Barry J. Pogson

Source to sink: regulation of carotenoid biosynthesis in plants.

Carotenoids are a diverse group of colourful pigments naturally found in plants, algae, fungi and bacteria. They play essential roles in development, photosynthesis, root-mycorrhizal interactions and the production of phytohormones, such as abscisic acid and strigolactone. Carotenoid biosynthesis is regulated throughout the life cycle of a plant with dynamic changes in composition matched to prevailing developmental requirements and in response to external environmental stimuli. There are key regulatory nodes in the pathway that control the flux of metabolites into the pathway and alter flux through the pathway. The molecular nature of the mechanisms regulating carotenoid biosynthesis, including evidence for metabolite feedback, transcription and epigenetic control as well as their accumulation, storage and degradation will be the focus of this review.

Identification of the Carotenoid Isomerase Provides Insight into Carotenoid Biosynthesis, Prolamellar Body Formation, and Photomorphogenesis

Carotenoids are essential photoprotective and antioxidant pigments synthesized by all photosynthetic organisms. Most carotenoid biosynthetic enzymes were thought to have evolved independently in bacteria and plants. For example, in bacteria, a single enzyme (CrtI) catalyzes the four desaturations leading from the colorless compound phytoene to the red compound lycopene, whereas plants require two desaturases (phytoene and ζ-carotene desaturases) that are unrelated to the bacterial enzyme. We have demonstrated that carotenoid desaturation in plants requires a third distinct enzyme activity, the carotenoid isomerase (CRTISO), which, unlike phytoene and ζ-carotene desaturases, apparently arose from a progenitor bacterial desaturase. The Arabidopsis CRTISO locus was identified by the partial inhibition of lutein synthesis in light-grown tissue and the accumulation of poly-cis-carotene precursors in dark-grown tissue of crtISO mutants. After positional cloning, enzymatic analysis of CRTISO expressed in Escherichia coli confirmed that the enzyme catalyzes the isomerization of poly-cis-carotenoids to all-trans-carotenoids. Etioplasts of dark-grown crtISO mutants accumulate acyclic poly-cis-carotenoids in place of cyclic all-trans-xanthophylls and also lack prolamellar bodies (PLBs), the lattice of tubular membranes that defines an etioplast. This demonstrates a requirement for carotenoid biosynthesis to form the PLB. The absence of PLBs in crtISO mutants demonstrates a function for this unique structure and carotenoids in facilitating chloroplast development during the first critical days of seedling germination and photomorphogenesis.

The Absence of ALTERNATIVE OXIDASE1a in Arabidopsis Results in Acute Sensitivity to Combined Light and Drought Stress

Treatment of Arabidopsis (Arabidopsis thaliana) alternative oxidase1a (aox1a) mutant plants with moderate light under drought conditions resulted in a phenotypic difference compared with ecotype Columbia (Col-0), as evidenced by a 10-fold increase in the accumulation of anthocyanins in leaves, alterations in photosynthetic efficiency, and increased superoxide radical and reduced root growth at the early stages of seedling growth. Analysis of metabolite profiles revealed significant changes upon treatment in aox1a plants typical of combined stress treatments, and these were less pronounced or absent in Col-0 plants. These changes were accompanied by alteration in the abundance of a variety of transcripts during the stress treatment, providing a molecular fingerprint for the stress-induced phenotype of aox1a plants. Transcripts encoding proteins involved in the synthesis of anthocyanins, transcription factors, chloroplastic and mitochondrial components, cell wall synthesis, and sucrose and starch metabolism changed, indicating that effects were not confined to mitochondria, where the AOX1a protein is located. Microarray and quantitative reverse transcription-polymerase chain reaction analysis revealed that transcripts typically induced upon stress treatment or involved in antioxidant defense systems, especially chloroplast-located antioxidant defense components, had altered basal levels in untreated aox1a plants, suggesting a significant change in the basal equilibrium of signaling pathways that regulate these components. Taken together, these results indicate that aox1a plants have a greatly altered stress response even when mitochondria or the mitochondrial electron transport chain are not the primary target of the stress and that AOX1a plays a broad role in determining the normal redox balance in the cell.

