Biology subjects

Rojas-Pierce, M.

Publications and source records attributed to Rojas-Pierce, M..

6 recordsLinked to original sources

bioRxiv2025

The Effect of Plasma Activated Water on the Rhizosphere Composition of Arabidopsis and Solanum lycopersicum

The emerging field of plasma agriculture investigates the potential benefit of non-thermal plasma (NTP) for agricultural practices. NTP-treated water, referred to as plasma activated water (PAW), has been proposed as a sustainable alternative to conventional nitrogen (N) fertilizers. Growing demand for N fertilizer is concomitant with increased global food demands. PAW contains nitrate (NO3-) and reactive oxygen species, such as hydrogen peroxide (H2O2), which are fixed from atmospheric molecules via NTP. While early studies report positive effects of PAW on plant growth, its influence on plant-associated microbial communities remains poorly understood. Here, we compared the impacts of PAW or NO3- solutions on the rhizosphere microbial community of Arabidopsis thaliana and Solanum lycopersicum. PAW was generated by a radio frequency (RF) glow discharge plasma source and contained no measurable ROS, while the control solution contained an equivalent concentration of NO3-. No significant differences in alpha diversity were detected in either plant species microbiome after 5 weeks of treatment when grown in non-commercial potting substrate. Significant dissimilarity was found in terms of beta diversity, but the relative abundance of the sequenced genera suggested no functional differences in rhizosphere communities. Overall, PAW treatment did not adversely impact the rhizosphere microbiome in either Arabidopsis or tomato. These results support the use of PAW as an alternative N-fertilizer, though outcomes may differ for PAW solutions containing ROS.

plant biology↗

bioRxiv2025

Guard Cell-Enriched Phosphoproteome Reveals Phosphorylation of Endomembrane Proteins in Closed Stomata

Control of the stomatal aperture is multifaceted, involving a complex interplay of environmental cues and intracellular signaling pathways. It is well established that changes in ion gradients drive water movement into and out of the guard cell, thereby altering cell volume and modulating the opening or closing of the stomatal pore. These rapid responses are often regulated by phosphorylation cascades to efficiently transmit environmental status and either reduce water loss or enhance carbon assimilation. The role of endomembrane trafficking networks in stomatal dynamics is not well characterized. Here, we investigated the regulation of stomatal opening and closing by generating a proteome and phosphoproteome of guard cell-enriched tissue. This deep proteome captured a protein profile that was similar to previously characterized guard cell proteomes. The guard cell-enriched tissue with closed stomata showed greater levels of phosphorylation of proteins related to endomembrane trafficking and vacuoles when compared to both whole leaf tissue with closed stomata and guard cell-enriched tissue with open stomata. These results support the hypothesis that phosphorylation of endomembrane proteins may contribute to the regulation of stomatal movements.

plant biology↗

bioRxiv2025

Regulation of Vacuole Fusion in Stomata by Dephosphorylation of the HOPS subunit VPS39

Understanding how plants regulate water loss is important for improving crop productivity. Tight control of stomatal opening and closing is essential for the uptake of CO2 while mitigating water vapor loss. The opening of stomata is regulated in part by homotypic vacuole fusion, which is mediated by conserved homotypic vacuole protein sorting (HOPS) and vacuolar SNARE (soluble N-ethylmaleimide sensitive factor attachment protein receptors) complexes. HOPS tethers apposing vacuole membranes and promotes the formation of trans-SNARE complexes to mediate fusion. In yeast, HOPS dissociates from the assembled SNARE complex to complete vacuole fusion, but little is known about this process in plants. HOPS-specific subunits VACUOLE PROTEIN SORTING39 (VPS39) and VPS41 are required for homotypic plant vacuole fusion, and a computational model predicted that post-translational modifications of HOPS may be needed for plant stomatal vacuole fusion. Here, we characterized a viable T-DNA insertion allele of VPS39 which demonstrated a critical role of VPS39 in stomatal vacuole fusion. We found that VPS39 has increased levels of phosphorylation when stomata are closed versus open, and that VPS39 function in stomata and embryonic development requires dynamic changes in phosphorylation. Our data are consistent with VPS39 phosphorylation altering vacuole dynamics in response to environmental cues, similar to well-established phosphorylation cascades that regulate ion transport during stomatal opening. SIGNIFICANCE STATEMENTVacuole fusion is important for stomata opening but how it is regulated in response of stomata opening signals is not characterized. This research demonstrated the role of the HOPS complex in vacuole fusion in stomata, and it identified phosphorylation sites in the HOPS subunit VPS39 that are critical for vacuole fusion. One Ser residue was enriched in closed stomata and represents a putative site for control of vacuole fusion downstream of stomata opening signals.

plant biology↗

bioRxiv2025

Non-Thermal Plasma Activated Water is an Effective Nitrogen Fertilizer Alternative for Arabidopsis thaliana

