Arabidopsis FORGETTER1 mediates stress-induced chromatin memory through nucleosome remodeling.

Plants as sessile organisms can adapt to environmental stress to mitigate its adverse effects. As part of such adaptation they maintain an active memory of heat stress for several days that promotes a more efficient response to recurring stress. We show that this heat stress memory requires the activity of the FORGETTER1 (FGT1) locus, with fgt1 mutants displaying reduced maintenance of heat-induced gene expression. FGT1 encodes the Arabidopsis thaliana orthologue of Strawberry notch (Sno), and the protein globally associates with the promoter regions of actively expressed genes in a heat-dependent fashion. FGT1 interacts with chromatin remodelers of the SWI/SNF and ISWI families, which also display reduced heat stress memory. Genomic targets of the BRM remodeler overlap significantly with FGT1 targets. Accordingly, nucleosome dynamics at loci with altered maintenance of heat-induced expression are affected in fgt1. Together, our results suggest that by modulating nucleosome occupancy, FGT1 mediates stress-induced chromatin memory.DOI:http://dx.doi.org/10.7554/eLife.17061.001

In nature, plants often encounter chronic or recurring stressful conditions. Recent results indicate that plants can remember a past exposure to stress to be better prepared for a future stress incident. However, the molecular basis of this is poorly understood. Here, we report the involvement of chromatin modifications in the maintenance of acquired thermotolerance (heat stress [HS] memory). HS memory is associated with the accumulation of histone H3 lysine 4 di- and trimethylation at memory-related loci. This accumulation outlasts their transcriptional activity and marks them as recently transcriptionally active. High accumulation of H3K4 methylation is associated with hyper-induction of gene expression upon a recurring HS. This transcriptional memory and the sustained accumulation of H3K4 methylation depend on HSFA2, a transcription factor that is required for HS memory, but not initial heat responses. Interestingly, HSFA2 associates with memory-related loci transiently during the early stages following HS. In summary, we show that transcriptional memory after HS is associated with sustained H3K4 hyper-methylation and depends on a hit-and-run transcription factor, thus providing a molecular framework for HS memory.

Respiratory burst oxidase homologue‐dependent H2O2 is essential during heat stress memory in heat sensitive tomato

To withstand high temperatures that would be lethal to a plant in the naïve state, land plants must establish heat stress memory. The acquisition of heat stress tolerance via heat stress memory in algae has only been observed in the red alga 'Bangia' sp. ESS1. In this study, we further evaluated the intrinsic ability of this alga to establish heat stress memory by monitoring hydrogen peroxide (H2O2) production and examining the relationship between heat stress memory and the expression of genes encoding nitrogen transporters, since heat stress generally reduces nitrogen absorption. Next, genes encoding nitrogen transporters were selected from our unpublished transcriptome data of 'Bangia' sp. ESS1. We observed a reduction in H2O2 content when heat stress memory was established in the alga. In addition, six ammonium transporter genes, a single-copy nitrate transporter gene and two urea transporter genes were identified. Two of these nitrogen transporter genes were induced by heat stress but not by heat stress memory, two genes showed heat stress memory-dependent expression, and one gene was induced by both treatments. Heat stress memory therefore differentially regulated the expression of the nitrogen transporter genes by reducing heat stress-inducible gene expression and inducing heat stress memory-dependent gene expression. These findings point to the functional diversity of nitrogen transporter genes, which play different roles under various heat stress conditions. The characteristic effects of heat stress memory on the expression of individual nitrogen transporter genes might represent an indispensable strategy for reducing the threshold of sensitivity to recurrent high-temperature conditions and for maintaining nitrogen absorption under such conditions in 'Bangia' sp. ESS1.

