Biology subjects

Benitez-Alfonso, Y.

Publications and source records attributed to Benitez-Alfonso, Y..

7 recordsLinked to original sources

bioRxiv2025

POLYGALACTURONASES REGULATED BY AUXIN facilitate root cell elongation in Arabidopsis thaliana via pectin remodeling

Root cell elongation, the main driver of root growth, is tightly associated with cell wall remodeling, particularly through pectin modifications, which facilitate cell wall loosening and strengthening while maintaining structural integrity. Root cell elongation is precisely regulated by the phytohormone auxin, which has long been known to inhibit this process. The molecular pathways through which auxin influences cell wall modifications remain poorly understood. In this study, we explore the transcriptional regulation of cell wall-related genes by auxin in Arabidopsis thaliana roots. The nuclear auxin pathway altered the expression of numerous cell-wall related genes, suggesting dynamic modification of the cell wall during root cell elongation. We identified novel root-specific polygalacturonases (PGs), enzymes involved in pectin degradation, which we termed POLYGALACTURONASES REGULATED BY AUXIN (PGRAs). PGRAs are expressed specifically in the root epidermis, beginning at the elongation zone. Our results demonstrate that induction of PGRA1 expression initially promotes root cell elongation, while long term overexpression inhibits root growth. Auxin downregulates PGRA1 in the elongation zone, and plants lacking PGRAs fail to increase root growth rate in response to reduced auxin levels. This suggests that auxin downregulates PGRA expression to prevent PGRA-mediated pectin remodeling, thereby contributing to inhibition of root cell elongation. We established a novel link between auxin signaling and pectin modifications in the control of cell growth. These findings provide new insights into the molecular mechanisms through which auxin regulates root cell elongation, highlighting the role of pectin matrix modifications in this process.

plant biology↗

bioRxiv2025

Modelling symplasmic transport: from biophysical basics to steeper root gradients

Development and spatial pattern formation are inherently linked. The coordinated determination of cell fates is crucial in any developmental process and requires extensive intercellular communication. Plants cells exchange many molecular signals via the symplasmic pathway, i.e., via plasmodesmata: narrow channels connecting the cytoplasm of neighbouring cells. Regulation of symplasmic transport is vital for normal plant development, and mutations that disrupt this regulation are often embryo or seedling lethal. In many tissues, symplasmic transport of small molecules is diffusion driven, resulting in a non-selective and bidirectional transport, although net directionality could arise from gradients. This has led to the (dogmatic) belief that symplasmic transport can only be detrimental to pattern formation, because signalling molecules cannot be confined, and gradients would fade. Here, we develop a detailed biophysical description of symplasmic transport to explore how plasmodesmata affect gradients in a linear tissue. We then apply the model in more complex tissue contexts, observing and explaining, e.g., that symplasmic transport may result in steeper gradients in the root apical meristem. In conclusion, our model provides a reference framework for estimating the consequences of symplasmic transport and explains how symplasmic transport can contribute to more robust developmental patterning.

plant biology↗

bioRxiv2024

Structurally diverse calloses/β-1,3-glucans in plant cell wall microdomains

Cell walls underpin the mechanics of cell growth, intercellular signalling, and defence against pathogenic organisms. {beta}-(1,3)-glucans (also known as callose) are polysaccharides found in plants, fungi, and some bacterial species. In developing plant organs, callose accumulates around intercellular channels (plasmodesmata) controlling cell-to-cell communication. We developed monoclonal antibodies for the detection of {beta}-(1,3)-glucans and using these identified distinct populations of callose differing in size and secondary structure. Callose sub-populations were in proximal but not overlapping cell wall microdomains implying distinct spatial and functional microenvironments. We also unveiled callose interaction with xyloglucan; another plant glycan regulating cell wall functions. This work challenges previous views demonstrating structural heterogeneity in plant callose and supporting interactions between glycans with roles in the regulation of cell wall properties and functions.

plant biology↗

bioRxiv2024

Callose Deficiency Modulates Plasmodesmata frequency and the intercellular space In Rice Anthers

Fertilization relies on pollen mother cells able to transit from mitosis-to-meiosis to supply gametes. This process involves remarkable changes at the molecular, cellular and physiological levels including (but not limited to) remodelling of cell wall. During meiosis onset, cellulose content at the pollen mother cell walls gradually declines with the concurrent deposition of the polysaccharide callose in anther locules. We aim to understand the biological significance of cellulose-to-callose turnover in pollen mother cells walls using electron microscopic analyses of rice flowers. Our observations indicate that in wild type anthers, the mitosis-to-meiosis transition coincides with a gradual reduction in the number of cytoplasmic connections called plasmodesmata. A mutant in the Oryza sativa callose synthase GSL5, impaired in callose accumulation in premeiotic and meiotic anthers, displayed a reduction in plasmodesmata frequency among pollen mother cells and tapetal cells suggesting a role for callose in plasmodesmata maintenance. In addition, a significant increase in cell-cell distance between pollen mother cells and impaired premeiotic cell shaping was observed in the mutant. The results suggest that cellulose-to-callose turnover during mitosis-meiosis transition is necessary to maintain cell-to-cell connections and optimal intercellular spacing among anther locular cells explaining the regulatory influence of callose metabolism during meiosis in flowering plants. HighlightsO_LICellulose-to-callose switch during meiosis onset in pollen mother cell walls correlates with changes in cytoplasmic connections called plasmodesmata, among meiocytes and between meiocytes and surrounding somatic cells. C_LIO_LIA mutant in GSL5 (glucan synthase like 5) affects callose deposition but no other cell wall component (e.g., cellulose) in premeiotic and meiotic anthers, except at dyad stage. C_LIO_LIImpaired callose synthesis is proposed to alter plasmodesmata frequency, the extracellular spacing and the shaping of pollen mother cells during mitosis-to-meiosis transition. C_LIO_LIIncrease in plasmodesmata frequency negatively correlates with cell-cell distance between pollen mother cells in both wildtype and a callose mutant. C_LI

