Chloroplast translational regulation uncovers nonessential photosynthesis genes as key players in plant cold acclimation

Plants evolved efficient multifaceted acclimation strategies to cope with low temperatures. Chloroplasts respond to temperature stimuli and participate in temperature sensing and acclimation. However, very little is known about the involvement of chloroplast genes and their expression in plant chilling tolerance. Here we systematically investigated cold acclimation in tobacco seedlings over 2 days of exposure to low temperatures by examining responses in chloroplast genome copy number, transcript accumulation and translation, photosynthesis, cell physiology, and metabolism. Our time-resolved genome-wide investigation of chloroplast gene expression revealed substantial cold-induced translational regulation at both the initiation and elongation levels, in the virtual absence of changes at the transcript level. These cold-triggered dynamics in chloroplast translation are widely distinct from previously described high light-induced effects. Analysis of the gene set responding significantly to the cold stimulus suggested nonessential plastid-encoded subunits of photosynthetic protein complexes as novel players in plant cold acclimation. Functional characterization of one of these cold-responsive chloroplast genes by reverse genetics demonstrated that the encoded protein, the small cytochrome b6f complex subunit PetL, crucially contributes to photosynthetic cold acclimation. Together, our results uncover an important, previously underappreciated role of chloroplast translational regulation in plant cold acclimation

Acclimation in plants - the Green Hub consortium

Acclimation is the capacity to adapt to environmental changes within the lifetime of an individual. This ability allows plants to cope with the continuous variation in ambient conditions to which they are exposed as sessile organisms. Because environmental changes and extremes are becoming even more pronounced due to the current period of climate change, enhancing the efficacy of plant acclimation is a promising strategy for mitigating the consequences of global warming on crop yields. At the cellular level, the chloroplast plays a central role in many acclimation responses, acting both as a sensor of environmental change and as a target of cellular acclimation responses. In this Perspective article, we outline the activities of the Green Hub consortium funded by the German Science Foundation. The main aim of this research collaboration is to understand and strategically modify the cellular networks that mediate plant acclimation to adverse environments, employing Arabidopsis, tobacco (Nicotiana tabacum) and Chlamydomonas as model organisms. These efforts will contribute to 'smart breeding' methods designed to create crop plants with improved acclimation properties. To this end, the model oilseed crop Camelina sativa is being used to test modulators of acclimation for their potential to enhance crop yield under adverse environmental conditions. Here we highlight the current state of research on the role of gene expression, metabolism and signalling in acclimation, with a focus on chloroplast-related processes. In addition, further approaches to uncovering acclimation mechanisms derived from systems and computational biology, as well as adaptive laboratory evolution with photosynthetic microbes, are highlighted

Acclimation in plants – the Green Hub consortium

Acclimation is the capacity to adapt to environmental changes within the lifetime of an individual. This ability allows plants to cope with the continuous variation in ambient conditions to which they are exposed as sessile organisms. Because environmental changes and extremes are becoming even more pronounced due to the current period of climate change, enhancing the efficacy of plant acclimation is a promising strategy for mitigating the consequences of global warming on crop yields. At the cellular level, the chloroplast plays a central role in many acclimation responses, acting both as a sensor of environmental change and as a target of cellular acclimation responses. In this Perspective article, we outline the activities of the Green Hub consortium funded by the German Science Foundation. The main aim of this research collaboration is to understand and strategically modify the cellular networks that mediate plant acclimation to adverse environments, employing Arabidopsis, tobacco (Nicotiana tabacum) and Chlamydomonas as model organisms. These efforts will contribute to ‘smart breeding’ methods designed to create crop plants with improved acclimation properties. To this end, the model oilseed crop Camelina sativa is being used to test modulators of acclimation for their potential to enhance crop yield under adverse environmental conditions. Here we highlight the current state of research on the role of gene expression, metabolism and signalling in acclimation, with a focus on chloroplast‐related processes. In addition, further approaches to uncovering acclimation mechanisms derived from systems and computational biology, as well as adaptive laboratory evolution with photosynthetic microbes, are highlighted.Deutsche Forschungsgemeinschaft http://dx.doi.org/10.13039/501100001659Peer Reviewe

Charakterisierung essentieller Faktoren des Nukleinsäuremetabolismus von Chloroplasten