When it suddenly gets extremely hot in summer – or when a growth chamber accidentally overheats – plants get stressed. As heat can cause permanent damage and is sometimes even lethal, plants have several mechanisms to protect themselves. They have a certain basal ability to survive temperatures above the optimum for growth, and when extreme directly preceded by a period of mild heat, they can survive better by acquiring thermotolerance (Yeh et al., 2012). In nature, however, high temperatures often occur repeatedly. To cope with this, plants have evolved an intricate mechanism: after experiencing heat once, they can deal better with high temperatures when the heat recurs (Bäurle, 2016). The idea that plants might be able to remember stressful events to cope better with similar situations later has been around for a while (e.g. Itai and Benzioni, 1976). However, evidence for it did not start to accumulate until the mid-2000s. The idea was rather enigmatic. Plants, non-cognitive organisms without a nervous system, were somehow able to construct a memory of past events. In search of the underlying mechanisms, the focus quickly turned towards the role of chromatin (Chinnusamy and Zhu, 2009). Epigenetic alterations such as histone modifications and DNA methylation can alter gene expression patterns, which can then be stably propagated. This might be convenient for acquiring a molecular memory. Indeed, epigenetic regulation turned out to play a key role in a range of different stress memories, such as for drought, salinity, cold and heat (Kim et al., 2015). The research towards heat-stress memory is fueled by imminent climate change, with more frequent and intense heatwaves. Isabel Bäurle in Potsdam, Germany, is one of the scientists investigating the underlying pathways. Her group is especially interested in the role of epigenetic and chromatin regulation in the adaptation of plants to stress. For example, in the last several years they have uncovered the role of FORGETTER 1 (FGT1), a protein that interacts with chromatin remodelers of the SWI/SNF and ISWI families, that mediates heat-stress memory by modulating nucleosome occupancy (Brzezinka et al., 2016). In this issue of The Plant Journal, her team identified two genes that are crucial for heat-stress memory. Surprisingly, these genes encode a protein phosphatase and a phospholipase, and are not directly involved in epigenetic regulation. The protein phosphatase was identified in a mutagenesis screen and designated as FORGETTER 2 (FGT2). When the authors performed heat-stress experiments with the mutant plants, they observed that they were specifically defective in heat-stress memory, but not in the initial acquisitison of thermotolerance (Figure ). Protein phosphatases are enzymes that remove phosphate groups from proteins. By altering the phosphorylation status, they control whether a protein is active or not. The class of type-2C protein phosphatases to which FGT2 belongs is known to play prominent roles in plant stress responses (Singh et al., 2015). Until now, however, they have never been implicated in heat-stress memory. When the authors isolated interaction partners of FGT2 they found phospholipase Dα2 (PLDα2). Mutant analyses confirmed that the gene encoding this enzyme was also critical for heat-stress memory. Moreover, the fgt2 pldα2 double mutant reacted in the same way to heat stress as the fgt2 single mutant, suggesting that the two genes act in the same genetic pathway. Phospholipases of the D class hydrolyze phospholipids and release phosphatidic acid. As biological membranes are mainly made up of phospholipids, the action of phospholipases plays an important role in their stability and structure. Furthermore, both phospholipids and phosphatidic acid are known to function as signaling molecules. Bäurle and her team showed that PLDα2 resided in the cytoplasm, whereas FGT2 attaches to the plasma membrane, probably through lipid anchoring. They suggest a model in which FGT2 and PLDα2 interact at the plasma membrane–cytosol interface. FGT2 might be responsible for the dephosphorylation of PLDα2, thereby controlling its activity (Figure 1). The phospholipase activity of PLDα2 might alter the lipid composition of the cell membrane or produce signaling molecules that induce heat-stress memory. The exact downstream pathway that eventually leads to this ‘memory’ of heat stress remains enigmatic. It is possible that chromatin remodeling is taking place further downstream, but it might as well be an independent mechanism. As Bäurle’s team is particularly focused on the role of epigenetic regulation in plant stress, discovering that FGT2 and PLDα2 are involved is remarkable. Bäurle calls it the beauty of forward genetics; it can yield new and unexpected findings that do not fit in your model (yet). It also underlines the importance of using unbiased approaches. She hopes that their new findings will inspire the work of others studying stress responses, protein phosphatases, phospholipases, membrane dynamics and stress memory, in foreseeable and unforeseeable ways. All good reasons to heat up your growth chambers.

