealed a novel role for this glutaredoxin (GRX) gene clade in regulating cell elongation. In summary, our study suggests a conserved function for GRX genes in conferring root cell elongation and enhancing resilience of pearl millet to its Sahelian environment.

plant biology↗

Genotypic variation in soil penetration by maize roots is negatively related to ethylene-induced thickening

Radial expansion is a classic response of roots to mechanical impedance that has generally been assumed to aid penetration. We analysed the response of maize nodal roots to impedance to test the hypothesis that radial expansion is not related to the ability of roots to cross a compacted soil layer. Genotypes varied in their ability to cross the compacted layer, and those with a steeper approach to the compacted layer or less radial expansion in the compacted layer were more likely to cross the layer and achieve greater depth. Root radial expansion was due to cortical cell size expansion, while cortical cell file number remained constant. Genotypes and nodal root classes that exhibited radial expansion upon encountering the compacted soil layer also thickened in response to exogenous ethylene in hydroponic culture, i.e. radial expansion in response to ethylene was correlated with the thickening response to impedance in soil. We propose that ethylene insensitive roots, i.e. those that do not thicken and are able to overcome impedance, have a competitive advantage under mechanically impeded conditions as they can maintain their elongation rates. We suggest that prolonged exposure to ethylene could function as a stop signal for axial root growth.

plant biology↗

Topically delivered 22 nt siRNAs enhance RNAi silencing of endogenous genes in two species

22 nt miRNAs or siRNAs have been shown to specifically induce production of transitive (secondary) siRNAs for targeted mRNAs. An abrasion method to deliver dsRNAs into leaf cells of intact plants was used to investigate the activities of 21 and 22 nt siRNAs in silencing genes in Nicotiana benthamiana and Amaranthus cruentus. We confirmed that both 21 and 22 nt siRNAs were able to silence a green fluorescent protein (GFP) transgene in treated leaves of N. benthamiana, but systemic silencing of GFP occurred only when the guide strand contained 22 nt. Silencing in the treated leaves of N. benthamiana was demonstrated for 3 endogenous genes: magnesium cheletase subunit I (CHL-I), magnesium cheletase subunit H (CHL-H), and GUN4. However, systemic silencing of these endogenous genes was not observed. Very high levels of transitive siRNAs were produced for GFP in response to treatment with 22 nt siRNAs, but only low levels were produced in response to a 21 nt siRNA. The endogenous genes tested also had more transitive siRNAs produced in response to 22 nt siRNAs, but the response varied from weak (CHL-I) to strong (CHL-H). 22 nt siRNAs produced greater local silencing phenotypes than 21 nt siRNAs for GFP, CHL-H and GUN4 in N. benthamiana. The special activity of 22 nt siRNAs in producing a greater local phenotype and induction of elevated levels of transitive siRNAs was also shown in A. cruentus for the CHL-H gene. These experiments suggest a functional role for transitive siRNAs in amplifying the RNAi response.

plant biology↗

External mechanical cues reveal core molecular pathway behind tissue bending in plants

Tissue folding is a central building block of plant and animal morphogenesis. In dicotyledonous plants, hypocotyl folds to form hook after seedling germination that protects their aerial stem cell niche during emergence from soil. Auxin response factors and auxin transport are classically thought to play a key role in this process. Here we show that the microtubule-severing enzyme katanin contributes to hook formation. However, by exposing hypocotyls to external mechanical cues mimicking the natural soil environment, we reveal that auxin response factors ARF7/ARF19, auxin influx carriers, and katanin are dispensable for apical hook formation, indicating that these factors primarily play the role of catalyzers of tissue bending in the absence of external mechanical cues. Instead, our results reveal the key roles of the non-canonical TMK-mediated auxin pathway, PIN efflux carriers and cellulose microfibrils as components of the core pathway behind hook formation in presence or absence of external mechanical cues.

plant biology↗

Identification of nitrogen-dependent QTL and underlying genes for root system architecture in hexaploid wheat

The root system architecture (RSA) of a crop has a profound effect on the uptake of nutrients and consequently the potential yield. However, little is known about the genetic basis of RSA and resource adaptive responses in wheat (Triticum aestivum L.). Here, a high-throughput germination paper plant phenotyping system was used to identify seedling traits in a wheat doubled haploid mapping population, Savannah x Rialto. Significant genotypic and nitrate-N treatment variation was found across the population for seedling traits with distinct trait grouping for root size-related traits and root distribution-related traits. Quantitative trait locus (QTL) analysis identified a total of 59 seedling trait QTLs. Across two nitrate treatments, 27 root QTLs were specific to the nitrate treatment. Transcriptomic analyses for one of the QTLs on chromosome 2D found under low nitrate conditions was pursued revealing gene enrichment in N-related biological processes and 17 candidate up-regulated genes with possible involvement in a root angle response. Together, these findings provide genetic insight into root system architecture and plant adaptive responses to nitrate and provide targets that could help improve N capture in wheat.

plant biology↗

Wheat shovelomics I: A field phenotyping approach for characterising the structure and function of root systems in tillering species

Wheat represents a major crop, yet the current rate of yield improvement is insufficient to meet its projected global food demand. Breeding root systems more efficient for water and nitrogen capture represents a promising avenue for accelerating yield gains. Root crown phenotyping, or shovelomics, relies on excavation of the upper portions of root systems in the field and measuring root properties such as numbers, angles, densities and lengths. We report a new shovelomics method that images the whole wheat root crown, then partitions it into the main shoot and tillers for more intensive phenotyping. Root crowns were phenotyped using the new method from the Rialto x Savannah population consisting of both parents and 94 doubled-haploid lines. For the whole root crown, the main shoot, and tillers, root phenes including nodal root number, growth angle, length, and diameter were measured. Substantial variation and heritability were observed for all phenes. Principal component analysis revealed latent constructs that imply pleiotropic genetic control of several related root phenes. Correlational analysis revealed that nodal root number and growth angle correlate among the whole crown, main shoot, and tillers, indicating shared genetic control among those organs. We conclude that this phenomics approach will be useful for breeding ideotype root systems in tillering species.

plant biology↗

The expa1-1 mutant reveals a new biophysical lateral root organogenesis checkpoint

In plants, post-embryonic formation of new organs helps shape the adult organism. This requires the tight regulation of when and where a new organ is formed, and a coordination of the underlying cell divisions. To build a root system, new lateral roots are continuously developing, and this process requires asymmetric cell division in adjacent pericycle cells. Characterization of an expansin a1 (expa1) mutant has revealed a novel checkpoint during lateral root formation. Specifically, a minimal pericycle width was found to be necessary and sufficient to trigger asymmetric pericycle cell divisions during auxin-driven lateral root formation. We conclude that a localized radial expansion of adjacent pericycle cells is required to position the asymmetric cell divisions and generate a core of small daughter cells, which is a prerequisite for lateral root organogenesis.\n\nSIGNFICANCE STATEMENTOrgan formation is an essential process in plants and animals, driven by cell division and cell identity establishment. Root branching, where lateral roots form along the primary root axis, increases the root system and aids capture of water and nutrients. We have discovered that tight control of cell width is necessary to co-ordinate asymmetric cell divisions in cells that give rise to a new lateral root organ. While biomechanical processes have been shown to play a role in plant organogenesis, including lateral root formation, our data give new mechanistic insights into the cell size checkpoint during lateral root initiation.

plant biology↗