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Although photoreceptors are expressed throughout all plant organs, most studies have focused on their function in aerial parts with laboratory-grown plants. Photoreceptor function in naturally dark-grown roots of plants in their native habitats is lacking. We characterized patterns of photoreceptor expression in field- and glasshouse-grown Nicotiana attenuata plants, silenced the expression of PhyB1/B2/A/Cry2 whose root transcripts levels were greater/equal to those of shoots, and by micrografting combined empty vector transformed shoots onto photoreceptor-silenced roots, creating chimeric plants with "blind" roots but "sighted" shoots. Micrografting procedure was robust in both field and glasshouse, as demonstrated by transcript accumulation patterns, and a spatially-explicit lignin visual reporter chimeric line. Field- and glasshouse-grown plants with PhyB1B2, but not PhyA or Cry2, -blind roots, were delayed in stalk elongation compared with control plants, robustly for two field seasons. Wild-type plants with roots directly exposed to FR phenocopied the growth of irPhyB1B2-blind root grafts. Additionally, root-expressed PhyB1B2 was required to activate the positive photomorphogenic regulator, HY5, in response to aboveground light. We conclude that roots of plants growing deep into the soil in nature sense aboveground light, and possibly soil temperature, via PhyB1B2 to control key traits, such as stalk elongation.

The circadian clock is known to increase plant growth and fitness, and is thought to prepare plants for photosynthesis at dawn and dusk; whether this happens in nature was unknown. We transformed the native tobacco, Nicotiana attenuata to silence two core clock components, NaLHY (irLHY) and NaTOC1 (irTOC1). We characterized growth and light- and dark-adapted photosynthetic rates (Ac) throughout a 24 h day in empty vector-transformed (EV), irLHY, and irTOC1 plants in the field, and in NaPhyA- and NaPhyB1-silenced plants in the glasshouse. The growth rates of irLHY plants were lower than those of EV plants in the field. While irLHY plants reduced Ac earlier at dusk, no differences between irLHY and EV plants were observed at dawn in the field. irLHY, but not EV plants, responded to light in the night by rapidly increasing Ac. Under controlled conditions, EV plants rapidly increased Ac in the day compared to dark-adapted plants at night; irLHY plants lost these time-dependent responses. The role of NaLHY in gating photosynthesis is independent of the light-dependent reactions and red light perceived by NaPhyA, but not NaPhyB1. In summary, the circadian clock allows plants not to respond photosynthetically to light at night by anticipating and gating red light-mediated in native tobacco.

The highly conserved core circadian clock component TIMING OF CAB EXPRESSION 1 (TOC1) contextualizes environmental stress responses in plants, for example by gating abscisic acid (ABA) signaling and suppressing thermoresponsive growth. Selective interaction of TOC1 with PHYTOCHROME B (PHYB) under far-red-enriched light suggests a connection between circadian gating of light responses and sensitivity to ABA, an important regulator of growth and stress responses, including under drought. However, the fitness consequences of TOC1 function, particularly in the root, are poorly understood. Here, we used the desert annual, Nicotiana attenuata, to investigate the function of TOC1 in shoots and roots for maintaining fitness under drought, in both field and glasshouse experiments. Despite marked decreases in leaf water loss, TOC1-deficient (irTOC1) lines failed to maintain fitness in response to drought stress as measured by total seed capsule production. Restoring TOC1 transcript levels in shoots via micrografting was sufficient to restore wild-type drought responses under field conditions. Microarrays identified a co-expression module in leaves strongly linking red and far-red light signaling to drought responses in a TOC1-dependent manner, but experiments with phytochrome-deficient lines revealed that the effects of TOC1 deficiency under drought cannot be attributed to changes in red/far-red light perception alone. Taken together, these results elucidate the sophisticated, tissue-dependent role of the circadian clock in maintaining fitness in the face of long-term abiotic stresses such as drought.

Phytochromes mainly function in photoautotrophic organisms to adjust growth in response to fluctuating light signals. The different isoforms of plant phytochromes often display both conserved and divergent roles, presumably to fine-tune plant responses to environmental signals and optimize fitness. Here we describe the distinct, yet partially redundant, roles of phytochromes NaPHYA, NaPHYB1 and NaPHYB2 in a wild tobacco species, Nicotiana attenuata using RNAi-silenced phytochrome lines. Consistent with results reported from other species, silencing the expression of NaPHYA or NaPHYB2 in N. attenuata had mild or no influence on plant development as long as NaPHYB1 was functional; whereas silencing the expression of NaPHYB1 alone strongly altered flowering time and leaf morphology. The contribution of NaPHYB2 became significant only in the absence of NaPHYB1; plants silenced for both NaPHYB1 and NaPHYB2 largely skipped the rosette-stage of growth to rapidly produce long, slender stalks that bore flowers early: hallmarks of the shade-avoidance responses. The phenotyping of phytochrome-silenced lines, combined with sequence and transcript accumulation analysis, suggest the independent functional diversification of the phytochromes, and a dominant role of NaPHYB1 and NaPHYB2 in N. attenuata's vegetative and reproductive development.

