SVALKA-POLYCOMB REPRESSIVE COMPLEX2 Module Controls C-REPEAT BINDING FACTOR3 Induction during Cold Acclimation.

C-REPEAT BINDING FACTORS (CBFs) are highly conserved plant transcription factors that promote cold tolerance. In Arabidopsis (Arabidopsis thaliana), three CBFs (CBF1-3) play a critical role in cold acclimation, and the expression of their corresponding genes is rapidly and transiently induced during this adaptive response. Cold induction of CBFs has been extensively studied and shown to be tightly controlled, yet the molecular mechanisms that restrict the expression of each CBF after their induction during cold acclimation are poorly understood. Here, we present genetic and molecular evidence that the decline in the induction of CBF3 during cold acclimation is epigenetically regulated through the Polycomb Repressive Complex (PRC) 2. We show that this complex promotes the deposition of the repressive mark H3K27me3 at the coding region of CBF3, silencing its expression. Our results indicate that the cold-inducible long noncoding RNA SVALKA is essential for this regulation by recruiting PRC2 to CBF3. These findings unveil a SVALKA-PRC2 regulatory module that ensures the precise timing of CBF3 induction during cold acclimation and the correct development of this adaptive response.

H4 acetylation by the NuA4 complex is required for plastid transcription and chloroplast biogenesis.

Chloroplast biogenesis is crucial in plant development, as it is essential for the transition to autotrophic growth. This process is light-induced and relies on the orchestrated transcription of nuclear and plastid genes, enabling the effective assembly and regulation of the photosynthetic machinery. Here we reveal a new regulation level for this process by showing the involvement of chromatin remodelling in the nuclear control of plastid gene expression for proper chloroplast biogenesis and function. The two Arabidopsis homologues of yeast EPL1 protein, components of the NuA4 histone acetyltransferase complex, are essential for plastid transcription and correct chloroplast development and performance. We show that EPL1 proteins are light-regulated and necessary for concerted expression of nuclear genes encoding most components of chloroplast transcriptional machinery, directly mediating H4K5ac deposition at these loci and promoting the expression of plastid genes required for chloroplast biogenesis. These data unveil a NuA4-mediated mechanism regulating chloroplast biogenesis that links the transcription of nuclear and plastid genomes during chloroplast development.

A MRG-operated chromatin switch at SOC1 attenuates abiotic stress responses during the floral transition.

Plants react to environmental challenges by integrating external cues with endogenous signals to optimize survival and reproductive success. However, the mechanisms underlying this integration remain obscure. While stress conditions are known to impact plant development, how developmental transitions influence responses to adverse conditions has not been addressed. Here, we reveal a molecular mechanism of stress response attenuation during the onset of flowering in Arabidopsis (Arabidopsis thaliana). We show that Arabidopsis MORF-RELATED GENE (MRG) proteins, components of the NuA4 histone acetyltransferase complex that bind trimethylated-lysine 36 in histone H3 (H3K36me3), function as a chromatin switch on the floral integrator SUPPRESSOR OF OVEREXPRESSION OF CONSTANS 1 (SOC1) to coordinate flowering initiation with plant responsiveness to hostile environments. MRG proteins are required to activate SOC1 expression during flowering induction by promoting histone H4 acetylation. In turn, SOC1 represses a broad array of genes that mediate abiotic stress responses. We propose that during the transition from vegetative to reproductive growth, the MRG-SOC1 module constitutes a central hub in a mechanism that tunes down stress responses to enhance the reproductive success and plant fitness at the expense of costly efforts for adaptation to challenging environments.

Insights Into the Function of the NuA4 Complex in Plants.

