An interplay between bZIP16, bZIP68, and GBF1 regulates nuclear photosynthetic genes during photomorphogenesis in Arabidopsis.

The development of a seedling into a photosynthetically active plant is a crucial process. Despite its importance, we do not fully understand the regulatory mechanisms behind the establishment of functional chloroplasts. We herein provide new insight into the early light response by identifying the function of three basic region/leucine zipper (bZIP) transcription factors: bZIP16, bZIP68, and GBF1. These proteins are involved in the regulation of key components required for the establishment of photosynthetically active chloroplasts. The activity of these bZIPs is dependent on the redox status of a conserved cysteine residue, which provides a mechanism to finetune light-responsive gene expression. The blue light cryptochrome (CRY) photoreceptors provide one of the major light-signaling pathways, and bZIP target genes overlap with one-third of CRY-regulated genes with an enrichment for photosynthesis/chloroplast-associated genes. bZIP16, bZIP68, and GBF1 were demonstrated as novel interaction partners of CRY1. The interaction between CRY1 and bZIP16 was stimulated by blue light. Furthermore, we demonstrate a genetic link between the bZIP proteins and cryptochromes as the cry1cry2 mutant is epistatic to the cry1cry2bzip16bzip68gbf1 mutant. bZIP16, bZIP68, and GBF1 regulate a subset of photosynthesis associated genes in response to blue light critical for a proper greening process in Arabidopsis.

GENOMES UNCOUPLED1 plays a key role during the de-etiolation process in Arabidopsis.

One of the most dramatic challenges in the life of a plant occurs when the seedling emerges from the soil and exposure to light triggers expression of genes required for establishment of photosynthesis. This process needs to be tightly regulated, as premature accumulation of light-harvesting proteins and photoreactive Chl precursors causes oxidative damage when the seedling is first exposed to light. Photosynthesis genes are encoded by both nuclear and plastid genomes, and to establish the required level of control, plastid-to-nucleus (retrograde) signalling is necessary to ensure correct gene expression. We herein show that a negative GENOMES UNCOUPLED1 (GUN1)-mediated retrograde signal restricts chloroplast development in darkness and during early light response by regulating the transcription of several critical transcription factors linked to light response, photomorphogenesis, and chloroplast development, and consequently their downstream target genes in Arabidopsis. Thus, the plastids play an essential role during skotomorphogenesis and the early light response, and GUN1 acts as a safeguard during the critical step of seedling emergence from darkness.

A fully assembled plastid-encoded RNA polymerase complex detected in etioplasts and proplastids in Arabidopsis.

The plastid-encoded genes of higher plants are transcribed by at least two types of RNA polymerases, the nuclear-encoded RNA polymerase (NEP) and the plastid-encoded RNA polymerase (PEP). In mature photosynthesizing leaves, the vast majority of the genes are transcribed by PEP. However, the regulatory mechanisms controlling plastid transcription during early light response is unclear. Chloroplast development is suggested to be associated with a shift in the usage of the primary RNA polymerase from NEP to PEP as the expression of the plastid-encoded photosynthesis genes is induced upon light exposure. Assembly of the PEP complex has been suggested as a rate-limiting step for full activation of plastid-encoded photosynthesis gene expression. However, two sigma factor mutants, sig2 and sig6, with reduced PEP activity, showed significantly lower expression of the plastid-encoded photosynthesis genes already in the dark and during the first hours of light exposure indicating that PEP activity is required for basal expression of plastid-encoded photosynthesis genes in the dark and during early light response. Furthermore, in etioplasts and proplastids a fully assembled PEP complex was revealed on Blue Native PAGE. Our results indicate that a full assembly of the PEP complex is possible in the dark and that PEP drives basal transcriptional activity of plastid-encoded photosynthesis genes in the dark. Assembly of the complex is most likely not a rate-limiting step for full activation of plastid-encoded photosynthesis gene expression which is rather achieved either by the abundance of the PEP complex or by some posttranslational regulation of the individual PEP components.

Emerging from the darkness: interplay between light and plastid signaling during chloroplast biogenesis.

