Functional dissection of phytochrome A in plants.

Plants lack behavioral responses to avoid dramatic environmental changes associated with the annual seasons. For survival, they have evolved complex sensory systems to sense fluctuations in light and optimize their architecture in response to changes in these cues. Phytochrome A (phyA) was initially identified as a photoreceptor that senses far-red light signals. It was then identified as playing a central role in promoting hypocotyl growth, fiber development, and flowering time in a variety of plants including Arabidopsis, rice, soybean and cotton. Under dark conditions, phyA is present in the cytoplasm in the physiologically inactive (Pr) form. Far-red light signals induce the transformation of Pr into the physiologically active (Pfr) form, after which Pfr-phyA is recognized by FAR-RED ELONGATED HYPOCOTYL 1 (FHY1) and FHY1-LIKE (FHL) and translocated to the nucleus, initiating a series of signaling cascades. The current review comprehensively summarizes recent advances in understanding the function of phyA in plants, including phyA-mediated shade avoidance and flowering time. Remaining issues and possible directions for future research on phyA are also discussed.

Applications of fluorescence spectroscopy in the investigation of plant phytochrome invivo.

Fluorometry is an effective research tool in biology and medicine; it is widely used in the study of the photosynthetic pigment apparatus in vivo. This method can be applied to the key plant photoreceptor phytochrome (phy). The fluorescence of phytochrome in plants was recorded for the first time in the group of the author, and a spectrofluorometric technique for its in vivo study was developed. The photophysical and photochemical properties of the pigment were described, and the photoreceptor was shown to be present in plants as two phenomenological types-active (at cryogenic temperatures) and water-soluble (Pr') and inactive and amphiphilic (Pr″). The scheme of the photoreaction explaining their photochemical distinctions was proposed. Phytochrome A was shown to comprise both types (phyA' and phyA″), whereas phytochrome B was only the second type. For phyA', distinct conformers have been detected. phyA' and phyA″ differ by the N-terminus of the molecule, possibly by serine phosphorylation. They mediate, respectively, the very low fluence and high irradiance photoresponses. Light, internal factors (kinase/phosphatase balance, pH), and hormones (jasmonate) were shown to affect the content and functions of the two phyA pools. All this points to the effectiveness of the developed method for invivo investigations of the phytochrome system. The data obtained can be applied in practical terms in agrobiology and light culture, as well as in the use of phytochrome as a new nanotool and a fluorescent probe.

PHYTOCHROME A controls the DNA damage response and cell death tolerance within the Arabidopsis root meristem.

The DNA damage response avoids mutations into dividing cells. Here, we analysed the role of photoreceptors on the restriction of root growth imposed by genotoxic agents and its relationship with cell viability and performance of meristems. Comparison of root growth of Arabidopsis WT, phyA-211, phyB-9, and phyA-211phyB-9 double mutants unveiled a critical role for phytochrome A (PhyA) in protecting roots from genotoxic stress, regeneration and cell replenishment in the meristematic zone. PhyA was located on primary root tips, where it influences genes related to the repair of DNA, including ERF115 and RAD51. Interestingly, phyA-211 mutants treated with zeocin failed to induce the expression of the repressor of cell cycle MYB3R3, which correlated with expression of the mitotic cyclin CycB1, suggesting that PhyA is required for safeguarding the DNA integrity during cell division. Moreover, the growth of the primary roots of PhyA downstream component HY5 and root growth analyses in darkness suggest that cell viability and DNA damage responses within root meristems may act independently from light and photomorphogenesis. These data support novel roles for PhyA as a key player for stem cell niche maintenance and DNA damage responses, which are critical for proper root growth.

Low temperature-mediated repression and far-red light-mediated induction determine morning FLOWERING LOCUS T expression levels.