Chromatin remodeling plays a key role in the establishment and maintenance of gene expression patterns essential for plant development and responses to environmental factors. Post-translational modification of histones, including acetylation, is one of the most relevant chromatin remodeling mechanisms that operate in eukaryotic cells. Histone acetylation is an evolutionarily conserved chromatin signature commonly associated with transcriptional activation. Histone acetylation levels are tightly regulated through the antagonistic activity of histone acetyltransferases (HATs) and histone deacetylases (HDACs). In plants, different families of HATs are present, including the MYST family, which comprises homologs of the catalytic subunit of the Nucleosome Acetyltransferase of H4 (NuA4) complex in yeast. This complex mediates acetylation of histones H4, H2A, and H2A.Z, and is involved in transcriptional regulation, heterochromatin silencing, cell cycle progression, and DNA repair in yeast. In Arabidopsis and, other plant species, homologs for most of the yeast NuA4 subunits are present and although the existence of this complex has not been demonstrated yet, compelling evidence supports the notion that this type of HAT complex functions from mosses to angiosperms. Recent proteomic studies show that several Arabidopsis homologs of NuA4 components, including the assembly platform proteins and the catalytic subunit, are associated in vivo with additional members of this complex suggesting that a NuA4-like HAT complex is present in plants. Furthermore, the functional characterization of some Arabidopsis NuA4 subunits has uncovered the involvement of these proteins in the regulation of different plant biological processes. Interestingly, for most of the mutant plants deficient in subunits of this complex characterized so far, conspicuous defects in flowering time are observed, suggesting a role for NuA4 in the control of this plant developmental program. Moreover, the participation of Arabidopsis NuA4 homologs in other developmental processes, such as gametophyte development, as well as in cell proliferation and stress and hormone responses, has also been reported. In this review, we summarize the current state of knowledge on plant putative NuA4 subunits and discuss the latest progress concerning the function of this chromatin modifying complex.

Gene Regulatory Networks Mediating Cold Acclimation: The CBF Pathway.

Under low nonfreezing temperature conditions, plants from temperate climates undergo physiological and biochemical adjustments that increase their tolerance to freezing temperatures. This response, termed cold acclimation, is largely regulated by changes in gene expression. Molecular and genetic studies have identified a small family of transcription factors, called C-repeat binding factors (CBFs), as key regulators of the transcriptomic rearrangement that leads to cold acclimation. The function of these proteins is tightly controlled, and an inadequate supply of CBF activity may be detrimental to the plant. Accumulated evidence has revealed an extremely intricate network of positive and negative regulators of cold acclimation that coalesce at the level of CBF promoters constituting a central hub where multiple internal and external signals are integrated. Moreover, CBF expression is also controlled at posttranscriptional and posttranslational levels further refining CBF regulation. Recently, natural variation studies in Arabidopsis have demonstrated that mutations resulting in changes in CBF expression have an adaptive value for wild populations. Intriguingly, CBF genes are also present in plant species that do not cold acclimate, which suggest that they may also have additional functions. For instance, CBFs are required for some cold-related abiotic stress responses. In addition, their involvement in plant development deserves further study. Although more studies are necessary to fully harness CBF biotechnological potential, these transcription factors are meant to be key for a rational design of crops with enhanced tolerance to abiotic stress.

The SlCBL10 Calcineurin B-Like Protein Ensures Plant Growth under Salt Stress by Regulating Na+ and Ca2+ Homeostasis.

Characterization of a new tomato (Solanum lycopersicum) T-DNA mutant allowed for the isolation of the CALCINEURIN B-LIKE PROTEIN 10 (SlCBL10) gene whose lack of function was responsible for the severe alterations observed in the shoot apex and reproductive organs under salinity conditions. Physiological studies proved that SlCBL10 gene is required to maintain a proper low Na+/Ca2+ ratio in growing tissues allowing tomato growth under salt stress. Expression analysis of the main responsible genes for Na+ compartmentalization (i.e. Na+/H+ EXCHANGERs, SALT OVERLY SENSITIVE, HIGH-AFFINITY K+ TRANSPORTER 1;2, H+-pyrophosphatase AVP1 [SlAVP1] and V-ATPase [SlVHA-A1]) supported a reduced capacity to accumulate Na+ in Slcbl10 mutant leaves, which resulted in a lower uploading of Na+ from xylem, allowing the toxic ion to reach apex and flowers. Likewise, the tomato CATION EXCHANGER 1 and TWO-PORE CHANNEL 1 (SlTPC1), key genes for Ca2+ fluxes to the vacuole, showed abnormal expression in Slcbl10 plants indicating an impaired Ca2+ release from vacuole. Additionally, complementation assay revealed that SlCBL10 is a true ortholog of the Arabidopsis (Arabidopsis thaliana) CBL10 gene, supporting that the essential function of CBL10 is conserved in Arabidopsis and tomato. Together, the findings obtained in this study provide new insights into the function of SlCBL10 in salt stress tolerance. Thus, it is proposed that SlCBL10 mediates salt tolerance by regulating Na+ and Ca2+ fluxes in the vacuole, cooperating with the vacuolar cation channel SlTPC1 and the two vacuolar H+-pumps, SlAVP1 and SlVHA-A1, which in turn are revealed as potential targets of SlCBL10.

