Early Sucrose Degradation and the Dominant Sucrose Cleavage Pattern Influence Lycoris sprengeri Bulblet Regeneration In Vitro

Bulblet formation and development determine the quantitative and qualitative traits, respectively, of bulb yield for most flowering bulbs. For Lycoris species, however, the underlying molecular mechanism remains elusive. Here, clonal bulblets of Lycoris sprengeri (Ls) derived from the same probulb were used as explants to establish efficient and inefficient in vitro regeneration systems by adjusting the 6-benzyladenine (BA) concentrations in media. BA application did not change the biological processes among groups but led to earlier decreases in sucrose and total soluble sugar (TSS) contents. Correlation analyses showed that the BA treatments changed the interaction between carbohydrate and endogenous hormone contents during bulblet regeneration. We found that two sucrose degradation enzyme-related genes, cell wall invertase (CWIN) and sucrose synthase, exhibited exactly opposite expression patterns during the competence stage. In addition, the regeneration system that obtained more bulblets showed significantly higher expression of LsCWIN2 than those that obtained fewer bulblets. Our data demonstrate the essential role of BA in accelerating sucrose degradation and the selection of a dominant sucrose cleavage pattern at the competence stage of in vitro bulblet regeneration. We propose that a relatively active CWIN-catalyzed pathway at the competence stage might promote bulblet regeneration, thus influencing bulb yield.

Knocking out AtCALS7 disturbs photoassimilate distribution and weakens defence against phytoplasma infection

Callose accumulation around sieve pores, under control of Callose synthase 7 (AtCALS7), has been interpreted as a mechanical response to limit pathogen spread in phytoplasma-infected plants. AtCALS7 is also involved in sieve-pore development and, hence, in mass-flow regulation, carbohydrate metabolism and distribution, and plant growth. Multiple roles of AtCALS7 during phytoplasma infection were investigated in healthy and phytoplasma-infected [Chrysanthemum Yellows (CY)-phytoplasma] wild-type and Atcals7ko Arabidopsis lines. In keeping with an increased phytoplasma titre in Atcals7ko plants, floral stalk height of infected wild-type and mutant plants was reduced by, respectively, 88 and 100% in comparison to their healthy controls, suggesting a higher investment of host resources in phytoplasma growth in the absence of AtCALS7. The apparently increased susceptibility of mutants was investigated by microscopic, metabolic and molecular analyses. Infection influenced the sieve-pore functionality in wild-type plants, which hardly affected plant growth, and plasmodesmata in the cortex, a phenomenon less prominent in mutants. Infection also increased the level of some sugars (glucose, sucrose, myoinositol), but to the highest extent in mutants. Finally, infection induced a similar upregulation of gene expression of enzymes involved in sucrose cleavage (AtSUS5, AtSUS6) in mutants and wild-type plants and an upregulation of carbohydrate transmembrane transporters (AtSWEET11, AtSTP13, AtSUC3) in mutants only. A more effective plasmodesmal closure seemingly suppressed spread of phytoplasma effectors, which rendered wild-type plants less susceptible to infection, because gene expression of enzymes channeling carbohydrates towards phytoplasmas is less promoted

Comparative transcriptome analysis reveals key genes potentially related to organic acid and sugar accumulation in loquat

