Biology subjects

Langdale, J. A.

Publications and source records attributed to Langdale, J. A..

9 recordsLinked to original sources

bioRxiv2024

Spatial transcriptomics reveals distinct lineage identities for major and minor vein initiation during maize leaf development

Leaves of flowering plants are characterised by diverse venation patterns. Patterning begins with the selection of vein-forming procambial initial cells from within the ground meristem of a developing leaf, a process which is considered to be auxin-dependent, and continues until veins are anatomically differentiated with functional xylem and phloem. At present, the mechanisms responsible for leaf venation patterning are primarily characterized in the model eudicot Arabidopsis thaliana which displays a reticulate venation network. However, evidence suggests that vein development may proceed via a different mechanism in monocot leaves where venation patterning is parallel. Here, we employed Molecular Cartography, a multiplexed in situ hybridization technique, to analyse the spatiotemporal localisation of a subset of auxin related genes and candidate regulators of vein patterning in maize leaves. We show how different combinations of auxin influx and efflux transporters are recruited during leaf and vein specification, and how major and minor vein ranks develop with distinct identities. The localisation of the procambial marker PIN1a and the spatial arrangement of procambial initial cells that give rise to major and minor vein ranks further suggests that vein spacing is pre-patterned across the medio-lateral leaf axis prior to accumulation of the PIN1a auxin transporter. In contrast, patterning in the adaxial-abaxial axis occurs progressively, with markers of xylem and phloem gradually becoming polarised as differentiation proceeds. Collectively our data suggest that both lineage- and position-based mechanisms may underpin vein patterning in maize leaves. SIGNIFICANCE STATEMENTDuring the development of multicellular organisms specialized cell-types differentiate from pluripotent stem cells, with cell identity acquired via lineage- or position-based mechanisms. In plants, most organs develop post-embryogenesis and as such developmental processes are influenced by the external environment. To adapt to different environmental contexts and yet still form recognizable structures, position-based differentiation mechanisms are deployed in which cells adopt a certain fate depending on the activity of neighbouring cells. Such is the prevalence of position-based mechanisms in plant development that a role for lineage is rarely contemplated. Here we show that stem cells which give rise to different vein types in maize leaves are transcriptionally distinct, possibly reflecting a role for lineage-based mechanisms in the differentiation of leaf veins.

plant biology↗

bioRxiv2023

THE WIP6 TRANSCRIPTION FACTOR TOO MANY LATERALS SPECIFIES VEIN TYPE IN C4 AND C3 GRASS LEAVES

Grass leaves are invariantly strap shaped with an elongated distal blade and a proximal sheath that wraps around the stem. Underpinning this uniform shape is a scaffold of leaf veins, most of which extend in parallel lines along the proximo-distal leaf axis. Differences between species are apparent both in the types of veins that develop and in the spacing between them across the medio-lateral leaf axis. A prominent engineering goal is to increase vein density and the proportion of bundle sheath cells surrounding the veins in leaves of C3 photosynthesizing species such as rice, in order to facilitate introduction of the more efficient C4 photosynthetic pathway. Here we discover that the WIP6 zinc finger transcription factor TOO MANY LATERALS (TML) specifies vein rank in both maize (C4) and rice (C3), species with distinct venation patterns. Loss of function tml mutations lead to the development of large lateral veins in positions normally occupied by smaller intermediate veins. The spatial localization of TML transcripts in wild-type leaves is consistent with a role in suppressing lateral vein formation in procambial cells that develop intermediate veins, specifically the class of intermediate veins that extend from the leaf blade into the leaf sheath. Attempts to manipulate TML function in rice were unsuccessful because transgene expression was silenced, suggesting that precise spatial and temporal regulation of TML expression is essential during the regeneration of shoot tissue from callus. Given that transcriptome analysis demonstrated altered profiles of genes associated with cytokinin and auxin signaling in loss of function maize mutants, the necessity for tight regulation of TML gene expression could be an indirect consequence of hormonal inbalances as opposed to ectopic activity of a specific downstream target. Importantly, however, loss of function mutants in rice display increased vascular and bundle sheath cell occupancy in the leaf. Collectively this work provides an understanding of how vein rank is specified in grass leaves and a first step towards an anatomical chassis for C4 engineering in rice.

plant biology↗

bioRxiv2022

Mutations in NAKED-ENDOSPERM IDD genes reveal functional interactions with SCARECROW and a maternal influence on leaf patterning in C4 grasses