Leaves of Psychotria leiocarpa accumulate a major shoot-specific indole alkaloid, N,β- d -glucopyranosyl vincosamide (GPV), which has antioxidant, but no apparent antifeedant or allelopathic activity. The species is tolerant to high doses of UV-B, possibly due to constitutive GPV accumulation in leaves. In seedlings, GPV accumulation is induced by white light in a photosynthesis independent fashion. To better understand the regulation of the in planta GPV pool, detailed alkaloid profiles were examined. Light shift tests confirmed that GPV accumulation is light dependent. GPV was tightly regulated in its organ distribution, with highest concentrations in reproductive structures. Far-red and blue wavelengths promoted GPV accumulation without clear correlation with carbohydrate or protein concentrations or dry biomass in seedlings, thus indicating direct effects of light. Since light conditions are often associated with higher oxidative stress, light-induced accumulation of GPV may contribute to maintain redox balance. In vitro, GPV was an overall more effective antioxidant compared to closely related alkaloids. Indirect evidence of potential antioxidant properties of GPV in vivo was obtained by application of GPV on leaves of UV sensitive species exposed to high doses of UV-B, which significantly improved tolerance to this stress. In contrast to the known constitutive accumulation of GPV in leaves, the alkaloid concentration proved to be highly dynamic, changing during development, reproductive organogenesis and seedling irradiance treatments. Data further support a role for GPV as an oxidative stress protectant and provide a means to improve alkaloid yields in plant biomass for pharmacological applications.

The highly conserved core circadian clock component TIMING OF CAB EXPRESSION1 (TOC1) contextualizes environmental stress responses in plants, for example by gating abscisic acid signaling and suppressing thermoresponsive growth. Selective interaction of TOC1 with PHYTOCHROME B under far-red–enriched light suggests a connection between circadian gating of light responses and sensitivity to ABA, an important regulator of growth and stress responses, including under drought. However, the fitness consequences of TOC1 function, particularly in the root, are poorly understood. Here, we used the desert annual, Nicotiana attenuata, to investigate the function of TOC1 in shoots and roots for maintaining fitness under drought, in both field and glasshouse experiments. Despite marked decreases in leaf water loss, TOC1-deficient lines failed to maintain fitness in response to drought stress as measured by total seed capsule production. Restoring TOC1 transcript levels in shoots via micrografting was sufficient to restore wild-type drought responses under field conditions. Microarrays identified a coexpression module in leaves strongly linking red and far-red light signaling to drought responses in a TOC1-dependent manner, but experiments with phytochrome-deficient lines revealed that the effects of TOC1 deficiency under drought cannot be attributed to changes in red/far-red light perception alone. Taken together, these results elucidate the sophisticated, tissue-dependent role of the circadian clock in maintaining fitness in the face of long-term abiotic stresses such as drought.

While jasmonic acid (JA) signaling is widely accepted as mediating plant resistance to herbivores, and the importance of the roots in plant defenses is recently being recognized, the role of root JA in the defense of above-ground parts remains unstudied. To restrict JA impairment to the roots, we micrografted wildtype Nicotiana attenuata shoots to the roots of transgenic plants impaired in JA signaling and evaluated ecologically relevant traits in the glasshouse and in nature. Root JA synthesis and perception are involved in regulating nicotine production in roots. Strikingly, systemic root JA regulated local leaf JA and abscisic acid (ABA) concentrations, which were associated with differences in nicotine transport from roots to leaves via the transpiration stream. Root JA signaling also regulated the accumulation of other shoot metabolites; together these account for differences in resistance against a generalist, Spodoptera littoralis, and a specialist herbivore, Manduca sexta. In N. attenuata's native habitat, silencing root JA synthesis increased the shoot damage inflicted by Empoasca leafhoppers, which are able to select natural jasmonate mutants. Silencing JA perception in roots also increased damage by Tupiocoris notatus. We conclude that attack from above-ground herbivores recruits root JA signaling to launch the full complement of plant defense responses.

The circadian clock is known to increase plant growth and fitness, and is thought to prepare plants for photosynthesis at dawn and dusk; whether this happens in nature was unknown. We transformed the native tobacco, Nicotiana attenuata to silence two core clock components, NaLHY (irLHY) and NaTOC1 (irTOC1). We characterized growth and light- and dark-adapted photosynthetic rates (A

Light affects not only the development and physiology of the shoots (stems, leaves, and flowers) of plants but also the underground root system. Light triggers shoot cells to release signals that travel to the root and affect the development and physiology of the root system. Like shoot cells, root cells also have photoreceptors that can be activated by light, leading Lee et al . to investigate if light actually reaches these underground parts of the plant. Exposing Arabidopsis thaliana shoots to light while protecting the roots from light activated the photoreceptor phyB in the roots. In the root, phyB activated Hy5, a transcription factor that mediates cellular responses to light and was important for growth of the primary root and for root gravitropism, the proper downward orientation of roots. Arabidopsis stems efficiently conducted only certain wavelengths of light to the root tissues, and these conducted wavelengths activated phyB directly in the roots. These findings demonstrate that roots not only receive information about light conditions through signaling molecules that travel from the shoot to the root in response to light but also directly perceive light that is conducted through the plant tissues.

