Periodic Reporting for period 1 - SUMOCell (Study of molecular events triggering the cytosolic enrichment of SUMO activating enzyme and its implications in seed quality.)
Période du rapport: 2022-11-01 au 2024-10-31
In nature, plants must adapt their development to multiple environmental challenges involving numerous plant pathogens and suboptimal growth conditions related to drought, heat or salinity, among the most prevalent. Plant survival to these challenges is associated with significant changes in post-translational modifications (PTMs) of plant proteins. PTMs are among the earliest and most rapid plant responses to changes in the environment and trigger downstream molecular and cellular responses influencing plant growth and survival in a precise and timely manner. PTMs, either constitutive or reversible, determine the function of proteins and expand the diversity of the cellular proteome. PTMs induce changes in protein activity, turnover, subcellular localization, and interactions with other molecules, enabling a fine-tuning of cellular responses.
SUMO (Small Ubiquitin-like MOdifer) is an essential PTM belonging to the Ubl family of modifiers. In plants, SUMO regulates multiple biological processes, ranging from development to responses arising from environmental challenges. This biological versatility of plant SUMOylation offers novel opportunities for improving agricultural productivity. SUMO modulates protein activity through regulation of subcellular localization, protein activity and stability, and protein-protein interactions. Several studies point to a nuclear enrichment of SUMO conjugation machinery, which is consistent with a preferential nuclear localization of the SUMO targets. Nonetheless, non–nuclear targets exist and the molecular mechanisms that mediate their modification remain elusive. Dr Lois’s group has uncovered a novel regulatory mechanism consisting of the processing of the C-terminus of the SUMO E1-activating enzyme large subunit, SAE2. Processed SAE2 partially localizes to the cytosol and the processing activity is prominent at the transition between embryo growth and seed maturation. Accumulation of SAE2 in the cytosol promotes cytosolic SUMOylation and behave as a signal for growth arrest, suggesting that SAE2 processing could contribute to finalizing embryogenesis. SUMOCell aims to uncover the signals that trigger SAE2 cytosolic localization and the cellular responses activated by cytosolic SUMOylation. First, with Duke University partner, we will study SAE2 subcellular dynamics using super-resolution microscopy, under hormone treatments and stress challenges. Second, we will study the physiological processes regulated by SAE2 localization to the cytosol. For that, we will perform single-cell RNAseq studies from cells enriched in cytosolic SAE2. Bioinformatics analysis will be carried out by a placement at Sequentia SL. Finally, we will analyse physiological parameters related to seed quality. Data obtained from cell studies, transcriptomics and seed performance will be integrated in a signalling model to understand the role of cytosolic SUMO conjugation in seed development, providing a multidisciplinar training program.
SUMO (Small Ubiquitin-like MOdifer) is an essential PTM belonging to the Ubl family of modifiers. In plants, SUMO regulates multiple biological processes, ranging from development to responses arising from environmental challenges. This biological versatility of plant SUMOylation offers novel opportunities for improving agricultural productivity. SUMO modulates protein activity through regulation of subcellular localization, protein activity and stability, and protein-protein interactions. Several studies point to a nuclear enrichment of SUMO conjugation machinery, which is consistent with a preferential nuclear localization of the SUMO targets. Nonetheless, non–nuclear targets exist and the molecular mechanisms that mediate their modification remain elusive. Dr Lois’s group has uncovered a novel regulatory mechanism consisting of the processing of the C-terminus of the SUMO E1-activating enzyme large subunit, SAE2. Processed SAE2 partially localizes to the cytosol and the processing activity is prominent at the transition between embryo growth and seed maturation. Accumulation of SAE2 in the cytosol promotes cytosolic SUMOylation and behave as a signal for growth arrest, suggesting that SAE2 processing could contribute to finalizing embryogenesis. SUMOCell aims to uncover the signals that trigger SAE2 cytosolic localization and the cellular responses activated by cytosolic SUMOylation. First, with Duke University partner, we will study SAE2 subcellular dynamics using super-resolution microscopy, under hormone treatments and stress challenges. Second, we will study the physiological processes regulated by SAE2 localization to the cytosol. For that, we will perform single-cell RNAseq studies from cells enriched in cytosolic SAE2. Bioinformatics analysis will be carried out by a placement at Sequentia SL. Finally, we will analyse physiological parameters related to seed quality. Data obtained from cell studies, transcriptomics and seed performance will be integrated in a signalling model to understand the role of cytosolic SUMO conjugation in seed development, providing a multidisciplinar training program.
The general objective of SUMOCell aims to uncover the signals that trigger SAE2 cytosolic localization, and the cellular responses activated by cytosolic SUMOylation. To achieve this general objective the following specific objectives, distributed in two work packages (WP), were proposed:
1. Cell imaging studies to identify the signals that promote SAE2 cytosolic enrichment (WP1): as it will be elaborated in more detail below, this objective, composed by 3 key tasks, have been fully achieved. First, a detailed study of the best imaging conditions for these experiments has been carried out. This involved the analysis of the most suitable cell type, plant growth conditions as well as the implementation of a methodology specific to the needs of the project for this work package. Secondly, after establishing the most optimal imaging conditions, experiments involving chemical screening, hormone treatments and abiotic challenges were conducted to identify the signals involved in the regulation of SAE2 subcellular localization and nucleo-cytoplasmic partitioning.
2. Study of the physiological processes regulated by cytosolic SAE2 (WP2): this second WP consists of four tasks of which three are to be carried out in the next phase of the grant (return phase) so this objective has been partially addressed at this point of the action. Regarding first task, a preliminary transcriptomic study has produced very promising results suggesting a role of cytosolic SAE2 in the regulation of ethylene response in Arabidopsis thaliana seed.
1. Cell imaging studies to identify the signals that promote SAE2 cytosolic enrichment (WP1): as it will be elaborated in more detail below, this objective, composed by 3 key tasks, have been fully achieved. First, a detailed study of the best imaging conditions for these experiments has been carried out. This involved the analysis of the most suitable cell type, plant growth conditions as well as the implementation of a methodology specific to the needs of the project for this work package. Secondly, after establishing the most optimal imaging conditions, experiments involving chemical screening, hormone treatments and abiotic challenges were conducted to identify the signals involved in the regulation of SAE2 subcellular localization and nucleo-cytoplasmic partitioning.
2. Study of the physiological processes regulated by cytosolic SAE2 (WP2): this second WP consists of four tasks of which three are to be carried out in the next phase of the grant (return phase) so this objective has been partially addressed at this point of the action. Regarding first task, a preliminary transcriptomic study has produced very promising results suggesting a role of cytosolic SAE2 in the regulation of ethylene response in Arabidopsis thaliana seed.