Die chloroplastidäre Genexpression ist durch charakteristische posttranskriptionelle Ereignisse, wie RNA-Prozessierung, RNA-Stabilität, RNA-Edierung oder RNA-Spleißen gekennzeichnet. Diese Prozesse werden fast ausnahmslos durch kernkodierte Proteine realisiert. PPR-Proteine (Pentatricopeptid repeat) stellen unter diesen kernkodierten Faktoren die größte Proteinfamilie dar. Das plastidäre Protein P67 gehört zur kleinen Untergruppe der PPR-Proteine mit SMR-Domäne (small MutS-related), deren molekulare Funktion im organellären Nukleinsäuremetabolismus bislang unverstanden ist. P67 zeigt eine nahe Verwandtschaft zu GUN1, einem zentralen Bestandteil retrograder Signalwege. Der hier analysierte P67-Knockout in Mais verursacht hellgrüne Phänotypen, eine drastische Reduktion der plastidären ATPase und Keimlingsletalität, was die essentielle Beteiligung von P67 an den Prozessen der Chloroplastenbiogenese und der Expression der plastidär kodierten ATPase-Untereinheiten vermuten lässt. Mögliche Implikationen eines fehlenden Phänotyps von Mutanten des P67-Orthologs aus Arabidopsis thaliana werden diskutiert. Eine Ausnahmestellung unter den Proteinen des chloroplastidären RNA-Metabolismus nimmt der einzige plastidär kodierte RNA-Reifungsfaktor MatK ein. Die genomische Position des matK-Gens im Intron der trnK-UUU ist in allen grünen Landpflanzen konserviert. MatK ist mit bakteriellen Maturasen verwandt, die spezifisch den Spleißprozess ihres Heimatintrons unterstützen. Dagegen deuten genetische und phylogenetische Studien zusätzliche MatK-Funktionen in trans an. In der vorliegenden Arbeit wird die spezifische Interaktion von MatK mit sieben Gruppe-IIA-Intron enthaltenden Transkripten in vivo gezeigt. Darunter befinden sich vier tRNA-Vorläufer (trnK-UUU mit dem matK-Heimatintron sowie trnV-UAC, trnI-GAU, trnA-UGC) und drei proteinkodierende Vorläufertranskripte (rpl2, rps12, atpF). Die Feinkartierung der MatK-Bindung im trnK-Heimatintron zeigt eine Assoziation mit multiplen Regionen. Organelläre Gruppe-II-Introns gelten als Vorläufer der spleißosomalen Introns. Die Assoziation mit multiplen Gruppe-II-Introns macht MatK somit zu einem interessanten Modell für die Evolution der transaktiven Spleißaktivität im Kern. Analysen der Expression von MatK und seinen Zielen deuten auf ein komplexes Muster möglicher regulativer Interaktionen hin.Chloroplast gene expression is characterized by posttranscriptional events including RNA cleavage, RNA stability, RNA editing, and RNA splicing. The underlying processing machinery is almost exclusively encoded in the nucleus. PPR proteins (pentatricopeptide repeat) form the biggest protein family among these factors and are major players of the aforementioned posttranscriptional processes. The plastidial protein P67 is a member of a small subgroup of PPR proteins with SMR domain (small MutS-related). Molecular functions of this protein family in organellar nucleic acid metabolism are yet unknown. P67 is a close relative of GUN1, an essential component of the chloroplast to nucleus retrograde signalling pathway. It is shown here that a P67 knockout in maize causes pale green phenotypes, a dramatic reduction in ATPase levels, and seedling lethality. This indicates an essential role of P67 for chloroplast biogenesis and expression of the plastid encoded ATPase. The finding that mutants of the P67-orthologe in Arabidopsis lack a phenotype is discussed against the background of physiological differences between maize and Arabidopsis. A special case among proteins involved in plastid RNA metabolism is MatK - the only plastid encoded RNA maturation factor. The genomic position of the matK gene in the trnK-UUU intron is conserved throughout autotrophic land plants. MatK is related to bacterial maturases - highly specific splice factors supporting splice processes of their respective home introns. There is, however, indirect genetic and phylogenetic evidence that MatK acts also in trans as a common plastidial splice factor serving various group II introns. This study shows that MatK interacts specifically with seven group IIA introns in vivo. Among them are four tRNA precursor transcripts (trnK-UUU including the matK home intron as well as trnV-UAC, trnI-GAU, trnA-UGC) and three protein-coding precursors (rpl2, rps12, atpF). Fine mapping of MatK binding sites within the trnK home intron uncovers protein RNA interactions with diverse intron regions. Organellar introns have been suggested as evolutionary ancestors of nuclear spliceosomal introns. Consequently, association of MatK with multiple group II intron ligands makes the plastidial maturase an attractive model for an early trans-acting nuclear splice activity. Analyses of the expression of MatK and its targets revealed a complex pattern of possible regulatory interactions