Many environmental conditions fluctuate and organisms need to respond effectively. This is especially true for temperature cues that can change in minutes to seasons and often follow a diurnal rhythm. Plants cannot migrate and most cannot regulate their temperature. Therefore, a broad array of responses have evolved to deal with temperature cues from freezing to heat stress. A particular response to mildly elevated temperatures is called thermomorphogenesis, a suite of morphological adaptations that includes thermonasty, formation of thin leaves and elongation growth of petioles and hypocotyl. Thermomorphogenesis allows for optimal performance in suboptimal temperature conditions by enhancing the cooling capacity. When temperatures rise further, heat stress tolerance mechanisms can be induced that enable the plant to survive the stressful temperature, which typically comprises cellular protection mechanisms and memory thereof. Induction of thermomorphogenesis, heat stress tolerance and stress memory depend on gene expression regulation, governed by diverse epigenetic processes. In this Tansley review we update on the current knowledge of epigenetic regulation of heat stress tolerance and elevated temperature signalling and response, with a focus on thermomorphogenesis regulation and heat stress memory. In particular we highlight the emerging role of H3K4 methylation marks in diverse temperature signalling pathways.

Exposure to moderate heat stress (HS) primes plants to better withstand future exposure to more severe HS conditions. In Arabidopsis (Arabidopsis thaliana), the primed state is maintained for several days, referred to as HS memory. HEAT SHOCK TRANSCRIPTION FACTOR A2 (HSFA2) and HSFA3 promote this HS memory by jointly mediating transcriptional memory in a subset of HS-inducible genes. Why only 2 of the 21 HSFs in Arabidopsis function specifically in HS memory is unknown. Here, we investigated this question through a promoter and domain swap analysis between HSFA2 and HSFA1D, a regulator of the acute HS response (HSR), aiming to uncover the requirements for HS memory HSFs. We determined that conferring the expression pattern of HSFA2 to HSFA1D is not sufficient for restoring HS memory. In chimeric proteins, the C-terminal regions and the DNA-binding domains (DBDs) of the 2 HSFs are interchangeable, while the presence of the HSFA1D repression domain abrogates HS memory. Interestingly, the oligomerization domain (OD) of HSFA1D cannot replace its HSFA2 counterpart in mediating physiological HS memory, suggesting a role for distinct interacting HSF complexes. The OD-exchanged chimeric HSF hyper-induced memory genes after a single HS, suggesting an altered response of the protein to HS. In summary, our study provides insights into the roles of individual HSF domains in specifying functions in HS memory or the acute HSR, thus providing avenues for tailoring HSFs to the demands of a changing climate.

Moderate and temporary heat stresses (HS) prime plants to tolerate, and survive, a subsequent severe HS. Such acquired thermotolerance can be maintained for several days under normal growth conditions, and create a HS memory. We recently demonstrated that plastid-localized small heat shock protein HSP21 is a key component of HS memory in Arabidopsis thaliana. A sustained high abundance of HSP21 during the HS recovery phase extends HS memory. The level of HSP21 is negatively controlled by plastid-localized metalloprotease FtsH6 during HS recovery. Here, we demonstrate that autophagy, a cellular recycling mechanism, exerts additional control over HSP21 degradation. Genetic and chemical disruption of both, metalloprotease activity and autophagy trigger superior HSP21 accumulation, thereby improving memory. Furthermore, we provide evidence that autophagy cargo receptor ATG8-INTERACTING PROTEIN1 (ATI1) is associated with HS memory. ATI1 bodies colocalize with both autophagosomes and HSP21, and their abundance and transport to the vacuole increase during HS recovery. Together, our results provide new insights into the control module for the regulation of HS memory, in which two distinct protein degradation pathways act in concert to degrade HSP21, thereby enabling cells to recover from the HS effect at the cost of reducing the HS memory.

