Involvement of Polyamines in Ribosome Biogenesis and Identification of Potential Genes Associated with Spermine Resistance in Arabidopsis thaliana
[eng] Polyamines are small polycationic molecules that contain amino groups and are found in all living organisms. This study is aimed at acquiring a comprehensive understanding of the diverse roles of polyamines in plants, specifically in A. thaliana, in order to gain a better understanding of thei...
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| Tipo de recurso: | tesis doctoral |
| Estado: | Versión publicada |
| Fecha de publicación: | 2023 |
| País: | España |
| Institución: | Universidad de Barcelona |
| Repositorio: | Dipòsit Digital de la UB |
| OAI Identifier: | oai:diposit.ub.edu:2445/210801 |
| Acceso en línea: | https://hdl.handle.net/2445/210801 http://hdl.handle.net/10803/690808 |
| Access Level: | acceso abierto |
| Palabra clave: | Poliamines Arabidopsis thaliana Genètica vegetal Nutrició vegetal Polyamines Plant genetics Plant nutrition |
| Sumario: | [eng] Polyamines are small polycationic molecules that contain amino groups and are found in all living organisms. This study is aimed at acquiring a comprehensive understanding of the diverse roles of polyamines in plants, specifically in A. thaliana, in order to gain a better understanding of their mechanisms of action. Studies have revealed that polyamines increase general proteins synthesis and are necessary for efficient translation. While some progress has been made regarding the participation of polyamines in ribosome biogenesis in bacteria and, recently, also in mammalians, little is known about their contribution and mode of action in plants. Given our interest on plant defense, the first part of this study was focused on analyzing the ribosome complexes of plants treated with the polyamines putrescine (Put) and spermine (Spm) in combination or not with the purified pathogen associated molecular pattern (PAMP) flagellin22 (flg22). Our results evidenced that treatments with polyamines and flg22 lead to increases in polysome abundance, resulting in a global enhancement of translational activity. Riboproteomic analyses revealed the accumulation of specific ribosomal proteins in response to Put, but not to Spm. Put was also found in polysome fractions where Spm was absent, suggesting a specific effect for Put on ribosome function. Additionally, when analyzing mutants deficient in Put and Spm biosynthesis, we observed a differential accumulation of ribonucleoprotein complex binding proteins. The data suggests a new role for putrescine in plants and provide valuable information for future investigations on how polyamines may have modulatory and functional effects on the ribosome machinery. The second part of the study was focused on identifying mutants insensitive to Spm. For this, two strategies were followed: forward genetics through random EMS mutagenesis and reverse genetics through loss-of-function analyses. As part of our screening process, we administered exogenous Spm supplementation to identify mutants that display insensitivity to Spm. This is possible because exogenously supplied Spm inhibits the growth of A. thaliana seedlings in vitro, enabling us to pinpoint mutants that display resistance to Spm. The mutagenesis screen identified the Ammonium Transporter 2 (AMT2), the Putative Laccase 9 (LAC9), and a hypothetical protein (At5g28090) as candidate genes responsible for Spm tolerance in roots. Such tolerance was associated with reduced ROS production under Spm treatment. In addition, Copper-Amine Oxidase ɤ2 (CuAOɤ2) and mutations in ethylene pathway genes, EIN1, EIN2, EIN3, and EIN3EIL1, were identified as candidates underlying Spm tolerance in roots through reverse genetics. CuAOɤ2 mutation might be related to reduced ROS production. The lack of rescue using ethylene inhibitors and lack of reconstitution of the triple response by Spm feeding suggests an effect of Spm on ethylene signaling. Last, we conducted an RNA-seq experiment that revealed a correlation between exogenous Spm supplementation and responses to Fe deficiency. This correlation was further validated through RT-qPCR analysis, and the fact that Fe supplementation restored growth in photosynthetic tissues under Spm supplementation, along with the induction of coumarin production by exogenous Spm. Additionally, we observed more severe Fe deficiency symptoms in spms mutants, indicating that Spm homeostasis can significantly impact the availability of Fe. The observed interaction between Spm and Fe deficiency holds promising implications in the modulation of the nutritional status of plants by polyamines. Overall, these studies open new frontiers of exploration and underscores the significance of polyamines in shaping various fundamental aspects of plant biology with potential applications in agriculture. |
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