Leaf heteroblasty in Passiflora edulis as revealed by metabolic profiling and expression analyses of the microRNAs miR156 and miR172

Juvenile-to-adult phase transition is marked by changes in leaf morphology, mostly due to the temporal development of the shoot apical meristem, a phenomenon known as heteroblasty. Sugars and microRNA-controlled modules are components of the heteroblastic process in Arabidopsis thaliana leaves. However, our understanding about their roles during phase-changing in other species, such as Passiflora edulis, remains limited. Unlike Arabidopsis, P. edulis (a semi-woody perennial climbing vine) undergoes remarkable changes in leaf morphology throughout juvenile-to-adult transition. Nonetheless, the underlying molecular mechanisms are unknown.Here we evaluated the molecular mechanisms underlying the heteroblastic process by analysing the temporal expression of microRNAs and targets in leaves as well as the leaf metabolome during P. edulis development.Metabolic profiling revealed a unique composition of metabolites associated with leaf heteroblasty. Increasing levels of glucose and α-trehalose were observed during juvenile-to-adult phase transition. Accumulation of microRNA156 (miR156) correlated with juvenile leaf traits, whilst miR172 transcript accumulation was associated with leaf adult traits. Importantly, glucose may mediate adult leaf characteristics during de novo shoot organogenesis by modulating miR156-targeted PeSPL9 expression levels at early stages of shoot development.Altogether, our results suggest that specific sugars may act as co-regulators, along with two microRNAs, leading to leaf morphological modifications throughout juvenile-to-adult phase transition in P. edulis

ATLANTIC EPIPHYTES: a data set of vascular and non-vascular epiphyte plants and lichens from the Atlantic Forest

Epiphytes are hyper-diverse and one of the frequently undervalued life forms in plant surveys and biodiversity inventories. Epiphytes of the Atlantic Forest, one of the most endangered ecosystems in the world, have high endemism and radiated recently in the Pliocene. We aimed to (1) compile an extensive Atlantic Forest data set on vascular, non-vascular plants (including hemiepiphytes), and lichen epiphyte species occurrence and abundance; (2) describe the epiphyte distribution in the Atlantic Forest, in order to indicate future sampling efforts. Our work presents the first epiphyte data set with information on abundance and occurrence of epiphyte phorophyte species. All data compiled here come from three main sources provided by the authors: published sources (comprising peer-reviewed articles, books, and theses), unpublished data, and herbarium data. We compiled a data set composed of 2,095 species, from 89,270 holo/hemiepiphyte records, in the Atlantic Forest of Brazil, Argentina, Paraguay, and Uruguay, recorded from 1824 to early 2018. Most of the records were from qualitative data (occurrence only, 88%), well distributed throughout the Atlantic Forest. For quantitative records, the most common sampling method was individual trees (71%), followed by plot sampling (19%), and transect sampling (10%). Angiosperms (81%) were the most frequently registered group, and Bromeliaceae and Orchidaceae were the families with the greatest number of records (27,272 and 21,945, respectively). Ferns and Lycophytes presented fewer records than Angiosperms, and Polypodiaceae were the most recorded family, and more concentrated in the Southern and Southeastern regions. Data on non-vascular plants and lichens were scarce, with a few disjunct records concentrated in the Northeastern region of the Atlantic Forest. For all non-vascular plant records, Lejeuneaceae, a family of liverworts, was the most recorded family. We hope that our effort to organize scattered epiphyte data help advance the knowledge of epiphyte ecology, as well as our understanding of macroecological and biogeographical patterns in the Atlantic Forest. No copyright restrictions are associated with the data set. Please cite this Ecology Data Paper if the data are used in publication and teaching events. © 2019 The Authors. Ecology © 2019 The Ecological Society of Americ

Autophagy deficiency compromises alternative pathways of respiration following energy deprivation in Arabidopsis thaliana

Under heterotrophic conditions, carbohydrate oxidation inside the mitochondrion is the primary energy source for cellular metabolism. However, during energy-limited conditions, alternative substrates are required to support respiration. Amino acid oxidation in plant cells plays a key role in this by generating electrons that can be transferred to the mitochondrial electron transport chain via the electron transfer flavoprotein/ubiquinone oxidoreductase system. Autophagy, a catabolic mechanism for macromolecule and protein recycling, allows the maintenance of amino acid pools and nutrient remobilization. Although the association between autophagy and alternative respiratory substrates has been suggested, the extent to which autophagy and primary metabolism interact to support plant respiration remains unclear. To investigate the metabolic importance of autophagy during development and under extended darkness, Arabidopsis (Arabidopsis thaliana) mutants with disruption of autophagy (atg mutants) were used. Under normal growth conditions, atg mutants showed lower growth and seed production with no impact on photosynthesis. Following extended darkness, atg mutants were characterized by signatures of early senescence, including decreased chlorophyll content and maximum photochemical efficiency of photosystem II coupled with increases in dark respiration. Transcript levels of genes involved in alternative pathways of respiration and amino acid catabolism were up-regulated in atg mutants. The metabolite profiles of dark-treated leaves revealed an extensive metabolic reprogramming in which increases in amino acid levels were partially compromised in atg mutants. Although an enhanced respiration in atg mutants was observed during extended darkness, autophagy deficiency compromises protein degradation and the generation of amino acids used as alternative substrates to the respiration