Leaves comprise a number of different cell-types that are patterned in the context of either the epidermal or inner cell layers. In grass leaves, two distinct anatomies develop in the inner leaf tissues depending on whether the leaf carries out C3 or C4 photosynthesis. In both cases a series of parallel veins develops that extends from the leaf base to the tip but in ancestral C3 species veins are separated by a greater number of intervening mesophyll cells than in derived C4 species. We have previously demonstrated that the GRAS transcription factor SCARECROW (SCR) regulates the number of photosynthetic mesophyll cells that form between veins in the leaves of the C4 species maize, whereas it regulates the formation of stomata in the epidermal leaf layer in the C3 species rice. Here we show that SCR is required for inner leaf patterning in the C4 species Setaria viridis but in this species the presumed ancestral stomatal patterning role is also retained. Through a comparative mutant analysis between maize, setaria and rice we further demonstrate that loss of NAKED-ENDOSPERM (NKD) INDETERMINATE DOMAIN (IDD) protein function exacerbates loss of function scr phenotypes in the inner leaf tissues of maize and setaria but not rice. Specifically, in both setaria and maize, scr;nkd mutants exhibit an increased proportion of fused veins with no intervening mesophyll cells, whereas inner leaf tissues are patterned normally in scr;nkd mutants of rice. Thus, combined action of SCR and NKD may control how many mesophyll cells are specified between veins in the leaves of C4 but not C3 grasses. Finally, we identified a maternal effect in maize in which maternally derived NKD can affect patterning of cells in leaf primordia that are initiated during embryogenesis. Together our results provide insight into the evolution of cell patterning in grass leaves, demonstrate a novel patterning role for IDD genes in C4 leaves and suggest that NKD can influence embryonic leaf development non-cell autonomously from the surrounding maternal tissue. Summary statementMutations in NKD IDD genes enhance loss of function scr phenotypes in the leaves of C4 grasses maize and Setaria viridis but not in the C3 grass rice, and reveal a maternal effect on cell-type patterning in leaves that are initiated during embryogenesis.

plant biology↗

bioRxiv2021

SCARECROW is deployed in distinct developmental contexts during rice and maize leaf development

The flexible deployment of developmental regulators is an increasingly appreciated aspect of plant development and evolution. The GRAS transcription factor SCARECROW (SCR) regulates the development of the endodermis in Arabidopsis and maize roots, but during leaf development it regulates the development of distinct cell-types; bundle-sheath in Arabidopsis and mesophyll in maize. In rice, SCR is implicated in stomatal patterning, but it is unknown whether this function is additional to a role in inner leaf patterning. Here, we demonstrate that two duplicated SCR genes function redundantly in rice. Contrary to previous reports, we show that these genes are necessary for stomatal development, with stomata virtually absent from leaves that are initiated after germination of mutants. The stomatal regulator OsMUTE is down-regulated in Osscr1;Osscr2 mutants indicating that OsSCR acts early in stomatal development. Notably, Osscr1;Osscr2 mutants do not exhibit the inner leaf patterning perturbations seen in Zmscr1;Zmscr1h mutants and Zmscr1;Zmscr1h mutants do not exhibit major perturbations in stomatal patterning. Taken together, these results indicate that SCR was deployed in different developmental contexts after the divergence of rice and maize around 50 million years ago. Summary statementThe transcription factor SCARECROW patterns stomata in rice leaves whereas in maize it predominantly patterns the inner leaf.

plant biology↗

bioRxiv2021

Acquisition of cell identity in the brown alga Ectocarpus: which of time, cell shape or position matters most?

During development, cells undergo simultaneous changes of different types that together depict cell "identity". In the multicellular brown alga Ectocarpus sp., while ageing, cells change shape and relative position within the filament. Understanding how these factors act and interact to specify cell identity requires markers of cell identity and the ability to genetically separate age, shape and position. Here we used laser capture microdissection (LCM) to isolate specific cell types from young sporophytes of Ectocarpus, and performed differential RNA-seq analysis. Transcriptome profiles of cell types in the wild-type strain provided signatures of the five cell types that can be identified by shape and position. In two mutants, where the relationship between cell shape, position and age are altered, transcriptome signatures revealed that little differential expression could be identified when only shape was perturbed. More generally, although the two mutants are characterised by opposite morphological phenotypes, their transcriptomes were remarkably similar. We concluded that despite the robustness of cell differentiation during WT development, neither the shape nor the position of the cell could serve as a faithful gauge for tracking differentiation.

developmental biology↗

bioRxiv2021

Conditional Stomatal Closure in a Fern Shares Molecular Features with Flowering Plant Active Stomatal Responses

Stomata evolved as plants transitioned from water to land, enabling carbon dioxide uptake and water loss to be controlled. In flowering plants, the most recently divergent land plant lineage, stomatal pores actively close in response to drought. In this response, the phytohormone abscisic acid (ABA) triggers signalling cascades that lead to ion and water loss in the guard cells of the stomatal complex, causing a reduction in turgor and pore closure. Whether this stimulus-response coupling pathway acts in other major land plant lineages is unclear, with some investigations reporting that stomatal closure involves ABA but others concluding that closure is passive. Here we show that in the model fern Ceratopteris richardii active stomatal closure is conditional on sensitisation by pre-exposure to either low humidity or exogenous ABA and is promoted by ABA. RNA-seq analysis and de novo transcriptome assembly reconstructed the protein coding complement of the C. richardii genome with coverage comparable to other plant models, enabling transcriptional signatures of stomatal sensitisation and closure to be identified. In both cases, changes in abundance of homologs of ABA, Ca2+ and ROS-related signalling components were observed, suggesting that the closure response pathway is conserved in ferns and flowering plants. These signatures further suggested that sensitisation is achieved by lowering the threshold required for a subsequent closure-inducing signal to trigger a response. We conclude that the canonical signalling network for active stomatal closure functioned in at least a rudimentary form in the stomata of the last common ancestor of ferns and flowering plants. Significance StatementStomata are valve-like pores that control the uptake of CO2 and the loss of water vapour in almost all land plants. In flowering plants, stomatal opening and closure is actively regulated by a stimulus-response coupling network. Whether active stomatal responses are present in other land plant lineages such as ferns has been hotly debated. Here we show that stomatal responses in the fern Ceratopteris richardii are active but depend on their past growth environment, and demonstrate that fern stomatal closure and sensitisation are associated with the altered expression of genes whose homologs function in the canonical stomatal regulatory network of flowering plants. Genetic pathways for active stomatal regulation therefore most likely evolved before the divergence of ferns and flowering plants.

