TY - JOUR A1 - Watanabe, Mutsumi A1 - Tohge, Takayuki A1 - Balazadeh, Salma A1 - Erban, Alexander A1 - Giavalisco, Patrick A1 - Kopka, Joachim A1 - Mueller-Roeber, Bernd A1 - Fernie, Alisdair R. A1 - Hoefgen, Rainer T1 - Comprehensive Metabolomics Studies of Plant Developmental Senescence JF - Plant Senescence: Methods and Protocols N2 - Leaf senescence is an essential developmental process that involves diverse metabolic changes associated with degradation of macromolecules allowing nutrient recycling and remobilization. In contrast to the significant progress in transcriptomic analysis of leaf senescence, metabolomics analyses have been relatively limited. A broad overview of metabolic changes during leaf senescence including the interactions between various metabolic pathways is required to gain a better understanding of the leaf senescence allowing to link transcriptomics with metabolomics and physiology. In this chapter, we describe how to obtain comprehensive metabolite profiles and how to dissect metabolic shifts during leaf senescence in the model plant Arabidopsis thaliana. Unlike nucleic acid analysis for transcriptomics, a comprehensive metabolite profile can only be achieved by combining a suite of analytic tools. Here, information is provided for measurements of the contents of chlorophyll, soluble proteins, and starch by spectrophotometric methods, ions by ion chromatography, thiols and amino acids by HPLC, primary metabolites by GC/TOF-MS, and secondary metabolites and lipophilic metabolites by LC/ESI-MS. These metabolite profiles provide a rich catalogue of metabolic changes during leaf senescence, which is a helpful database and blueprint to be correlated to future studies such as transcriptome and proteome analyses, forward and reverse genetic studies, or stress-induced senescence studies. KW - Senescence KW - Metabolomics KW - Arabidopsis KW - GC/MS KW - LC/MS KW - HPLC KW - IC Y1 - 2018 SN - 978-1-4939-7672-0 SN - 978-1-4939-7670-6 U6 - https://doi.org/10.1007/978-1-4939-7672-0_28 SN - 1064-3745 SN - 1940-6029 VL - 1744 SP - 339 EP - 358 PB - Humana Press CY - Totowa ER - TY - JOUR A1 - Zhao, Liming A1 - Xia, Yan A1 - Wu, Xiao-Yuan A1 - Schippers, Jos H. M. A1 - Jing, Hai-Chun T1 - Phenotypic analysis and molecular markers of leaf senescence JF - Plant Senescence: Methods and Protocols N2 - The process of leaf senescence consists of the final stage of leaf development. It has evolved as a mechanism to degrade macromolecules and micronutrients and remobilize them to other developing parts of the plant; hence it plays a central role for the survival of plants and crop production. During senescence, a range of physiological, morphological, cellular, and molecular events occur, which are generally referred to as the senescence syndrome that includes several hallmarks such as visible yellowing, loss of chlorophyll and water content, increase of ion leakage and cell death, deformation of chloroplast and cell structure, as well as the upregulation of thousands of so-called senescence-associated genes (SAGs) and downregulation of photosynthesis-associated genes (PAGs). This chapter is devoted to methods characterizing the onset and progression of leaf senescence at the morphological, physiological, cellular, and molecular levels. Leaf senescence normally progresses in an age-dependent manner but is also induced prematurely by a variety of environmental stresses in plants. Focused on the hallmarks of the senescence syndrome, a series of protocols is described to asses quantitatively the senescence process caused by developmental cues or environmental perturbations. We first briefly describe the senescence process, the events associated with the senescence syndrome, and the theories and methods to phenotype senescence. Detailed protocols for monitoring senescence in planta and in vitro, using the whole plant and the detached leaf, respectively, are presented. For convenience, most of the protocols use the model plant species Arabidopsis and rice, but they can be easily extended to other plants. KW - Leaf senescence KW - Visible yellowing KW - Chlorophyll KW - Ion leakage KW - Cell death KW - Senescence-associated genes (SAGs) KW - Arabidopsis KW - Rice Y1 - 2018 SN - 978-1-4939-7672-0 SN - 978-1-4939-7670-6 U6 - https://doi.org/10.1007/978-1-4939-7672-0_3 SN - 1064-3745 SN - 1940-6029 VL - 1744 SP - 35 EP - 48 PB - Humana Press Inc. CY - Totowa ER - TY - JOUR A1 - Streubel, Susanna A1 - Fritz, Michael Andre A1 - Teltow, Melanie A1 - Kappel, Christian A1 - Sicard, Adrien T1 - Successive duplication-divergence mechanisms at the RCO locus contributed to leaf shape diversity in the Brassicaceae JF - Development : Company of Biologists N2 - Gene duplication is a major driver for the increase of biological complexity. The divergence of newly duplicated paralogs may allow novel functions to evolve, while maintaining the ancestral one. Alternatively, partitioning the ancestral function among paralogs may allow parts