Differential impact of amino acids on OXPHOS system activity following carbohydrate starvation in Arabidopsis cell suspensions

Plant respiration mostly depends on the activity of glycolysis and the oxidation of organic acids in the tricarboxylic acid cycle to synthesize ATP. However, during stress situations plant cells also use amino acids as alternative substrates to donate electrons through the electron-transfer flavoprotein (ETF)/ETF:ubiquinone oxidoreductase (ETF/ETFQO) complex to the mitochondrial electron transport chain (mETC). Given this, we investigated changes of the oxidative phosphorylation (OXPHOS) system in Arabidopsis thaliana cell culture under carbohydrate starvation supplied with a range of amino acids. Induction of isovaleryl-CoA dehydrogenase (IVDH) activity was observed under carbohydrate starvation which was associated with increased amounts of IVDH protein detected by immunoblotting. Furthermore, activities of the protein complexes of the mETC were reduced under carbohydrate starvation. We also observed that OXPHOS system activity behavior is differently affected by different amino acids and that proteins associated with amino acids catabolism are upregulated in cells following carbohydrate starvation. Collectively, our results support the contention that ETF/ETFQO is an essential pathway to donate electrons to the mETC and that amino acids are alternative substrates to maintain respiration under carbohydrate starvation

Knockout mutants are due to altered organic acid accumulation and an increase in both stomatal and mesophyll conductance

Stomata control the exchange of CO 2 and water vapor in land plants. Thus, whereas a constant supply of CO 2 is required to maintain adequate rates of photosynthesis, the accompanying water losses must be tightly regulated to prevent dehydration and undesired metabolic changes. Accordingly, the uptake or release of ions and metabolites from guard cells is necessary to achieve normal stomatal function. The AtQUAC1, an R-type anion channel responsible for the release of malate from guard cells, is essential for efficient stomatal closure. Here, we demonstrate that mutant plants lacking AtQUAC1 accumulated higher levels of malate and fumarate. These mutant plants not only display slower stomatal closure in response to increased CO 2 concentration and dark but are also characterized by improved mesophyll conductance. These responses were accompanied by increases in both photosynthesis and respiration rates, without affecting the activity of photosynthetic and respiratory enzymes and the expression of other transporter genes in guard cells, which ultimately led to improved growth. Collectively, our results highlight that the transport of organic acids plays a key role in plant cell metabolism and demonstrate that AtQUAC1 reduce diffusive limitations to photosynthesis, which, at least partially, explain the observed increments in growth under well-watered conditions

Enhanced photosynthesis and growth in atquac1 knockout mutants are due to altered organic acid accumulation and an increase in both stomatal and mesophyll conductance

Stomata control the exchange of CO2 and water vapor in land plants. Thus, whereas a constant supply of CO2 is required to maintain adequate rates of photosynthesis, the accompanying water losses must be tightly regulated to prevent dehydration and undesired metabolic changes. Accordingly, the uptake or release of ions and metabolites from guard cells is necessary to achieve normal stomatal function. The AtQUAC1, an R-type anion channel responsible for the release of malate from guard cells, is essential for efficient stomatal closure. Here, we demonstrate that mutant plants lacking AtQUAC1 accumulated higher levels of malate and fumarate. These mutant plants not only display slower stomatal closure in response to increased CO2 concentration and dark but are also characterized by improved mesophyll conductance. These responses were accompanied by increases in both photosynthesis and respiration rates, without affecting the activity of photosynthetic and respiratory enzymes and the expression of other transporter genes in guard cells, which ultimately led to improved growth. Collectively, our results highlight that the transport of organic acids plays a key role in plant cell metabolism and demonstrate that AtQUAC1 reduce diffusive limitations to photosynthesis, which, at least partially, explain the observed increments in growth under well-watered conditions

An L,L- diaminopimelate aminotransferase mutation leads to metabolic shifts and growth inhibition in Arabidopsis

Lysine (Lys) connects the mitochondrial electron transport chain to amino acid catabolism and the tricarboxylic acid cycle. However, our understanding of how a deficiency in Lys biosynthesis impacts plant metabolism and growth remains limited. Here, we used a previously characterized Arabidopsis mutant (dapat) with reduced activity of the Lys biosynthesis enzyme L,L-diaminopimelate aminotransferase to investigate the physiological and metabolic impacts of impaired Lys biosynthesis. Despite displaying similar stomatal conductance and internal CO2 concentration, we observed reduced photosynthesis and growth in the dapat mutant. Surprisingly, whilst we did not find differences in dark respiration between genotypes, a lower storage and consumption of starch and sugars was observed in dapat plants. We found higher protein turnover but no differences in total amino acids during a diurnal cycle in dapat plants. Transcriptional and two-dimensional (isoelectric focalization/SDS-PAGE) proteome analyses revealed alterations in the abundance of several transcripts and proteins associated with photosynthesis and photorespiration coupled with a high glycine/serine ratio and increased levels of stress-responsive amino acids. Taken together, our findings demonstrate that biochemical alterations rather than stomatal limitations are responsible for the decreased photosynthesis and growth of the dapat mutant, which we hypothesize mimics stress conditions associated with impairments in the Lys biosynthesis pathway