Chloroplast biogenesis is a highly complex process that requires carefully coordinated communication between the nucleus and the chloroplast to integrate light signaling and information about the state of the plastid through retrograde signals. Most studies on plastid development have been performed using dark-grown seedlings and have focused on the transition from etioplast to chloroplast in response to light. Some advances are now also being made to understand the transition directly from proplastids to chloroplasts as it occurs in the shoot apical meristems. Recent reports have highlighted the importance of repressive mechanisms to block premature chloroplast development in dark, both at the transcriptional and post-transcriptional level. A group of new proteins with dual plastid and nuclear localization were shown to take part in the light triggered degradation of PHYTOCHROME INTERACTING FACTORs (PIFs) in the nucleus and thereby release the suppression of the nuclear photosynthesis associated genes. These dually localized proteins are also required to activate transcription of photosynthesis genes in the plastid in response to light, emphasizing the close link between the nucleus and the plastids during early light response. Furthermore, development of a fully functional chloroplast requires a plastid signal but the nature of this signal(s) is still unknown. GENOMES UNCOUPLED1 (GUN1) is a plastid protein pivotal for retrograde signal(s) during early seedling development, and recent reports have revealed multiple interactors of GUN1 from different plastid processes. These new GUN1 interactors could reveal the true molecular function of the enigmatic character, GUN1, under naturally occurring adverse growth conditions.

Establishment of Photosynthesis through Chloroplast Development Is Controlled by Two Distinct Regulatory Phases.

Chloroplasts develop from undifferentiated proplastids present in meristematic tissue. Thus, chloroplast biogenesis is closely connected to leaf development, which restricts our ability to study the process of chloroplast biogenesis per se. As a consequence, we know relatively little about the regulatory mechanisms behind the establishment of the photosynthetic reactions and how the activities of the two genomes involved are coordinated during chloroplast development. We developed a single cell-based experimental system from Arabidopsis (Arabidopsis thaliana) with high temporal resolution allowing for investigations of the transition from proplastids to functional chloroplasts. Using this unique cell line, we could show that the establishment of photosynthesis is dependent on a regulatory mechanism involving two distinct phases. The first phase is triggered by rapid light-induced changes in gene expression and the metabolome. The second phase is dependent on the activation of the chloroplast and generates massive changes in the nuclear gene expression required for the transition to photosynthetically functional chloroplasts. The second phase also is associated with a spatial transition of the chloroplasts from clusters around the nucleus to the final position at the cell cortex. Thus, the establishment of photosynthesis is a two-phase process with a clear checkpoint associated with the second regulatory phase allowing coordination of the activities of the nuclear and plastid genomes.

Dual and dynamic intracellular localization of Arabidopsis thaliana SnRK1.1.

Sucrose non-fermenting 1 (SNF1)-related protein kinase 1.1 (SnRK1.1; also known as KIN10 or SnRK1α) has been identified as the catalytic subunit of the complex SnRK1, the Arabidopsis thaliana homologue of a central integrator of energy and stress signalling in eukaryotes dubbed AMPK/Snf1/SnRK1. A nuclear localization of SnRK1.1 has been previously described and is in line with its function as an integrator of energy and stress signals. Here, using two biological models (Nicotiana benthamiana and Arabidopsis thaliana), native regulatory sequences, different microscopy techniques, and manipulations of cellular energy status, it was found that SnRK1.1 is localized dynamically between the nucleus and endoplasmic reticulum (ER). This distribution was confirmed at a spatial and temporal level by co-localization studies with two different fluorescent ER markers, one of them being the SnRK1.1 phosphorylation target HMGR. The ER and nuclear localization displayed a dynamic behaviour in response to perturbations of the plastidic electron transport chain. These results suggest that an ER-associated SnRK1.1 fraction might be sensing the cellular energy status, being a point of crosstalk with other ER stress regulatory pathways.

The role of retrograde signals during plant stress responses.