Structure of the actively translating plant 80S ribosome at 2.2 Å resolution

In plant cells, translation occurs in three compartments: the cytosol, the plastids and the mitochondria. While the structures of the (prokaryotic-type) ribosomes in plastids and mitochondria are well characterized, high-resolution structures of the eukaryotic 80S ribosomes in the cytosol have been lacking. Here the structure of translating tobacco (Nicotiana tabacum) 80S ribosomes was solved by cryo-electron microscopy with a global resolution of 2.2 Å. The ribosome structure includes two tRNAs, decoded mRNA and the nascent peptide chain, thus providing insights into the molecular underpinnings of the cytosolic translation process in plants. The map displays conserved and plant-specific rRNA modifications and the positions of numerous ionic cofactors, and it uncovers the role of monovalent ions in the decoding centre. The model of the plant 80S ribosome enables broad phylogenetic comparisons that reveal commonalities and differences in the ribosomes of plants and those of other eukaryotes, thus putting our knowledge about eukaryotic translation on a firmer footing

Fast and global reorganization of the chloroplast protein biogenesis network during heat acclimation

International audienceAbstract Photosynthesis is a central determinant of plant biomass production, but its homeostasis is increasingly challenged by heat. Little is known about the sensitive regulatory principles involved in heat acclimation that underly the biogenesis and repair of chloroplast-encoded core subunits of photosynthetic complexes. Employing time-resolved ribosome and transcript profiling together with selective ribosome proteomics, we systematically deciphered these processes in chloroplasts of Chlamydomonas reinhardtii. We revealed protein biosynthesis and altered translation elongation as central processes for heat acclimation and showed that these principles are conserved between the alga and the flowering plant Nicotiana tabacum. Short-term heat exposure resulted in specific translational repression of chlorophyll a-containing core antenna proteins of photosystems I and II. Furthermore, translocation of ribosome nascent chain complexes to thylakoid membranes was affected, as reflected by the increased accumulation of stromal cpSRP54-bound ribosomes. The successful recovery of synthesizing these proteins under prolonged acclimation of nonlethal heat conditions was associated with specific changes of the co-translational protein interaction network, including increased ribosome association of chlorophyll biogenesis enzymes and acclimation factors responsible for complex assembly. We hypothesize that co-translational cofactor binding and targeting might be bottlenecks under heat but become optimized upon heat acclimation to sustain correct co-translational protein complex assembly

AtRsgA from Arabidopsis thaliana is important for maturation of the small subunit of the chloroplast ribosome

Plastid ribosomes are very similar in structure and function to the ribosomes of their bacterial ancestors. Since ribosome biogenesis is not thermodynamically favorable under biological conditions it requires the activity of many assembly factors. Here we have characterized a homolog of bacterial RsgA in Arabidopsis thaliana and show that it can complement the bacterial homolog. Functional characterization of a strong mutant in Arabidopsis revealed that the protein is essential for plant viability, while a weak mutant produced dwarf, chlorotic plants that incorporated immature pre-16S ribosomal RNA into translating ribosomes. Physiological analysis of the mutant plants revealed smaller, but more numerous, chloroplasts in the mesophyll cells, reduction of chlorophyll a and b, depletion of proplastids from the rib meristem and decreased photosynthetic electron transport rate and efficiency. Comparative RNA sequencing and proteomic analysis of the weak mutant and wild-type plants revealed that various biotic stress-related, transcriptional regulation and post-transcriptional modification pathways were repressed in the mutant. Intriguingly, while nuclear- and chloroplast-encoded photosynthesis-related proteins were less abundant in the mutant, the corresponding transcripts were increased, suggesting an elaborate compensatory mechanism, potentially via differentially active retrograde signaling pathways. To conclude, this study reveals a chloroplast ribosome assembly factor and outlines the transcriptomic and proteomic responses of the compensatory mechanism activated during decreased chloroplast function.status: publishe