To assess the functional significance of adenosine salvage in plants, the cDNAs and genes encoding two isoforms of adenosine kinase (ADK) were isolated from Arabidopsis. The ADK1-and ADK2-coding sequences are very similar, sharing 92% and 89% amino acid and nucleotide identity, respectively. Each cDNA was overexpressed in Escherichia coli, and the catalytic activity of each isoform was determined. Both ADKs had similar catalytic properties with a K m and V max /K m for adenosine of 0.3 to 0.5 m and 5.4 to 22 L min Ϫ1 mg Ϫ1 protein, respectively. The K m and V max /K m for the cytokinin riboside N 6 (isopentenyl) adenosine are 3 to 5 m and 0.021 to 0.14 L min Ϫ1 mg Ϫ1 protein, respectively, suggesting that adenosine is the preferred substrate for both ADK isoforms. In Arabidopsis plants, both ADK genes are expressed constitutively, with the highest steady-state mRNA levels being found in stem and root. ADK1 transcript levels were generally higher than those of ADK2. ADK enzyme activity reflected relative ADK protein levels seen in immunoblots for leaves, flowers, and stems but only poorly so for roots, siliques, and dry seeds. The catalytic properties, tissue accumulation, and expression levels of these ADKs suggest that they play a key metabolic role in the salvage synthesis of adenylates and methyl recycling in Arabidopsis. They may also contribute to cytokinin interconversion.
Ultra-performance liquid chromatography-tandem mass spectrometry was used to establish the cytokinin profile of the bryophyte Physcomitrella patens (Hedw.) B.S.G.; of 40 analyzed cytokinins, 20 were detected. cis-Zeatin-riboside-O-glucoside, N 6 -(D 2 -isopentenyl)adenosine-5#-monophosphate (iPRMP), and trans-zeatin-riboside-O-glucoside were the most abundant intracellular cytokinins. In addition, the aromatic cytokinins N 6 -benzyladenosine (BAR), N 6 -benzyladenine, meta-, and orthotopolin were detected. Unexpectedly, the most abundant extracellular cytokinin was the nucleotide iPRMP, and its identity was confirmed by quadrupole time-of-flight mass spectrometry. The effects of overexpressing a heterologous cytokinin oxidase/ dehydrogenase (CKX; EC 1.4.3.18/1.5.99.12) gene (AtCKX2 from Arabidopsis [Arabidopsis thaliana]) on the intracellular and extracellular distribution of cytokinins was assessed. In cultures of CKX-transformed plants, ultra-performance liquid chromatography-tandem mass spectrometry measurements showed that there were pronounced reductions in the extracellular concentrations of-isopentenyl)adenosine (iPR), but their intracellular cytokinin concentrations were only slightly affected. In vitro and in vivo measured CKX activity was shown to be strongly increased in the transformants. Major phenotypic changes observed in the CKX-overexpressing plants included reduced and retarded budding, absence of sexual reproduction, and abnormal protonema cells. In bud-induction bioassays with wild-type Physcomitrella, the nucleotides iPRMP, trans-zeatin-riboside-5#-monophosphate, BAR monophosphate, and the cis-zeatin forms cZ and cZR had no detectable effects, while the activities displayed by other selected cytokinins were in the following order: iP . tZ . N 6 -benzyladenine . BAR . iPR . tZR . meta-topolin . dihydrozeatin . ortho-topolin. The results on wild type and CKX transgenics suggest that extracellular iP and iPR are the main cytokinins responsible for inducing buds in the bryophyte Physcomitrella. Cytokinin profile is discussed regarding the evolution of cytokinin biosynthetic pathways.Cytokinins play important roles as growth-regulating compounds in plants (Kieber, 2002). External applications of cytokinins to mosses have been shown to induce bud formation and, thus, the transition from filamentous, protonemic growth to the formation of gametophores (Bopp and Brandes, 1964;Reski and Abel, 1985). However, knowledge of the endogenous cytokinin profiles of mosses is incomplete, and it is unclear how their intracellular and extracellular distributions regulate developmental processes. We have therefore attempted to establish the cytokinin profile of Physcomitrella patens, a model organism for plant development and metabolism studies (Cove et al., 2006). Naturally occurring cytokinins are N 6 -substituted adenine derivatives bearing either an isoprenoid or an aromatic side chain. Isoprenoid forms include N 6 -(D 2 -isopentenyl)adenine (iP)-and zeatin (Z)-type cytokinins, which are characterized by the ...
SUMMARY All land plants (embryophytes) share a common ancestor that likely evolved from a filamentous freshwater alga. Elucidating the transition from algae to embryophytes – and the eventual conquering of Earth’s surface – is one of the most fundamental questions in plant evolutionary biology. Here, we investigated one of the organismal properties that might have enabled this transition: resistance to drastic temperature shifts. We explored the effect of heat stress in Mougeotia and Spirogyra, two representatives of Zygnematophyceae – the closest known algal sister lineage to land plants. Heat stress induced pronounced phenotypic alterations in their plastids, and high‐performance liquid chromatography‐tandem mass spectroscopy‐based profiling of 565 transitions for the analysis of main central metabolites revealed significant shifts in 43 compounds. We also analyzed the global differential gene expression responses triggered by heat, generating 92.8 Gbp of sequence data and assembling a combined set of 8905 well‐expressed genes. Each organism had its own distinct gene expression profile; less than one‐half of their shared genes showed concordant gene expression trends. We nevertheless detected common signature responses to heat such as elevated transcript levels for molecular chaperones, thylakoid components, and – corroborating our metabolomic data – amino acid metabolism. We also uncovered the heat‐stress responsiveness of genes for phosphorelay‐based signal transduction that links environmental cues, calcium signatures and plastid biology. Our data allow us to infer the molecular heat stress response that the earliest land plants might have used when facing the rapidly shifting temperature conditions of the terrestrial habitat.
SummaryIs there more than one pathway for cytokinin biosynthesis in Physcomitrella? Despite the apparent absence of adenylate-isopentenyltransferases, characterization of ipt1 knockout mutants points towards a second, tRNA-independent cytokinin biosynthesis pathway.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.
hi@scite.ai
10624 S. Eastern Ave., Ste. A-614
Henderson, NV 89052, USA
Copyright © 2024 scite LLC. All rights reserved.
Made with 💙 for researchers
Part of the Research Solutions Family.