Chloroplast and mitochondria not only provide the energy to the plant cell but due to the sensitivity of organellar processes to perturbations caused by abiotic stress, they are also key cellular sensors of environmental fluctuations. Abiotic stresses result in reduced photosynthetic efficiency and thereby reduced energy supply for cellular processes. Thus, in order to acclimate to stress, plants must re-program gene expression and cellular metabolism to divert energy from growth and developmental processes to stress responses. To restore cellular energy homeostasis following exposure to stress, the activities of the organelles must be tightly co-ordinated with the transcriptional re-programming in the nucleus. Thus, communication between the organelles and the nucleus, so-called retrograde signalling, is essential to direct the energy use correctly during stress exposure. Stress-triggered retrograde signals are mediated by reactive oxygen species and metabolites including ß-cyclocitral, MEcPP (2-C-methyl-d-erythritol 2,4-cyclodiphosphate), PAP (3'-phosphoadenosine 5'-phosphate), and intermediates of the tetrapyrrole biosynthesis pathway. However, for the plant cell to respond optimally to environmental stress, these stress-triggered retrograde signalling pathways must be integrated with the cytosolic stress signalling network. We hypothesize that the Mediator transcriptional co-activator complex may play a key role as a regulatory hub in the nucleus, integrating the complex stress signalling networks originating in different cellular compartments.

Circadian and Plastid Signaling Pathways Are Integrated to Ensure Correct Expression of the CBF and COR Genes during Photoperiodic Growth.

The circadian clock synchronizes a wide range of biological processes with the day/night cycle, and correct circadian regulation is essential for photosynthetic activity and plant growth. We describe here a mechanism where a plastid signal converges with the circadian clock to fine-tune the regulation of nuclear gene expression in Arabidopsis (Arabidopsis thaliana). Diurnal oscillations of tetrapyrrole levels in the chloroplasts contribute to the regulation of the nucleus-encoded transcription factors C-REPEAT BINDING FACTORS (CBFs). The plastid signal triggered by tetrapyrrole accumulation inhibits the activity of cytosolic HEAT SHOCK PROTEIN90 and, as a consequence, the maturation and stability of the clock component ZEITLUPE (ZTL). ZTL negatively regulates the transcription factor LONG HYPOCOTYL5 (HY5) and PSEUDO-RESPONSE REGULATOR5 (PRR5). Thus, low levels of ZTL result in a HY5- and PRR5-mediated repression of CBF3 and PRR5-mediated repression of CBF1 and CBF2 expression. The plastid signal thereby contributes to the rhythm of CBF expression and the downstream COLD RESPONSIVE expression during day/night cycles. These findings provide insight into how plastid signals converge with, and impact upon, the activity of well-defined clock components involved in circadian regulation.

Differential response of Scots pine seedlings to variable intensity and ratio of red and far-red light.

We investigated the response to increasing intensity of red (R) and far-R (FR) light and to a decrease in R:FR ratio in Pinus sylvestris L. (Scots pine) seedling. The results showed that FR high-irradiance response for hypocotyl elongation may be present in Scots pine and that this response is enhanced by increasing light intensity. However, both hypocotyl inhibition and pigment accumulation were more strongly affected by the R light compared with FR light. This is in contrast to previous reports in Arabidopsis thaliana (L.) Heynh. In the angiosperm, A. thaliana R light shows an overall milder effect on inhibition of hypocotyl elongation and on pigment biosynthesis compared with FR suggesting conifers and angiosperms respond very differently to the different light regimes. Scots pine shade avoidance syndrome with longer hypocotyls, shorter cotyledons and lower chlorophyll content in response to shade conditions resembles the response observed in A. thaliana. However, anthocyanin accumulation increased with shade in Scots pine, which again differs from what is known in angiosperms. Overall, the response of seedling development and physiology to R and FR light in Scots pine indicates that the regulatory mechanism for light response may differ between gymnosperms and angiosperms.

Summer mortalities and detection of ostreid herpesvirus microvariant in Pacific oyster Crassostrea gigas in Sweden and Norway.