Tomato plants increase their tolerance to low temperature in a chilling acclimation process entailing comprehensive transcriptional and metabolic adjustments.

Low temperature is a major environmental stress that seriously compromises plant development, distribution and productivity. Most crops are from tropical origin and, consequently, chilling sensitive. Interestingly, however, some tropical plants, are able to augment their chilling tolerance when previously exposed to suboptimal growth temperatures. Yet, the molecular and physiological mechanisms underlying this adaptive process, termed chilling acclimation, still remain practically unknown. Here, we demonstrate that tomato plants can develop a chilling acclimation response, which includes comprehensive transcriptomic and metabolic adjustments leading to increased chilling tolerance. More important, our results reveal strong resemblances between this response and cold acclimation, the process whereby plants from temperate regions raise their freezing tolerance after exposure to low, non-freezing temperatures. Both chilling and cold acclimation are regulated by a similar set of transcription factors and hormones, and share common defence mechanisms, including the accumulation of compatible solutes, the mobilization of antioxidant systems and the rearrangement of the photosynthetic machinery. Nonetheless, we have found some important divergences that may account for the freezing sensitivity of tomato plants. The data reported in this manuscript should foster new research into the chilling acclimation response with the aim of improving tomato tolerance to low temperature.

Post-translational regulation of cold acclimation response.

Cold acclimation is an adaptive response whereby plants from temperate regions increase their capacity to tolerate freezing in response to low-nonfreezing temperatures. Numerous studies have unveiled the large transcriptome re-programming that takes place during cold acclimation in diverse species, and a number of proteins have been identified as important regulators of this adaptive response. Post-translational mechanisms regulating the function of proteins involved in cold acclimation have been, however, much less studied. Several components of the signal transduction pathways mediating cold response have been described to be post-translationally modified. These post-translational modifications, including protein phosphorylation and dephosphorylation, ubiquitination, SUMOylation, N-glycosylation and lipid modification, determine key aspects of protein function such as sub-cellular localization, stability, activity or ability to interact with other proteins. Integrating these post-translational mechanisms within the appropriate spatio-temporal context of cold acclimation is essential to develop new crops with improved cold tolerance. Here, we review available evidence regarding the post-translational regulation of cold acclimation, discuss its relevance for the accurate development of this response, and highlight significant missing data.

Potassium transport systems in the moss Physcomitrella patens: pphak1 plants reveal the complexity of potassium uptake.

Potassium uptake is one of the most basic processes of plant physiology. However, a comprehensive description is lacking. At a cellular level fungi have provided a helpful but imperfect plant model, which we aim to improve using Physcomitrella patens. Blast searches in expressed sequence tag databases demonstrated that Physcomitrella expresses the same families of K(+) and Na(+) transport systems as flowering plants. We cloned two inward rectifier channels, PpAKT1-2, and four HAK-type transporters (PpHAK1-4). In both types of transport system, phylogenetic analyses revealed that despite their high sequence conservation they could not be included in Arabidopsis or rice (Oryza sativa) clusters. Both inward rectifier channels and one HAK transporter (PpHAK1) were expressed in yeast. PpAKT1 and activated mutants of PpAKT2 and PpHAK1 showed clear functions that were similar to those of homologous systems of flowering plants. A pphak1 null mutant line of Physcomitrella failed to deplete K(+) below 10 mum. Moreover, in a non-K(+)-limiting medium in which wild-type plants grew only as protonema, pphak1-1 plants produced leafy gametophores and contained 60% more K(+). We found that Physcomitrella takes up K(+) through several systems. PpHAK1 is the dominant system in plants that underwent K(+) starvation for long periods but an as-yet unidentified system, which is non-selective for K(+), Rb(+), and Cs(+), dominates in many other conditions. Finally, we discuss that, similar to PpHAK1, one of the functions of AtHAK5 may be to control cellular K(+) content and that a non-selective as-yet unidentified system also exists in Arabidopsis.