plant biology↗

bioRxiv2020

SCARECROW gene function is required for photosynthetic development in maize

C4 photosynthesis in grasses relies on a specialized leaf anatomy. In maize, this ‘Kranz’ leaf anatomy is patterned in part by the duplicated SCARECROW (SCR) genes ZmSCR1 and ZmSCR1h. Here we show that in addition to patterning defects, chlorophyll content and levels of transcripts encoding Golden2-like regulators of chloroplast development are significantly lower in Zmscr1;Zmscr1h mutants than in wild-type. These perturbations are not associated with changes in chloroplast number, size or ultrastructure. However, the maximum rates of carboxylation by ribulose bisphosphate carboxylase/oxygenase (RuBisCO, Vcmax) and phosphoenolpyruvate carboxylase (PEPC, Vpmax) are both reduced, leading to perturbed plant growth. The CO2 compensation point and 13C‰ of Zmscr1;Zmscr1h plants are both normal, indicating that a canonical C4 cycle is operating, albeit at reduced overall capacity. Taken together, our results reveal that the maize SCR genes, either directly or indirectly, play a role in photosynthetic development.Significance statement SCARECROW (SCR) is one of the best studied plant developmental regulators, however, its role in downstream plant physiology is less well-understood. Here, we have demonstrated that SCR is required to establish and/or maintain photosynthetic capacity in maize leaves.Competing Interest StatementThe authors have declared no competing interest.View Full Text

plant biology↗

bioRxiv2019

Optimization of a Modular Steroid-Inducible Gene Expression System for Use in Rice

Chemically inducible systems that provide both spatial and temporal control of gene expression are essential tools, with many applications in plant biology. Using Golden Gate modular cloning, we have created a monocot-optimized dexamethasone (DEX)-inducible pOp6/LhGR system and tested its efficacy in rice using the reporter enzyme {beta}-glucuronidase (GUS). The system is tightly regulated and highly sensitive to DEX application, with six hours of induction sufficient to induce high levels of GUS activity in transgenic callus. In seedlings, GUS activity was detectable in the root after in vitro application of just 0.01M DEX. However, transgenic plants manifested severe developmental perturbations when grown on higher concentrations of DEX. The direct cause of these growth defects is not known, but the rice genome contains sequences with high similarity to the LhGR target sequence lacO, suggesting non-specific activation of endogenous genes by DEX induction. These off-target effects can be minimized by quenching with isopropyl {beta}-D-1-thiogalactopyranoside (IPTG). The system is thus suitable for general use in rice, when the method of DEX application and relevant controls are tailored appropriately for each specific application.

plant biology↗

bioRxiv2019

Redundant SCARECROW genes pattern distinct cell layers in roots and leaves of maize

The highly efficient C4 photosynthetic pathway is facilitated by Kranz leaf anatomy. In Kranz leaves, closely spaced veins are encircled by concentric layers of photosynthetic bundle sheath (inner) and mesophyll (outer) cells. Here we demonstrate that in the C4 monocot maize, Kranz patterning is regulated by redundant function of SCARECROW 1 (ZmSCR1) and a previously uncharacterized homeolog ZmSCR1h. ZmSCR1 and ZmSCR1h transcripts accumulate in ground meristem cells of developing leaf primordia and in Zmscr1;Zmscr1h mutant leaves, most veins are separated by one rather than two mesophyll cells; many veins have sclerenchyma above and/or below instead of mesophyll cells; and supernumerary bundle sheath cells develop. The mutant defects are unified by compromised mesophyll cell development. In addition to Kranz defects, Zmscr1;Zmscr1h mutants fail to form an organized endodermal layer in the root. Collectively, these data indicate that ZmSCR1 and ZmSCR1h redundantly regulate cell-type patterning in both leaves and roots of maize. Leaf and root pathways are distinguished, however, by the cell layer in which they operate - mesophyll at a two-cell distance from leaf veins versus endodermis immediately adjacent to root vasculature.\n\nSummary statementTwo duplicated maize SCARECROW genes control the development of the endodermis in roots and the mesophyll in leaves

plant biology↗