The Pacific oyster Crassostrea gigas has recently expanded its range in Scandinavia. The expansion is presumably a result of northwards larval drift. Massive settlements were recorded in many areas along the Swedish west coast and southern Norway in 2013 and 2014. After the spawning season in 2014, the temperature of the surface water peaked at 24-26°C. After this period, high and sudden mortalities occurred in a Swedish hatchery and in wild populations along the Swedish west coast and south coast of Norway. Surveys and collected data showed that mortalities mainly occurred during 3 wk in September. All size classes were affected, and affected populations displayed a patchy distribution with heavily affected and unaffected populations in close proximity. Flat oysters Ostrea edulis and blue mussels Mytilus edulis were unaffected. Ostreid herpesvirus (OsHV) was detected in moribund Pacific oyster spat as well as in surviving adults. The virus was identified as OsHV-1 µvar. This is the first detection of this variant in Scandinavia, showing that OsHV-1 µvar is present in areas with recent establishments of Pacific oysters, and where there is no aquaculture of this species.

The CRYPTOCHROME1-dependent response to excess light is mediated through the transcriptional activators ZINC FINGER PROTEIN EXPRESSED IN INFLORESCENCE MERISTEM LIKE1 and ZML2 in Arabidopsis.

Exposure of plants to light intensities that exceed the electron utilization capacity of the chloroplast has a dramatic impact on nuclear gene expression. The photoreceptor Cryptochrome 1 (cry1) is essential to the induction of genes encoding photoprotective components in Arabidopsis thaliana. Bioinformatic analysis of the cry1 regulon revealed the putative cis-element CryR1 (GnTCKAG), and here we demonstrate an interaction between CryR1 and the zinc finger GATA-type transcription factors ZINC FINGER PROTEIN EXPRESSED IN INFLORESCENCE MERISTEM LIKE1 (ZML1) and ZML2. The ZML proteins specifically bind to the CryR1 cis-element as demonstrated in vitro and in vivo, and TCTAG was shown to constitute the core sequence required for ZML2 binding. In addition, ZML2 activated transcription of the yellow fluorescent protein reporter gene driven by the CryR1 cis-element in Arabidopsis leaf protoplasts. T-DNA insertion lines for ZML2 and its homolog ZML1 demonstrated misregulation of several cry1-dependent genes in response to excess light. Furthermore, the zml1 and zml2 T-DNA insertion lines displayed a high irradiance-sensitive phenotype with significant photoinactivation of photosystem II (PSII), indicated by reduced maximum quantum efficiency of PSII, and severe photobleaching. Thus, we identified the ZML2 and ZML1 GATA transcription factors as two essential components of the cry1-mediated photoprotective response.

Cyclin-dependent kinase E1 (CDKE1) provides a cellular switch in plants between growth and stress responses.

Plants must deal effectively with unfavorable growth conditions that necessitate a coordinated response to integrate cellular signals with mitochondrial retrograde signals. A genetic screen was carried out to identify regulators of alternative oxidase (rao mutants), using AOX1a expression as a model system to study retrograde signaling in plants. Two independent rao1 mutant alleles identified CDKE1 as a central nuclear component integrating mitochondrial retrograde signals with energy signals under stress. CDKE1 is also necessary for responses to general cellular stresses, such as H(2)O(2) and cold that act, at least in part, via anterograde pathways, and integrates signals from central energy/stress sensing kinase signal transduction pathways within the nucleus. Together, these results place CDKE1 as a central kinase integrating diverse cellular signals and shed light on a mechanism by which plants can effectively switch between growth and stress responses.

Plastid-to-nucleus communication, signals controlling the running of the plant cell.

The presence of genes encoding organellar proteins in both the nucleus and the organelle necessitates tight coordination of expression by the different genomes, and this has led to the evolution of sophisticated intracellular signaling networks. Organelle-to-nucleus signaling, or retrograde control, coordinates the expression of nuclear genes encoding organellar proteins with the metabolic and developmental state of the organelle. Complex networks of retrograde signals orchestrate major changes in nuclear gene expression and coordinate cellular activities and assist the cell during plant development and stress responses. It has become clear that, even though the chloroplast depends on the nucleus for its function, plastid signals play important roles in an array of different cellular processes vital to the plant. Hence, the chloroplast exerts significant control over the running of the cell. This article is part of a Special Issue entitled: Protein Import and Quality Control in Mitochondria and Plastids.