Cloning of the PpNHAD1 transporter of Physcomitrella patens, a chloroplast transporter highly conserved in photosynthetic eukaryotic organisms.

A cDNA that encodes a transporter from the NHAD family was identified in Physcomitrella patens. Computer-based searches using the amino acid sequence of PpNHAD1 revealed that, in addition to being expressed in flowering plants, highly conserved transporters of this family are expressed in red algae, green algae, mosses, liverworts, and photosynthetic stramenopiles, but not in heterotrophic stramenopiles. A chloroplast transit peptide was detected in PpNHAD1 and in most of the related sequences, indicating that PpNHAD1 is a chloroplast transporter. A PpNHAD1-GFP fusion localized to the chloroplast in Physcomitrella protoplasts, and truncation of the N-terminus of the protein dispersed the fluorescence signal outside the chloroplast. PpNHAD1 did not show functional expression in either yeast or bacterial mutants, but truncated proteins with shorter N-termini, PpNHAD1-1 and PpNHAD1-2, could be functionally expressed in bacteria. PpNHAD1-1 alleviated the Li(+) intolerance of a Na(+)-efflux Escherichia coli mutant at acidic pH values. Both PpNHAD1-1 and PpNHAD1-2 reduced the K(+) requirements of a K(+)-influx E. coli mutant more actively at high pH values. PpNHAD1 seems to be an important transporter that mediates ionic homeostasis in chloroplasts from red algae to flowering plants.

HKT1 mediates sodium uniport in roots. Pitfalls in the expression of HKT1 in yeast.

The function of HKT1 in roots is controversial. We tackled this controversy by studying Na+ uptake in barley (Hordeum vulgare) roots, cloning the HvHKT1 gene, and expressing the HvHKT1 cDNA in yeast (Saccharomyces cerevisiae) cells. High-affinity Na+ uptake was not detected in plants growing at high K+ but appeared soon after exposing the plants to a K(+)-free medium. It was a uniport, insensitive to external K+ at the beginning of K+ starvation and inhibitable by K+ several hours later. The expression of HvHKT1 in yeast was Na+ (or K+) uniport, Na(+)-K+ symport, or a mix of both, depending on the construct from which the transporter was expressed. The Na+ uniport function was insensitive to external K+ and mimicked the Na+ uptake carried out by the roots at the beginning of K+ starvation. The K+ uniport function only took place in yeast cells that were completely K+ starved and disappeared when internal K+ increased, which makes it unlikely that HvHKT1 mediates K+ uptake in roots. Mutation of the first in-frame AUG codon of HvHKT1 to CUC changed the uniport function into symport. The expression of the symport from either mutants or constructs keeping the first in-frame AUG took place only in K(+)-starved cells, while the uniport was expressed in all conditions. We discuss here that the symport occurs only in heterologous expression. It is most likely related to the K+ inhibitable Na+ uptake process of roots that heterologous systems fail to reproduce.

Sodium, potassium-atpases in algae and oomycetes.

We have investigated the presence of K(+)-transporting ATPases that belong to the phylogenetic group of animal Na(+),K(+)-ATPases in the Pythium aphanidermatum Stramenopile oomycete, the Porphyra yezoensis red alga, and the Udotea petiolata green alga, by molecular cloning and expression in heterologous systems. PCR amplification and search in EST databases allowed one gene to be identified in each species that could encode ATPases of this type. Phylogenetic analysis of the sequences of these ATPases revealed that they cluster with ATPases of animal origin, and that the algal ATPases are closer to animal ATPases than the oomycete ATPase is. The P. yezoensis and P. aphanidermatum ATPases were functionally expressed in Saccharomyces cerevisiae and Escherichia coli alkali cation transport mutants. The aforementioned cloning and complementary searches in silicio for H(+)- and Na(+),K(+)-ATPases revealed a great diversity of strategies for plasma membrane energization in eukaryotic cells different from typical animal, plant, and fungal cells.