Climate change requires optimizing stress responses in crops. Priming and memory of heat stress (HS) allow plants to improve their tolerance against high temperatures. Here, we investigate HS memory in cultivated barley (Hordeum vulgare) to assess whether the mechanisms underlying priming by and memory of HS are conserved in monocots. Mutation of barley HvHSFA2 and HvHSFA3 reduced HS memory. This correlated with altered transcriptional responses of heat-induced genes in the mutants after recurrent HS. Conversely, overexpression of HvHSFA2 increases HS tolerance with no penalty on productivity. While the biological role of HSFA2 and HSFA3 is conserved, their mechanistic functions appear to have diverged; both factors are globally required to boost induction of HS-responsive genes after recurrent HS. In summary, barley HS memory depends on the highly conserved HvHSFA2 and HvHSFA3, however, the underlying transcriptional wiring is different. Our findings provide a tangible route to improve HS tolerance in temperate cereals.

Plants can mitigate environmental stress conditions through acclimation. In the case of fluctuating stress conditions such as high temperatures, maintaining a stress memory enables a more efficient response upon recurring stress. In a genetic screen for Arabidopsis thaliana mutants impaired in the memory of heat stress (HS) we have isolated the FORGETTER2 (FGT2) gene, which encodes a type 2C protein phosphatase (PP2C) of the D-clade. Fgt2 mutants acquire thermotolerance normally; however, they are defective in the memory of HS. FGT2 interacts with phospholipase D α2 (PLDα2), which is involved in the metabolism of membrane phospholipids and is also required for HS memory. In summary, we have uncovered a previously unknown component of HS memory and identified the FGT2 protein phosphatase and PLDα2 as crucial players, suggesting that phosphatidic acid-dependent signaling or membrane composition dynamics underlie HS memory.

BackgroundThe escalating impacts of global warming intensify the detrimental effects of heat stress on crop growth and yield. Among the earliest and most vulnerable sites of damage is Photosystem II (PSII). Plants exposed to recurring high temperatures develop heat stress memory, a phenomenon that enables them to retain information from previous stress events to better cope with subsequent one. Understanding the components and regulatory networks associated with heat stress memory is crucial for the development of heat-resistant crops.ResultsPhysiological assays revealed that heat priming (HP) enabled tall fescue to possess higher Photosystem II photochemical activity when subjected to trigger stress. To investigate the underlying mechanisms of heat stress memory, we performed comparative proteomic analyses on tall fescue leaves at S0 (control), R4 (primed), and S5 (triggering), using an integrated approach of Tandem Mass Tag (TMT) labeling and Liquid Chromatography-Mass Spectrometry. A total of 3,851 proteins were detected, with quantitative information available for 3,835 proteins. Among these, we identified 1,423 differentially abundant proteins (DAPs), including 526 proteins that were classified as Heat Stress Memory Proteins (HSMPs). GO and KEGG enrichment analyses revealed that the HSMPs were primarily associated with the “autophagy” in R4 and with “PSII repair”, “HSP binding”, and “peptidase activity” in S5. Notably, we identified 7 chloroplast-localized HSMPs (HSP21, DJC77, EGY3, LHCA4, LQY1, PSBR and DEGP8, R4/S0 > 1.2, S5/S0 > 1.2), which were considered to be effectors linked to PSII heat stress memory, predominantly in cluster 4. Protein-protein interaction (PPI) analysis indicated that the ubiquitin-proteasome system, with key nodes at UPL3, RAD23b, and UCH3, might play a role in the selective retention of memory effectors in the R4 stage. Furthermore, we conducted RT-qPCR validation on 12 genes, and the results showed that in comparison to the S5 stage, the R4 stage exhibited reduced consistency between transcript and protein levels, providing additional evidence for post-transcriptional regulation in R4.ConclusionsThese findings provide valuable insights into the establishment of heat stress memory under recurring high-temperature episodes and offer a conceptual framework for breeding thermotolerant crops with improved PSII functionality.