Pseudomonas syringae infectivity correlates to altered transcript and metabolite levels of Arabidopsis mediator mutants.

Rapid metabolic responses to pathogens are essential for plant survival and depend on numerous transcription factors. Mediator is the major transcriptional co-regulator for integration and transmission of signals from transcriptional regulators to RNA polymerase II. Using four Arabidopsis Mediator mutants, med16, med18, med25 and cdk8, we studied how differences in regulation of their transcript and metabolite levels correlate to their responses to Pseudomonas syringae infection. We found that med16 and cdk8 were susceptible, while med25 showed increased resistance. Glucosinolate, phytoalexin and carbohydrate levels were reduced already before infection in med16 and cdk8, but increased in med25, which also displayed increased benzenoids levels. Early after infection, wild type plants showed reduced glucosinolate and nucleoside levels, but increases in amino acids, benzenoids, oxylipins and the phytoalexin camalexin. The Mediator mutants showed altered levels of these metabolites and in regulation of genes encoding key enzymes for their metabolism. At later stage, mutants displayed defective levels of specific amino acids, carbohydrates, lipids and jasmonates which correlated to their infection response phenotypes. Our results reveal that MED16, MED25 and CDK8 are required for a proper, coordinated transcriptional response of genes which encode enzymes involved in important metabolic pathways for Arabidopsis responses to Pseudomonas syringae infections.

Governance hurdles for expansion of low trophic mariculture production in Sweden.

The study examines the governance of low trophic species mariculture (LTM) using Sweden as a case study. LTM, involving species such as seaweeds and mollusks, offers ecosystem services and nutritious foods. Despite its potential to contribute to blue growth and Sustainable Development Goals, LTM development in the EU and OECD countries has stagnated. A framework for mapping governance elements (institutions, structures, and processes) and analyzing governance objective (effective, equitable, responsive, and robust) was combined with surveys addressed to the private entrepreneurs in the sector. Analysis reveals ineffective institutions due to lack of updated legislation and guidance, resulting in ambiguous interpretations. Governance structures include multiple decision-making bodies without a clear coordination agency. Licensing processes were lengthy and costly for the private entrepreneurs, and the outcomes were uncertain. To support Sweden's blue bioeconomy, LTM governance requires policy integration, clearer direction, coordinated decision-making, and mechanisms for conflict resolution and learning.

Unlocking the secret life of blue mussels: Exploring connectivity in the Skagerrak through biophysical modeling and population genomics.

Knowledge of functional dispersal barriers in the marine environment can be used to inform a wide variety of management actions, such as marine spatial planning, restoration efforts, fisheries regulations, and invasive species management. Locations and causes of dispersal barriers can be studied through various methods, including movement tracking, biophysical modeling, demographic models, and genetics. Combining methods illustrating potential dispersal, such as biophysical modeling, with realized dispersal through, e.g., genetic connectivity estimates, provides particularly useful information for teasing apart potential causes of observed barriers. In this study, we focus on blue mussels (Mytilus edulis) in the Skagerrak-a marginal sea connected to the North Sea in Northern Europe-and combine biophysical models of larval dispersal with genomic data to infer locations and causes of dispersal barriers in the area. Results from both methods agree; patterns of ocean currents are a major structuring factor in the area. We find a complex pattern of source-sink dynamics with several dispersal barriers and show that some areas can be isolated despite an overall high dispersal capability. Finally, we translate our finding into management advice that can be used to sustainably manage this ecologically and economically important species in the future.

Environmental assessment of diets: overview and guidance on indicator choice.