plant biology↗

bioRxiv2021

A phylogenetic and transcriptomic study of the β-1,3-glucanase family in tomato identifies candidate targets for fruit improvement

Tomato, Solanum lycopersicum, is one of the most cultivated fruits. However, between one-quarter and half of their production is lost during transport and storage. Modifications in cell walls, and specifically pectin composition, delay fruit softening but, so far, the impact of callose metabolism in this process has not been investigated. Callose accumulates in cell walls around plasmodesmata to modify symplasmic transport. It also plays a role in reinforcing cell walls in response to bruising or pathogen invasion. The aim of this work is to identify cell wall {beta}-1,3-glucanases expressed in tomato fruit that can be used as targets to modify callose accumulation during ripening. A phylogenetic analysis identified fifty candidate {beta} -1,3-glucanases in tomato distributed in three clusters (, {beta} and {gamma}) with evolutionary relations previously characterised in the model Arabidopsis thaliana. Analysis of tomato microarray data indicates different regulatory patterns: the expression of a subset of enzymes in cluster decreased during ripening, while enzymes in cluster {beta} and {gamma} displayed higher expression in white-red stages. qRT-PCR experiments confirm the differential regulation of enzymes in different clusters suggesting evolutionary divergences that correlate with differences in their predicted localization and function. The potential to exploit this information in the selection of targets to modify cell walls and fruit development is discussed.

plant biology↗

bioRxiv2021

Comparative meta-proteomic analysis for the identification of novel plasmodesmata proteins and regulatory cues.

A major route for cell-to-cell signaling is via cell wall-embedded pores termed plasmodesmata (PD) forming the symplasm. PD regulate many aspects of plant development and responses to the environment however, our understanding of what factors affect their structure and permeability is limited. In this paper, a meta-analysis is presented as a tool for the identification of conditions affecting PD transport and in silico generation of PD proteomes for species of interest. The custom-built pipeline searches the whole genome for protein structural features and conserved domains identified on experimental proteomes and use it to predict PD candidates in 22 compatible plant species. Using the in silico proteome and microarray analysis, interactions between PD genes and conditions affecting PD function are identified. High salinity and osmotic stress affect a significant number of PD candidate genes and we provide evidence that these conditions regulate symplasmic transport of GFP. Using the pipeline, the in silico PD proteome for Medicago truncatula was generated, as an example of a plant in which experimental data is not available. The identification of a candidate receptor like protein was experimentally validated in M. truncatula transgenic roots expressing fluorescently tagged protein fusion. Together the results highlight the power of our newly designed tool in the identification of new factors and proteins influencing PD in diverse plant species.

plant biology↗

bioRxiv2019

Plasma membrane associated Receptor Like Kinases relocalise to plasmodesmata in response to osmotic stress.

Plasmodesmata act as key elements in intercellular communication, coordinating processes related to plant growth, development and responses to environmental stresses. While many of the developmental, biotic and abiotic signals are primarily perceived at the plasma membrane (PM) by receptor proteins, plasmodesmata also cluster receptor-like activities and whether or not these two pathways interact is currently unknown.\n\nHere we show that specific PM-located Leucine-Rich-Repeat Receptor-Like-Kinases (LRR-RLKs), KIN7 and IMK2, which under optimal growth conditions are absented from plasmodesmata, rapidly relocate and cluster to the pores in response to osmotic stress. This process is remarkably fast, it is not a general feature of PM-associated proteins and is independent of sterol- and sphingolipid-membrane composition. Focusing on KIN7, previously reported to be involved in stress responses, we show that relocalisation upon mannitol depends on KIN7 phosphorylation. Loss-of-function mutation in KIN7 induces delay in lateral root (LR) development and the mutant is affected in the root response to mannitol stress. Callose-mediated plasmodesmata regulation is known to regulate LR development. We found that callose levels are reduced in kin7 mutant background with a root phenotype resembling ectopic expression of PdBG1, an enzyme that degrades callose at the pores. Both the LR and callose phenotypes can be complemented by expression of KIN7 -wild-type and -phosphomimic variants but not by KIN7 phosphodead mutant which fails to relocalise at plasmodesmata. Together the data indicate that re-organisation of RLKs to plasmodesmata is important for the regulation of callose and LR development as part of the plant response to osmotic stress.

plant biology↗