Valérie Legué

Summary: Plants are critical for sustaining human life and planetary health. However, their potential to enable humans to survive and thrive beyond Earth remains unrealized. This Viewpoint presents a collective vision outlining priorities associated with plant science to support a new frontier of human existence. These priorities are drawn from the International Space Life Sciences Working Group (ISLSWG) Plants for Space Exploration and Earth Applications workshop, held at the European Low Gravity Research Association (ELGRA) conference in September 2024. First, we highlight transformative advances gained from using the ‘laboratory of space’ in understanding how plants respond to gravity and other stressors. Second, we introduce a new crop Bioregenerative Life Support System (BLSS) readiness level (BRL) framework – extending the existing Crop Readiness Level (CRL) – to assist in overcoming challenges to establish resilient, sustainable crop production. Materializing the vision of plants as enablers of space exploration will require innovative approaches, including predictive modeling, synthetic biology, robust Earth‐based analogue systems, and reliable space‐based instruments to monitor biological processes. Success relies upon a unified international community to promote sharing of resources, facilities, expertise, and data to accelerate progress. Ultimately, this work will both advance human space exploration and provide solutions to enhance sustainable plant production on Earth.

A fundamental function of an organ is the ability to perceive mechanical cues. Yet, how this is accomplished is not fully understood, particularly in plant roots. In plants, the majority of studies dealing with the effects of mechanical stress have investigated the aerial parts. However, in natural conditions roots are also subjected to mechanical cues, for example when the root encounters a hard obstacle during its growth or when the soil settles. To investigate root cellular responses to root compression, we developed a microfluidic system associated with a microvalve allowing the delivery of controlled and reproducible mechanical stimulations to the root. In this study, examining plants expressing the R-GECO1-mTurquoise calcium reporter, we addressed the root cell deformation and calcium increase induced by the mechanical stimulation. Lateral pressure applied on the root induced a moderate elastic deformation of root cortical cells and elicited a multicomponent calcium signal at the onset of the pressure pulse, followed by a second one at the release of the pressure. This indicates that straining rather than stressing of tissues is relevant to trigger the calcium signal. Although the intensity of the calcium response increases with the pressure applied, successive pressure stimuli led to a remarkable attenuation of the calcium signal. The calcium elevation was restricted to the tissue under pressure and did not propagate. Strain sensing, spatial restriction and habituation to repetitive stimulation represent the fundamental properties of root signalling in response to local mechanical stimulation. These data linking mechanical properties of root cells to calcium elevation contribute to elucidating the pathway allowing the root to adapt to the mechanical cues generated by the soil.

Advancements in plant space biology are required for the realization of human space exploration missions, where the re-supply of resources from Earth is not feasible. Until a few decades ago, space life science was focused on the impact of the space environment on the human body. More recently, the interest in plant space biology has increased because plants are key organisms in Bioregenerative Life Support Systems (BLSS) for the regeneration of resources and fresh food production. Moreover, plants play an important role in psychological support for astronauts. The definition of cultivation requirements for the design, realization, and successful operation of BLSS must consider the effects of space factors on plants. Altered gravitational fields and radiation exposure are the main space factors inducing changes in gene expression, cell proliferation and differentiation, signalling and physiological processes with possible consequences on tissue organization and organogenesis, thus on the whole plant functioning. Interestingly, the changes at the cellular and molecular levels do not always result in organismic or developmental changes. This apparent paradox is a current research challenge. In this paper, the main findings of gravity- and radiation-related research on higher plants are summarized, highlighting the knowledge gaps that are still necessary to fill. Existing experimental facilities to simulate the effect of space factors, as well as requirements for future facilities for possible experiments to achieve fundamental biology goals are considered. Finally, the need for making synergies among disciplines and for establishing global standard operating procedures for analyses and data collection in space experiments is highlighted.

Throughout their life, plant root are submitted to mechanical stresses due to pressure exerted by the soil. So far, few studies addressed root cell deformation and calcium signaling elicited by soil compression. In this study, we designed a microchip inspired by pneumatic microvalve concept in order to deliver a lateral pressure to the root of a plant expressing the RGECO1-mTurquoise calcium reporter. Lateral pressure applied on the root induced a moderate elastic deformation of root cortical cells and elicited a multicomponent calcium signal at the onset of the pressure pulse, followed by a second one at the release of the pressure. This indicates that straining rather than stressing of tissues is relevant to trigger the calcium signal. The calcium elevation was restricted to the tissue under pressure and did not propagate. Additionally, the calcium signals exhibited a remarkable attenuation upon repetitive stimulations. Significance statement In natural conditions, roots are subjected to diurnal hydraulic pressure variations producing a periodic root diameter fluctuation moreover during its progression, the lateral confinement of the root increases, generating additional lateral forces. Thus it is crucial for the root to be able to perceive and transduce these stimulations in order to progress in the heterogeneous soil network. Our microfluidic set-up, mimicking lateral soil pressure, revealed new features of the early root cellular responses. A cytosolic calcium elevation is observed upon increase but also upon release of the pressure, corresponding to variations in the shape of the cells of the cortex. Although the intensity of the calcium response increases with the pressure applied, successive pressure stimuli lead to attenuation of the calcium signal. Strain sensing and habituation to repetitive stimulation represent the fundamental properties of the “sensing system” allowing the root to adapt to the stresses generated by the soil.