Organic acids and sugars are the primary components that determine the quality and flavor of loquat fruits. In the present study, major organic acids, sugar content, enzyme activities, and the expression of related genes were analyzed during fruit development in two loquat cultivars, ’JieFangZhong’ (JFZ) and ’BaiLi’ (BL). Our results showed that the sugar content increased during fruit development in the two cultivars; however, the organic acid content dramatically decreased in the later stages of fruit development. The differences in organic acid and sugar content between the two cultivars primarily occured in the late stage of fruit development and the related enzymes showed dynamic changes in activies during development. Phosphoenolpyruvate carboxylase (PEPC) and mNAD malic dehydrogenase (mNAD-MDH) showed higher activities in JFZ at 95 days after flowering (DAF) than in BL. However, NADP-dependent malic enzyme (NADP-ME) activity was the lowest at 95 DAF in both JFZ and BL with BL showing higher activity compared with JFZ. At 125 DAF, the activity of fructokinase (FRK) was significantly higher in JFZ than in BL. The activity of sucrose synthase (SUSY) in the sucrose cleavage direction (SS-C) was low at early stages of fruit development and increased at 125 DAF. SS-C activity was higher in JFZ than in BL. vAI and sucrose phosphate synthase (SPS) activities were similar in the two both cultivars and increased with fruit development. RNA-sequencing was performed to determine the candidate genes for organic acid and sugar metabolism. Our results showed that the differentially expressed genes (DEGs) with the greated fold changes in the later stages of fruit development between the two cultivars were phosphoenolpyruvate carboxylase 2 (PEPC2), mNAD-malate dehydrogenase (mNAD-MDH), cytosolic NADP-ME (cyNADP-ME2), aluminum-activated malate transporter (ALMT9), subunit A of vacuolar H+-ATPase (VHA-A), vacuolar H+-PPase (VHP1), NAD-sorbitol dehydrogenase (NAD-SDH), fructokinase (FK), sucrose synthase in sucrose cleavage (SS-C), sucrose-phosphate synthase 1 (SPS1), neutral invertase (NI), and vacuolar acid invertase (vAI). The expression of 12 key DEGs was validated by quantitative reverese transcription PCR (RT-qPCR). Our findings will help understand the molecular mechanism of organic acid and sugar formation in loquat, which will aid in breeding high-quality loquat cultivars.

Change in Sucrose Cleavage Pattern and Rapid Starch Accumulation Govern Lily Shoot-to-Bulblet Transition in vitro

In bulb crops, bulbing is a key progress in micropropagation and is the feature that most distinguishes bulbous crops from other plants. Generally, bulbing involves a shoot-to-bulblet transition; however, the underlying mechanism remains elusive. We explored this process by tracking the shoot-to-bulblet transition under different culture conditions. Rapid starch accumulation occurred at 15 days after transplanting (DAT) in the bulblet-inducing treatments as confirmed via histological observations and the significant elevation of starch synthesis related-gene transcription, including LohAGPS, LohAGPL, LohGBSS, LohSS, and LohSBE. However, for shoots that did not transition to bulblets and maintained the shoot status, much higher soluble sugars were detected. Interestingly, we observed a clear shift from invertase-catalyzed to sucrose synthase-catalyzed sucrose cleavage pattern based on the differential expression of LohCWIN and LohSuSy during the key transition stage (prior to and after bulbing at 0–15 DAT). Shoots that transitioned into bulblets showed significantly higher LohSuSy expression, especially LohSuSy4 expression, than shoots that did not transition. A symplastic phloem unloading pathway at the bulblet emergence stage (15 DAT) was verified via the 6(5)-carboxyfluorescein diacetate fluorescent tracer. We propose that starch is the fundamental compound in the shoot-to-bulblet transition and that starch synthesis is likely triggered by the switch from apoplastic to symplastic sucrose unloading, which may be related to sucrose depletion. Furthermore, this study is the first to provide a complete inventory of the genes involved in starch metabolism based on our transcriptome data. Two of these genes, LohAGPS1.2b and LohSSIIId, were verified by rapid amplification of cDNA ends cloning, and these data will provide additional support for Lilium research since whole genome is currently lacking.