Comprehensive but interpretable assessment of the environmental performance of diets involves choosing a set of appropriate indicators. Current knowledge and data gaps on the origin of dietary foodstuffs restrict use of indicators relying on site-specific information. This Personal View summarises commonly used indicators for assessing the environmental performance of diets, briefly outlines their benefits and drawbacks, and provides recommendations on indicator choices for actors across multiple fields involved in activities that include the environmental assessment of diets. We then provide recommendations on indicator choices for actors across multiple fields involved in activities that use environmental assessments, such as health and nutrition experts, policy makers, decision makers, and private-sector and public-sector sustainability officers. We recommend that environmental assessment of diets should include indicators for at least the five following areas: climate change, biosphere integrity, blue water consumption, novel entities, and impacts on natural resources (especially wild fish stocks), to capture important environmental trade-offs. If more indicators can be handled in the assessment, indicators to capture impacts related to land use quantity and quality and green water consumption should be used. For ambitious assessments, indicators related to biogeochemical flows, stratospheric ozone depletion, and energy use can be added.

Redox-mediated mechanisms regulate DNA binding activity of the G-group of basic region leucine zipper (bZIP) transcription factors in Arabidopsis.

Plant genes that contain the G-box in their promoters are responsive to a variety of environmental stimuli. Bioinformatics analysis of transcriptome data revealed that the G-box element is significantly enriched in promoters of high light-responsive genes. From nuclear extracts of high light-treated Arabidopsis plants, we identified the AtbZIP16 transcription factor as a component binding to the G-box-containing promoter fragment of light-harvesting chlorophyll a/b-binding protein2.4 (LHCB2.4). AtbZIP16 belongs to the G-group of Arabidopsis basic region leucine zipper (bZIP) type transcription factors. Although AtbZIP16 and its close homologues AtbZIP68 and AtGBF1 bind the G-box, they do not bind the mutated half-sites of the G-box palindrome. In addition, AtbZIP16 interacts with AtbZIP68 and AtGBF1 in the yeast two-hybrid system. A conserved Cys residue was shown to be necessary for redox regulation and enhancement of DNA binding activity in all three proteins. Furthermore, transgenic Arabidopsis lines overexpressing the wild type version of bZIP16 and T-DNA insertion mutants for bZIP68 and GBF1 demonstrated impaired regulation of LHCB2.4 expression. Finally, overexpression lines for the mutated Cys variant of bZIP16 provided support for the biological significance of Cys(330) in redox regulation of gene expression. Thus, our results suggest that environmentally induced changes in the redox state regulate the activity of members of the G-group of bZIP transcription factors.

The plastid redox insensitive 2 mutant of Arabidopsis is impaired in PEP activity and high light-dependent plastid redox signalling to the nucleus.

The photosynthetic apparatus is composed of proteins encoded by genes from both the nuclear and the chloroplastic genomes. The activities of the nuclear and chloroplast genomes must therefore be closely coordinated through intracellular signalling. The plastids produce multiple retrograde signals at different times of their development, and in response to changes in the environment. These signals regulate the expression of nuclear-encoded photosynthesis genes to match the current status of the plastids. Using forward genetics we identified PLASTID REDOX INSENSITIVE 2 (PRIN2), a chloroplast component involved in redox-mediated retrograde signalling. The allelic mutants prin2-1 and prin2-2 demonstrated a misregulation of photosynthesis-associated nuclear gene expression in response to excess light, and an inhibition of photosynthetic electron transport. As a consequence of the misregulation of LHCB1.1 and LHCB2.4, the prin2 mutants displayed a high irradiance-sensitive phenotype with significant photoinactivation of photosystem II, indicated by a reduced variable to maximal fluorescence ratio (F(v) /F(m) ). PRIN2 is localized to the nucleoids, and plastid transcriptome analyses demonstrated that PRIN2 is required for full expression of genes transcribed by the plastid-encoded RNA polymerase (PEP). Similarly to the prin2 mutants, the ys1 mutant with impaired PEP activity also demonstrated a misregulation of LHCB1.1 and LHCB2.4 expression in response to excess light, suggesting a direct role for PEP activity in redox-mediated retrograde signalling. Taken together, our results indicate that PRIN2 is part of the PEP machinery, and that the PEP complex responds to photosynthetic electron transport and generates a retrograde signal, enabling the plant to synchronize the expression of photosynthetic genes from both the nuclear and plastidic genomes.