Evidence for a SAL1-PAP Chloroplast Retrograde Pathway That Functions in Drought and High Light Signaling in Arabidopsis

This work describes a chloroplast-to-nucleus retrograde signaling pathway involving SAL1, a chloroplast and mitochondrial enzyme that degrades the phosphonucleotide 3′-phosphoadenosine 5′-phosphate (PAP). In the absence of SAL1 or in response to drought and high light, PAP accumulates and can move between the chloroplast and the nucleus, where it inhibits exoribonucleases, thereby inducing stress-responsive genes. Compartmentation of the eukaryotic cell requires a complex set of subcellular messages, including multiple retrograde signals from the chloroplast and mitochondria to the nucleus, to regulate gene expression. Here, we propose that one such signal is a phosphonucleotide (3′-phosphoadenosine 5′-phosphate [PAP]), which accumulates in Arabidopsis thaliana in response to drought and high light (HL) stress and that the enzyme SAL1 regulates its levels by dephosphorylating PAP to AMP. SAL1 accumulates in chloroplasts and mitochondria but not in the cytosol. sal1 mutants accumulate 20-fold more PAP without a marked change in inositol phosphate levels, demonstrating that PAP is a primary in vivo substrate. Significantly, transgenic targeting of SAL1 to either the nucleus or chloroplast of sal1 mutants lowers the total PAP levels and expression of the HL-inducible ASCORBATE PEROXIDASE2 gene. This indicates that PAP must be able to move between cellular compartments. The mode of action for PAP could be inhibition of 5′ to 3′ exoribonucleases (XRNs), as SAL1 and the nuclear XRNs modulate the expression of a similar subset of HL and drought-inducible genes, sal1 mutants accumulate XRN substrates, and PAP can inhibit yeast (Saccharomyces cerevisiae) XRNs. We propose a SAL1-PAP retrograde pathway that can alter nuclear gene expression during HL and drought stress.

Global Changes in Gene Expression in Response to High Light in Arabidopsis

A range of environmental conditions can lead to oxidative stress; thus, a prompt and effective response to oxidative stress is crucial for the survival of plants. Microarray and northern-blot analyses were performed toward the identification of the factors and signaling pathways that enable plants to limit oxidative damage caused by exposure to high light (HL). Arabidopsis plants grown under moderate light (100 μmol m−2 s−1) were exposed to HL (1,000 μmol m−2 s−1) for 1 h. The microarray analyses revealed that exposure of Arabidopsis to HL caused an increase in known antioxidant genes, as well as several unknown genes. Some of these unknown genes had homologies to possible regulatory genes and metabolic enzymes. Furthermore, it was found that a range of chaperones were up-regulated in the HL treatment and that this induction was specifically due to the HL stress. The temporal expression under HL and different oxidative stress conditions of a subset of HL-responsive genes was confirmed via northern-blot analysis. Results from the arrays were also compared with publicly available microarray data sets from a range of different stress conditions at the Arabidopsis Functional Genomics Consortium. This cross comparison enabled the identification of genes that may be induced by changes in redox poise. Finally, to determine if the genes that were differentially expressed by HL stress were under similar transcriptional control, we analyzed the promoter sequences for the presence of common motifs.

Carotenoid Metabolism in Plants

Carotenoids are mostly C40 terpenoids, a class of hydrocarbons that participate in various biological processes in plants, such as photosynthesis, photomorphogenesis, photoprotection, and development. Carotenoids also serve as precursors for two plant hormones and a diverse set of apocarotenoids. They are colorants and critical components of the human diet as antioxidants and provitamin A. In this review, we summarize current knowledge of the genes and enzymes involved in carotenoid metabolism and describe recent progress in understanding the regulatory mechanisms underlying carotenoid accumulation. The importance of the specific location of carotenoid enzyme metabolons and plastid types as well as of carotenoid-derived signals is discussed.

Remodeled Respiration in ndufs4 with Low Phosphorylation Efficiency Suppresses Arabidopsis Germination and Growth and Alters Control of Metabolism at Night

Respiratory oxidative phosphorylation is a cornerstone of cellular metabolism in aerobic multicellular organisms. The efficiency of this process is generally assumed to be maximized, but the presence of dynamically regulated nonphosphorylating bypasses implies that plants can alter phosphorylation efficiency and can benefit from lowered energy generation during respiration under certain conditions. We characterized an Arabidopsis (Arabidopsis thaliana) mutant, ndufs4 (for NADH dehydrogenase [ubiquinone] fragment S subunit 4), lacking complex I of the respiratory chain, which has constitutively lowered phosphorylation efficiency. Through analysis of the changes to mitochondrial function as well as whole cell transcripts and metabolites, we provide insights into how cellular metabolism flexibly adapts to reduced phosphorylation efficiency and why this state may benefit the plant by providing moderate stress tolerance. We show that removal of the single protein subunit NDUFS4 prevents assembly of complex I and removes its function from mitochondria without pleiotropic effects on other respiratory components. However, the lack of complex I promotes broad changes in the nuclear transcriptome governing growth and photosynthetic function. We observed increases in organic acid and amino acid pools in the mutant, especially at night, concomitant with alteration of the adenylate content. While germination is delayed, this can be rescued by application of gibberellic acid, and root growth assays of seedlings show enhanced tolerance to cold, mild salt, and osmotic stress. We discuss these observations in the light of recent data on the knockout of nonphosphorylating respiratory bypass enzymes that show opposite changes in metabolites and stress sensitivity. Our data suggest that the absence of complex I alters the adenylate control of cellular metabolism.