Vacuolar invertase activity shapes photosynthetic stress response of Arabidopsis thaliana and stabilizes central energy supply

Stabilization of the central carbohydrate and energy metabolism plays a key role in plant stress response. As the primary photosynthetic products, carbohydrates are substrate for numerous metabolic and stress-protective reactions. Further, they have been shown to be involved in diverse signalling processes finally affecting and regulating plant stress response on a whole plant level. Sucrose metabolism is known to be central to many stress-related processes and enzymes catalysing its biosynthesis, transport and degradation have been shown to significantly impact stress resistance and acclimation output. However, due to the cyclic structure of sucrose metabolism involving sucrose cleavage in multiple compartments as well as energy-demanding re-synthesis via hexose phosphorylation, it is challenging to derive an unambiguous picture of its contribution to stress reactions. In the present study, a combined stress experiment comprising cold and high-light identified metabolism of sucrose and fumaric acid to significantly separate the stress response of a cold susceptible and a tolerant natural accession of Arabidopsis thaliana. Kinetic modelling and simulation of subcellular rates of invertase-driven sucrose cleavage revealed a contrasting picture between the susceptible and the tolerant accession pointing to an important role of vacuolar invertase during initial stress response. Using a T-DNA insertion mutant with a dramatically reduced invertase activity provided evidence for a central role of the enzyme in stabilizing photosynthesis and the central energy metabolism during freezing and high-light stress. Reducing vacuolar invertase activity to about 3% of the wild type resulted in a strong increase of ADP and ATP levels indicating a severe effect on cytosolic and plastidial energy balance. Together with a significant decrease of maximum quantum yield of photosystem II (Fv/Fm) these results suggest that vacuolar invertase activity stabilizes cytosolic energy metabolism by supplying hexose equivalents being phosphorylated in the cytosol. Finally, the accompanying ATP consumption is essential for cytosolic phosphate balance which directly affects photosynthetic performance by the supply of ADP being crucial for photosynthetic ATP production.

Mycorrhizal Fungi Regulate Root Responses and Leaf Physiological Activities in Trifoliate Orange

Plant responses to mycorrhization are mediated through secretion of certain signal molecules deposited in mycorrhizosphere in response to environmental stimuli. Responses of four arbuscular mycorrhizal fungi (AMF), namely Claroideoglomus etunicatum, Diversispora versiformis, Funneliformis mosseae, and Rhizoglomus intraradices on root morphology, lateral root (LR) number, and leaf carbohydrates, nitric oxide (NO), and calmodulin (CaM) changes were studied using trifoliate orange. Inoculation response of D. versiformis, F. mosseae, and R. intraradices registered significantly higher plant growth performance (plant height, stem diameter, leaf number, and shoot and root biomass), root morphological traits (total length, projected area, surface area, and volume), and LR number (first-, second-, third-, and forth-order), compared to un-inoculated response. Higher concentrations of CaM, NO, glucose, and fructose and lower sucrose level in leaves were observed in AMF-seedlings than in non-AMF seedlings. Correlation studies further revealed, root morphological traits and LR numbers were significantly negatively correlated with sucrose whereas positively correlated with glucose, fructose, NO, and CaM level in leaves. These results suggested, AMF-induced root modification is routed through sucrose cleavage and partly through changes in NO and CaM.

Differences in Sucrose Metabolism in Peach Fruit Stored at Chilling Stress versus Nonchilling Stress Temperatures

To explore differences in sucrose metabolism between peach fruit subjected to chilling stress (5 °C) and nonchilling stress (10 °C), sucrose concentration as well as the activities and gene expression levels for enzymes associated with sucrose metabolism were compared. Fruits stored at 5 °C accumulated higher concentrations of H2O2 and developed severe chilling injury (CI) compared with fruit kept at 10 °C. Activities and gene expression levels for enzymes related to sucrose metabolism, such as acid invertase (AI), neutral invertase (NI), sucrose synthase (SS), and sucrose phosphate synthase (SPS) were higher in fruit stored at 5 °C than at 10 °C throughout or late in storage. A sharp increase in net sucrose cleavage activity dramatically decreased sucrose concentration and increased reducing sugars at 5 °C. The sucrose concentration at 10 °C increased over the first 21 days and then declined slightly, and was higher than in fruit at 5 °C throughout storage. The increase in net sucrose cleavage activity at 5 °C is contrary to the expectation that biochemical reactions ordinarily proceed more rapidly with increasing temperature. We conclude that chilling stress stimulates the activities and transcription levels of enzymes involved in sucrose metabolism, resulting in increased sucrose cleavage.