of that role to follow independent evolutionary trajectories. We studied the REDUCED COMPLEXITY (RCO) locus, which contains three paralogs that have evolved through two independent events of gene duplication, and which underlies repeated events of leaf shape evolution within the Brassicaceae. In particular, we took advantage of the presence of three potentially functional paralogs in Capsella to investigate the extent of functional divergence among them. We demonstrate that the RCO copies control growth in different areas of the leaf. Consequently, the copies that are retained active in the different Brassicaceae lineages contribute to define the leaf dissection pattern. Our results further illustrate how successive gene duplication events and subsequent functional divergence can increase trait evolvability by providing independent evolutionary trajectories to specialized functions that have an additive effect on a given trait. KW - Plant development KW - Gene duplication KW - Leaf shape KW - Morphological evolution KW - Capsella KW - Arabidopsis Y1 - 2018 U6 - https://doi.org/10.1242/dev.164301 SN - 0950-1991 SN - 1477-9129 VL - 145 IS - 8 PB - Company of Biologists CY - Cambridge ER - TY - JOUR A1 - Kamranfar, Iman A1 - Xue, Gang-Ping A1 - Tohge, Takayuki A1 - Sedaghatmehr, Mastoureh A1 - Fernie, Alisdair R. A1 - Balazadeh, Salma A1 - Mueller-Roeber, Bernd T1 - Transcription factor RD26 is a key regulator of metabolic reprogramming during dark-induced senescence JF - New phytologist : international journal of plant science N2 - Leaf senescence is a key process in plants that culminates in the degradation of cellular constituents and massive reprogramming of metabolism for the recovery of nutrients from aged leaves for their reuse in newly developing sinks. We used molecular-biological and metabolomics approaches to identify NAC transcription factor (TF) RD26 as an important regulator of metabolic reprogramming in Arabidopsis thaliana. RD26 directly activates CHLOROPLAST VESICULATION (CV), encoding a protein crucial for chloroplast protein degradation, concomitant with an enhanced protein loss in RD26 over-expressors during senescence, but a reduced decline of protein in rd26 knockout mutants. RD26 also directly activates LKR/SDH involved in lysine catabolism, and PES1 important for phytol degradation. Metabolic profiling revealed reduced c-aminobutyric acid (GABA) in RD26 overexpressors, accompanied by the induction of respective catabolic genes. Degradation of lysine, phytol and GABA is instrumental for maintaining mitochondrial respiration in carbon-limiting conditions during senescence. RD26 also supports the degradation of starch and the accumulation of mono-and disaccharides during senescence by directly enhancing the expression of AMY1, SFP1 and SWEET15 involved in carbohydrate metabolism and transport. Collectively, during senescence RD26 acts by controlling the expression of genes across the entire spectrum of the cellular degradation hierarchy. KW - Arabidopsis KW - fatty acid KW - primary metabolism KW - protein and amino acid degradation KW - respiration KW - senescence Y1 - 2018 U6 - https://doi.org/10.1111/nph.15127 SN - 0028-646X SN - 1469-8137 VL - 218 IS - 4 SP - 1543 EP - 1557 PB - Wiley CY - Hoboken ER - TY - JOUR A1 - Liu, Qinsong A1 - Vain, Thomas A1 - Viotti, Corrado A1 - Doyle, Siamsa M. A1 - Tarkowska, Danuse A1 - Novak, Ondrej A1 - Zipfel, Cyril A1 - Sitbon, Folke A1 - Robert, Stephanie A1 - Hofius, Daniel T1 - Vacuole integrity maintained by DUF300 proteins is required for brassinosteroid signaling regulation JF - Molecular plant N2 - Brassinosteroid (BR) hormone signaling controls multiple processes during plant growth and development and is initiated at the plasma membrane through the receptor kinase BRASSINOSTEROID INSENSITIVE1 (BRI1) together with co-receptors such as BRI1-ASSOCIATED RECEPTOR KINASE1 (BAK1). BRI1 abundance is regulated by endosomal recycling and vacuolar targeting, but the role of vacuole-related proteins in BR receptor dynamics and BR responses remains elusive. Here, we show that the absence of two DUF300 domain-containing tonoplast proteins, LAZARUS1 (LAZ1) and LAZ1 HOMOLOG1 (LAZ1H1), causes vacuole morphology defects, growth inhibition, and constitutive activation of BR signaling. Intriguingly, tonoplast accumulation of BAK1 was substantially increased and appeared causally linked to enhanced BRI1 trafficking and degradation in laz1 laz1h1 plants. Since unrelated vacuole mutants exhibited normal BR responses, our findings indicate that DUF300 proteins play distinct roles in the regulation of BR signaling by maintaining vacuole integrity required to balance subcellular BAK1 pools and BR receptor distribution. KW - brassinosteroid signaling KW - vacuole integrity KW - DUF300 proteins KW - tonoplast KW - Arabidopsis Y1 - 2018 U6 - https://doi.org/10.1016/j.molp.2017.12.015 SN - 1674-2052 SN - 1752-9867 VL - 11 IS - 4 SP - 553 EP - 567 PB - Cell Press CY - Cambridge ER -