As sessile life forms, plants are repeatedly confronted with adverse environmental conditions, which can impair development, growth, and reproduction. During evolution, plants have established mechanisms to orchestrate the delicate balance between growth and stress tolerance, to reset cellular biochemistry once stress vanishes, or to keep a molecular memory, which enables survival of a harsher stress that may arise later. Although there are several examples of memory in diverse plants species, the molecular machinery underlying the formation, duration, and resetting of stress memories is largely unknown so far. We report here that autophagy, a central self-degradative process, assists in resetting cellular memory of heat stress (HS) in Arabidopsis thaliana. Autophagy is induced by thermopriming (moderate HS) and, intriguingly, remains high long after stress termination. We demonstrate that autophagy mediates the specific degradation of heat shock proteins at later stages of the thermorecovery phase leading to the accumulation of protein aggregates after the second HS and a compromised heat tolerance. Autophagy mutants retain heat shock proteins longer than wild type and concomitantly display improved thermomemory. Our findings reveal a novel regulatory mechanism for HS memory in plants.

As sessile organisms, plants are constantly exposed to a wide spectrum of stress conditions such as high temperature, which causes protein misfolding. Misfolded proteins are highly toxic and must be efficiently removed to reduce cellular proteotoxic stress if restoration of native conformations is unsuccessful. Although selective autophagy is known to function in protein quality control by targeting degradation of misfolded and potentially toxic proteins, its role and regulation in heat stress responses have not been analyzed in crop plants. In the present study, we found that heat stress induced expression of autophagy-related (ATG) genes and accumulation of autophagosomes in tomato plants. Virus-induced gene silencing (VIGS) of tomato ATG5 and ATG7 genes resulted in increased sensitivity of tomato plants to heat stress based on both increased development of heat stress symptoms and compromised photosynthetic parameters of heat-stressed leaf tissues. Silencing of tomato homologs for the selective autophagy receptor NBR1, which targets ubiquitinated protein aggregates, also compromised tomato heat tolerance. To better understand the regulation of heat-induced autophagy, we found that silencing of tomato ATG5, ATG7, or NBR1 compromised heat-induced expression of not only the targeted genes but also other autophagy-related genes. Furthermore, we identified two tomato genes encoding proteins highly homologous to Arabidopsis WRKY33 transcription factor, which has been previously shown to interact physically with an autophagy protein. Silencing of tomato WRKY33 genes compromised tomato heat tolerance and reduced heat-induced ATG gene expression and autophagosome accumulation. Based on these results, we propose that heat-induced autophagy in tomato is subject to cooperative regulation by both WRKY33 and ATG proteins and plays a critical role in tomato heat tolerance, mostly likely through selective removal of heat-induced protein aggregates.

Plants encounter biotic and abiotic stresses many times during their life cycle and this limits their productivity. Moderate heat stress (HS) primes a plant to survive higher temperatures that are lethal in the naïve state. Once temperature stress subsides, the memory of the priming event is actively retained for several days preparing the plant to better cope with recurring HS. Recently, chromatin regulation at different levels has been implicated in HS memory. Here, we report that the chromatin protein BRUSHY1 (BRU1)/TONSOKU/MGOUN3 plays a role in the HS memory in Arabidopsis thaliana. BRU1 is also involved in transcriptional gene silencing and DNA damage repair. This corresponds with the functions of its mammalian orthologue TONSOKU-LIKE/NFΚBIL2. During HS memory, BRU1 is required to maintain sustained induction of HS memory-associated genes, whereas it is dispensable for the acquisition of thermotolerance. In summary, we report that BRU1 is required for HS memory in A.thaliana, and propose a model where BRU1 mediates the epigenetic inheritance of chromatin states across DNA replication and cell division.

As one of the most important food and feed crops worldwide, maize suffers much more tremendous damages under heat stress compared to other plants, which seriously inhibits plant growth and reduces productivity. To mitigate the heat-induced damages and adapt to high temperature environment, plants have evolved a series of molecular mechanisms to sense, respond and adapt high temperatures and heat stress. In this review, we summarized recent advances in molecular regulations underlying high temperature sensing, heat stress response and memory in maize, especially focusing on several important pathways and signals in high temperature sensing, and the complex transcriptional regulation of ZmHSFs (Heat Shock Factors) in heat stress response. In addition, we highlighted interactions between ZmHSFs and several epigenetic regulation factors in coordinately regulating heat stress response and memory. Finally, we laid out strategies to systematically elucidate the regulatory network of maize heat stress response, and discussed approaches for breeding future heat-tolerance maize.