Nitrogen (N) fixation with non-thermal plasmas has been proposed as a sustainable alternative to meet growing N fertilizer demands for agriculture. This technology generates Plasma Activated Water (PAW) with a range of chemical compositions, including different concentrations of nitrate (NO-) and hydrogen peroxide (H2O2), among other compounds. Potential use of PAW as an effective crop fertilizer necessitates a robust understanding of the underlying biology of the plant, which is not yet available. The lack of a unified standard in PAW production and the varying chemical make-up that results from different devices and protocols hampers comparative studies and adoption of this technology. The objective of this study was to compare the efficacy of two PAW solutions with differing concentrations of H2O2 produced from a Radio Frequency (RF) glow discharge plasma source. The effect of these solutions on plant growth, ROS accumulation, gene expression and heat stress response were compared to N-equivalent controls in the model plant Arabidopsis to assess their potential as an alternative N fertilizer. While PAW solutions lacking detectable H2O2 enhanced seedling growth, those containing approximately 0.3 {micro}M of H2O2 did not. ROS accumulation in root tissues was similar between PAW and chemically equivalent solutions, suggesting H2O2 is the primary ROS present in the PAW at the time of treatment. Gene expression studies showed induction of genes involved in N uptake and assimilation in PAW-treated seedlings. Pre- treatment of seedlings with PAW solutions containing H2O2 improved root growth under heat stress which indicates that this treatment may induce plant stress response pathways. Finally, mature plants showed similar growth when fertilized with PAW lacking H2O2 or NO3- control regimes for over 5 weeks indicating equivalency in chemical composition, plant nutrient uptake and utilization. Overall, these results demonstrate that PAW is an effective alternative to NO3- fertilizers for plant cultivation but the levels of H2O2 need to be carefully controlled.

plant biology↗

bioRxiv2023

Model-based inference of a plant-specific dual role for HOPS in regulating guard cell vacuole fusion

Stomata are the pores on a leaf surface that regulate gas exchange. Each stoma consists of two guard cells whose movements regulate pore opening and thereby control CO2 fixation and water loss. Guard cell movements depend in part on the remodeling of vacuoles, which have been observed to change from a highly fragmented state to a fused morphology during stomata opening. This change in morphology requires a membrane fusion mechanism that responds rapidly to environmental signals, allowing plants to respond to diurnal and stress cues. With guard cell vacuoles being both large and responsive to external signals, stomata represent a unique system in which to delineate mechanisms of membrane fusion. Fusion of vacuole membranes is a highly conserved process in eukaryotes, with key roles played by two multi-subunit complexes: HOPS (homotypic fusion and vacuolar protein sorting) and SNARE (soluble NSF attachment protein receptor). HOPS is a vacuole tethering factor that is thought to chaperone SNAREs from apposing vacuole membranes into a fusion-competent complex capable of rearranging membranes. To resolve a counter-intuitive observation regarding the role of HOPS in regulating plant vacuole morphology, we derived a quantitative model of vacuole fusion dynamics and used it to generate testable predictions about HOPS-SNARE interactions. We derived our model by applying simulation-based inference to integrate prior knowledge about molecular interactions with limited, qualitative observations of emergent vacuole phenotypes. By constraining the model parameters to yield the emergent outcomes observed for stoma opening - as induced by two distinct chemical treatments - we predicted a dual role for HOPS and identified a stalled form of the SNARE complex that differs from phenomena reported in yeast. We predict that HOPS has contradictory actions at different points in the fusion signaling pathway, promoting the formation of SNARE complexes, but limiting their activity. Author summaryPlants "breathe" through pores in their leaves where each pore is formed by two specialized cells called guard cells. To open these pores, guard cells change in volume. This volume change is controlled by water-filled organelles called vacuoles that morph from multiple small entities to a few large ones capable of taking up more water to reshape the cell. Specialized proteins in vacuole membranes make this change happen by pulling vacuoles together until they fuse. Some of these proteins reside in membranes, but others must be drawn to the membrane from the cells cytoplasm. Specific lipid molecules in the membrane play an important role in recruiting those proteins to the vacuole membrane. We previously made an unexpected finding that removing this lipid induces plant vacuole fusion. To make sense of this observation, we used a mathematical model to piece together our knowledge of the proteins involved in this process and what we know about the chemical treatments that cause vacuoles to morph. Using computer simulations, we uncovered new rules about how molecules interact in membranes to accomplish the task of vacuole fusion in plants. We think the rules uncovered through mathematical modeling allow plants to respond quickly to environmental cues.

systems biology↗

bioRxiv2022

Microgravity enhances the phenotype of Arabidopsis zigzag-1 and reduces the Wortmannin-induced vacuole fusion in root cells

The spaceflight environment of the International Space Station poses a multitude of stresses on plant growth including reduced gravity. Plants exposed to microgravity and other conditions on the ISS display root skewing, changes in gene expression and protein abundance that may result in changes in cell wall composition, antioxidant accumulation and modification of growth anisotropy. Systematic studies that address the effects of microgravity on cellular organelles are lacking but altered numbers and sizes of vacuoles have been detected in previous flights. The prominent size of plant vacuoles makes them ideal models to study organelle dynamics in space. Here, we used Arabidopsis zigzag1 (zig-1) as a sensitized genotype to study the effect of microgravity on plant vacuole fusion. Wortmannin was used to induce vacuole fusion in seedlings and a formaldehyde-based fixation protocol was developed to visualize plant vacuole morphology after sample return, using confocal microscopy. Our results indicate that microgravity enhances the zig-1 phenotype by reducing hypocotyl growth and vacuole fusion in some cells. This study demonstrates the feasibility of chemical inhibitor treatments for plant cell biology experiments in space.

plant biology↗