Phytochromes mainly function in photoautotrophic organisms to adjust growth in response to fluctuating light signals. The different isoforms of plant phytochromes often display both conserved and divergent roles, presumably to fine-tune plant responses to environmental signals and optimize fitness. Here we describe the distinct, yet partially redundant, roles of phytochromes NaPHYA, NaPHYB1 and NaPHYB2 in a wild tobacco species, Nicotiana attenuata using RNAi-silenced phytochrome lines. Consistent with results reported from other species, silencing the expression of NaPHYA or NaPHYB2 in N. attenuata had mild or no influence on plant development as long as NaPHYB1 was functional; whereas silencing the expression of NaPHYB1 alone strongly altered flowering time and leaf morphology. The contribution of NaPHYB2 became significant only in the absence of NaPHYB1; plants silenced for both NaPHYB1 and NaPHYB2 largely skipped the rosette-stage of growth to rapidly produce long, slender stalks that bore flowers early: hallmarks of the shade-avoidance responses. The phenotyping of phytochrome-silenced lines, combined with sequence and transcript accumulation analysis, suggest the independent functional diversification of the phytochromes, and a dominant role of NaPHYB1 and NaPHYB2 in N. attenuata's vegetative and reproductive development.

Psychollatine is an unusual indole alkaloid produced by Psychotria umbellata Vell, an Atlantic Forest understorey tree. Leaves, young inflorescences and fruit pulp displayed high amounts of psychollatine (from 2.5 to 4.5% dry wt). Seeds accumulated lower quantities (0.2% dry wt), and roots, trace amounts (0.06% dry wt). Alkaloid accumulation was not seasonally affected and leaf content was not significantly enhanced by wounding, exposure to salicylic acid, hydrogen peroxide, or UV. The alkaloid content in leaves decreased upon cutting exposure to auxin. Under strong UV, the amount of chlorophylls did not decrease compared to control contents at 48 and 72 h. Post-harvest psychollatine content in leaves was stable in different temperatures, except at 65 °C. Psychollatine was an effective singlet oxygen, superoxide, and peroxide quencher. These antioxidant properties and the constitutively high amount of psychollatine indicate a role for the alkaloid in oxidative stress responses.

Psychollatine is a monoterpene indole alkaloid produced and accumulated by Psychotria umbellata Vell. (Rubiaceae) leaves in relatively high amounts (approximately 3% of the dry weight). The alkaloid has been shown to display opioid-like analgesic, anxiolytic, antidepressive and antipsychotic activities in rodents. In vitro assays suggested a protective role for this molecule in plant oxidative stress responses. This work reports antioxidant properties of psychollatine and the crude foliar extract from P. umbellata in strains of Saccharomyces cerevisiae proficient and deficient in antioxidant defenses exposed to H2O2 and paraquat. The antimutagenic activity of P. umbellata and its main alkaloid were assayed in S. cerevisiae N123 strain in presence of H2O2. Moreover, the antioxidant capacity of these substances on the hydroxyl radical (OH ) was investigated, using the hypoxanthine/xanthine oxidase assay. Psychollatine and the crude foliar extract of P. umbellata showed protective effect against oxidative stress in yeast, acting both as antioxidant and antimutagenic agents.

Psychotria umbellata Vell. (Rubiaceae), a Brazilian coastal woody species, produces umbellatine (also known as psychollatine), an analgesic indole alkaloid. An in vitro embryogenic regeneration protocol capable of yielding alkaloid-accumulating plants was developed. Rhizogenic calli, which were obtained from stem segments derived from rooted apical cuttings, were cultured on Murashige and Skoog's (MS) medium containing either 1 mg l(-1) NAA (naphthalene acetic acid) and no kinetin, or 5 mg l(-1) NAA + 1 mg l(-1) kinetin. Calli did not accumulate umbellatine. Segments of rhizogenic callus were cultured on complete MS medium with various concentrations of kinetin and sucrose. Plant regeneration was best in the light with 0.25 mg l(-1) of kinetin and 1.5% sucrose. After 3 months of acclimatization in soil mixture, plant survival was 81%. Leaves of 10-month-old regenerated plants yielded umbellatine concentrations equivalent to those of adult forest-grown plants.

The monoterpene indole alkaloid brachycerine from Psychotria brachyceras has been shown to be induced by UV and to have in vitro antioxidant activity, indicating a possible protective role against the secondary effects of this radiation. In this work, we have studied the antioxidant properties of brachycerine and a crude foliar extract from P. brachyceras by using Saccharomyces cerevisiae strains proficient and deficient in antioxidant defenses. The mutagenic and antimutagenic potential of these substances were assayed in S.cerevisiae N123 strain in the presence and absence of H2O2. In addition, we tested the antioxidant capacity of brachycerine and a crude foliar extract from P. brachyceras on hydroxyl radicals (OH) using the hypoxanthine/xanthine oxidase assay. The results show that brachycerine and the crude foliar extract of P. brachyceras have antioxidant and antimutagenic effects in yeast and probably this action is mainly due to the scavenging of OH radicals.