Photoprotection in a zeaxanthin- and lutein-deficient double mutant of Arabidopsis

When light absorption by a plant exceeds its capacity for light utilization, photosynthetic light harvesting is rapidly downregulated by photoprotective thermal dissipation, which is measured as nonphotochemical quenching of chlorophyll fluorescence (NPQ). To address the involvement of specific xanthophyll pigments in NPQ, we have analyzed mutants affecting xanthophyll metabolism in Arabidopsis thaliana. An npq1 lut2 double mutant was constructed, which lacks both zeaxanthin and lutein due to defects in the violaxanthin de-epoxidase and lycopene ∈-cyclase genes. The npq1 lut2 strain had normal Photosystem II efficiency and nearly wild-type concentrations of functional Photosystem II reaction centers, but the rapidly reversible component of NPQ was completely inhibited. Despite the defects in xanthophyll composition and NPQ, the npq1 lut2 mutant exhibited a remarkable ability to tolerate high light.

Signaling from the endoplasmic reticulum activates brassinosteroid signaling and promotes acclimation to stress in Arabidopsis.

BR signaling may integrate stress responses and growth processes to optimize growth under challenging environmental conditions. Integrating Stress and Growth Brassinosteroids (BRs) are plant hormones that control various aspects of plant growth and development. BR signaling has also been implicated in plants’ ability to tolerate environmental stresses. Regulated intramembrane proteolysis (RIP) that occurs in the Golgi liberates membrane-bound transcription factors in response to various forms of cellular stress, such as endoplasmic reticulum (ER) stress, heat, and salt stress. Che et al. identified two transcription factors, bZIP17 and bZIP28, that were cleaved through endomembrane RIP and activated by heat and ER stress to mediate tolerance to stress. This RIP pathway enhanced BR signaling, suggesting a mechanism by which plants may optimize growth during challenging environmental conditions. The ability to acclimate to stresses enables plants to grow and develop under adverse environmental conditions. Regulated intramembrane proteolysis (RIP) triggered by endoplasmic reticulum (ER) stress mediates some forms of stress signaling. Brassinosteroids (BRs) have been implicated in plant adaptation to stress, but no mechanisms for activation have been discovered. Here, we reveal a connection between ER stress signaling and BR-mediated growth and stress acclimation. Arabidopsis transcription factors bZIP17 and bZIP28 were translocated from the ER through the Golgi, where they were proteolytically cleaved by site 2 protease and released to translocate into the nucleus. Stresses, including heat and inhibition of protein glycosylation, increased translocation of these two bZIPs to the nucleus. These nuclear-localized bZIPs not only activated ER chaperone genes but also activated BR signaling, which was required for stress acclimation and growth. Thus, these bZIPs link ER stress and BR signaling, which may be a mechanism by which plant growth and stress responses can be integrated.

Systemic and Intracellular Responses to Photooxidative Stress in Arabidopsis

As the sun tracks daily through the sky from east to west, different parts of the canopy are exposed to high light (HL). The extent of and mechanisms by which a systemic acquired acclimation (SAA) response might preacclimate shaded leaves that will be subsequently exposed to full sunlight is largely undefined. We investigated the role of an Arabidopsis thaliana zinc finger transcription factor, ZAT10, in SAA. ZAT10 overexpression resulted in enhanced tolerance to photoinhibitory light and exogenous H2O2, increased expression of antioxidative genes whose products are targeted to multiple subcellular compartments. Partial HL exposure of a leaf or leaves rapidly induced ZAT10 mRNA in distal, shaded photosynthetic tissues, including the floral stem, cauline leaves, and rosette, but not in roots. Fully 86% of fivefold HL-upregulated and 71% of HL-downregulated genes were induced and repressed, respectively, in distal, shaded leaves. Between 15 and 23% of genes whose expression changed in the HL and/or distal tissues were coexpressed in the ZAT10 overexpression plants, implicating ZAT10 in modulating the expression of SAA-regulated genes. The SAA response was detectable in plants with mutations in abscisic acid, methyl jasmonate, or salicylic acid synthesis or perception, and systemic H2O2 diffusion was not detected. Hence, SAA is distinct from pathogen-stimulated systemic acquired resistance and apparently involves a novel signal or combination of signals that preacclimate photosynthetic tissues to HL.