Recalcitrant adventitious root (AR) development is a major hurdle in propagating commercially important woody plants. Although significant progress has been made to identify genes involved in subsequent steps of AR development, the molecular basis of differences in apparent recalcitrance to form AR between easy-to-root and difficult-to-root genotypes remains unknown. To address this, we generated cambium tissue-specific transcriptomic data from stem cuttings of hybrid aspen, T89 (difficult-to-root) and hybrid poplar OP42 (easy-to-root), and used transgenic approaches to verify the role of several transcription factors in the control of adventitious rooting. Increased peroxidase activity was positively correlated with better rooting. We found differentially expressed genes encoding reactive oxygen species scavenging proteins to be enriched in OP42 compared with T89. A greater number of differentially expressed transcription factors in cambium cells of OP42 compared with T89 was revealed by a more intense transcriptional reprograming in the former. PtMYC2, a potential negative regulator, was less expressed in OP42 compared with T89. Using transgenic approaches, we demonstrated that PttARF17.1 and PttMYC2.1 negatively regulate adventitious rooting. Our results provide insights into the molecular basis of genotypic differences in AR and implicate differential expression of the master regulator MYC2 as a critical player in this process.

Summary: Damage can be signalled by extracellular ATP (eATP) using plasma membrane (PM) receptors to effect cytosolic free calcium ion ([Ca²⁺]_cyt) increase as a second messenger. The downstream PM Ca²⁺ channels remain enigmatic. Here, the Arabidopsis thaliana Ca²⁺ channel subunit CYCLIC NUCLEOTIDE‐GATED CHANNEL2 (CNGC2) was identified as a critical component linking eATP receptors to downstream [Ca²⁺]_cyt signalling in roots. Extracellular ATP‐induced changes in single epidermal cell PM voltage and conductance were measured electrophysiologically, changes in root [Ca²⁺]_cyt were measured with aequorin, and root transcriptional changes were determined by quantitative real‐time PCR. Two cngc2 loss‐of‐function mutants were used: cngc2‐3 and defence not death1 (which expresses cytosolic aequorin). Extracellular ATP‐induced transient depolarization of Arabidopsis root elongation zone epidermal PM voltage was Ca²⁺ dependent, requiring CNGC2 but not CNGC4 (its channel co‐subunit in immunity signalling). Activation of PM Ca²⁺ influx currents also required CNGC2. The eATP‐induced [Ca²⁺]_cyt increase and transcriptional response in cngc2 roots were significantly impaired. CYCLIC NUCLEOTIDE‐GATED CHANNEL2 is required for eATP‐induced epidermal Ca²⁺ influx, causing depolarization leading to [Ca²⁺]_cyt increase and damage‐related transcriptional response.

Recalcitrant adventitious root (AR) development is a major hurdle in propagating commercially important woody plants. Although significant progress has been made to identify genes involved in subsequent steps of AR development, the molecular basis of differences in apparent recalcitrance to form AR between easy-to-root and difficult-to-root genotypes remains unknown. To address this, we generated cambium tissue-specific transcriptomic data from stem cuttings of hybrid aspen, T89 (difficult-to-root) and hybrid poplar OP42 (easy-to-root) and used transgenic approaches to verify the role of several transcription factors (TF) in the control of adventitious rooting. Increased peroxidase activity is positively correlated with better rooting. We found differentially expressed genes encoding Reactive Oxygen Species (ROS) scavenging proteins to be enriched in OP42 compared to T89. A higher number of differentially expressed TF in OP42 compared to T89 cambium cells was revealed by a more intense transcriptional reprograming in the former. PtMYC2 , a potential negative regulator, was less expressed in OP42 compared to T89. Using transgenic approaches, we have demonstrated that PttARF17.1 and PttMYC2.1 negatively regulate adventitious rooting. Our results provide insights into the molecular basis of genotypic differences in AR and implicate differential expression of the master regulator MYC2 as a critical player in this process.

Editorial: Higher Plants, Algae and Cyanobacteria in Space Environments

Analysis of the growth and orientation of roots of Arabidopsis mutants with differing root cap sizes and shapes indicates that the form of the cap affects root responses to variations in the strength of the growth medium.

Adenosine 5′-triphosphate (ATP) is an important extracellular signaling agent, operating in growth regulation, stomatal conductance, and wound response. With the first receptor for extracellular ATP now identified in plants (P2K1/DORN1) and a plasma membrane NADPH oxidase revealed as its target, the search continues for the components of the signaling cascades they command. The Arabidopsis root elongation zone epidermal plasma membrane has recently been shown to contain cation transport pathways (channel conductances) that operate downstream of P2K1 and could contribute to extracellular ATP (eATP) signaling. Here, patch clamp electrophysiology has been used to delineate two further conductances from the root elongation zone epidermal plasma membrane that respond to eATP, including one that would permit chloride transport. This perspective addresses how these conductances compare to those previously characterized in roots and how they might operate together to enable early events in eATP signaling, including elevation of cytosolic-free calcium as a second messenger. The role of the reactive oxygen species (ROS) that could arise from eATP’s activation of NADPH oxidases is considered in a qualitative model that also considers the regulation of plasma membrane potential by the concerted action of the various cation and anion conductances. The molecular identities of the channel conductances in eATP signaling remain enigmatic but may yet be found in the multigene families of glutamate receptor-like channels, cyclic nucleotide-gated channels, annexins, and aluminum-activated malate transporters.