In order to flower in the appropriate season, plants monitor light and temperature changes and alter downstream pathways that regulate florigen genes such as Arabidopsis (Arabidopsis thaliana) FLOWERING LOCUS T (FT). In Arabidopsis, FT messenger RNA levels peak in the morning and evening under natural long-day conditions (LDs). However, the regulatory mechanisms governing morning FT induction remain poorly understood. The morning FT peak is absent in typical laboratory LDs characterized by high red:far-red light (R:FR) ratios and constant temperatures. Here, we demonstrate that ZEITLUPE (ZTL) interacts with the FT repressors TARGET OF EATs (TOEs), thereby repressing morning FT expression in natural environments. Under LDs with simulated sunlight (R:FR = 1.0) and daily temperature cycles, which are natural LD-mimicking environmental conditions, FT transcript levels in the ztl mutant were high specifically in the morning, a pattern that was mirrored in the toe1 toe2 double mutant. Low night-to-morning temperatures increased the inhibitory effect of ZTL on morning FT expression by increasing ZTL protein levels early in the morning. Far-red light counteracted ZTL activity by decreasing its abundance (possibly via phytochrome A (phyA)) while increasing GIGANTEA (GI) levels and negatively affecting the formation of the ZTL-GI complex in the morning. Therefore, the phyA-mediated high-irradiance response and GI play pivotal roles in morning FT induction. Our findings suggest that the delicate balance between low temperature-mediated ZTL activity and the far-red light-mediated functions of phyA and GI offers plants flexibility in fine-tuning their flowering time by controlling FT expression in the morning.

Negative regulation of tobacco cold stress tolerance by NtPhyA.

Cold stress is a non-biological stressor that adversely affects tobacco yield and leaf quality. Plant photoreceptor proteins, which function as dual light-temperature sensors, play a vital role in temperature changes, making them crucial for responses to non-biological stressors. However, the regulatory mechanisms of PhyA in tobacco remain poorly understood. Therefore, in this study, we aimed to clone the NtPhyA gene from tobacco and generate overexpression (OE-NtPhyA) and mutant (KO-NtPhyA) constructs of NtPhyA. By assessing the physiological and biochemical responses of the mutants under cold stress and performing transcriptome sequencing, we determined the signalling mechanism of NtPhyA under cold stress. Comparative analysis with wild-type (WT) NtPhyA revealed that KO-NtPhyA exhibited increased seed germination rates and reduced wilting under cold stress. In additional, the degree of damage to leaf cells, cell membranes, and stomatal structures was mitigated, and the levels of reactive oxygen species (ROS) were significantly decreased. Antioxidant enzyme activity, net photosynthetic rate, and Fv/Fm were significantly enhanced in KO-NtPhyA, whereas the opposite effects were observed in OE-NtPhyA. These findings indicate that KO-NtPhyA augments tobacco tolerance to cold stress, implying a negative regulatory role of NtPhyA in tobacco during cold stress. Transcriptome analysis revealed that NtPhyA governs the expression of a cascade of genes involved in the response to oxygen-containing compounds, hydrogen peroxide (H2O2), ROS, temperature stimuli, photosystem II oxygen-evolving complex assembly, water channel activity, calcium channel activity, and carbohydrate transport. Collectively, our findings indicate that NtPhyA activates downstream gene expression to enhance the resilience of tobacco to cold stress.

Genome-wide analysis of the GRAS gene family in Liriodendron chinense reveals the putative function in abiotic stress and plant development.

Introduction: GRAS genes encode plant-specific transcription factors that play essential roles in plant growth and development. However, the members and the function of the GRAS gene family have not been reported in Liriodendron chinense. L. chinense, a tree species in the Magnolia family that produces excellent timber for daily life and industry. In addition, it is a good relict species for plant evolution research. Methods: Therefore, we conducted a genome-wide study of the LcGRAS gene family and identified 49 LcGRAS genes in L. chinense. Results: We found that LcGRAS could be divided into 13 sub-groups, among which there is a unique branch named HAM-t. We carried out RNA sequencing analysis of the somatic embryos from L. chinense and found that LcGRAS genes are mainly expressed after heart-stage embryo development, suggesting that LcGRAS may have a function during somatic embryogenesis. We also investigated whether GRAS genes are responsive to stress by carrying out RNA sequencing (RNA-seq) analysis, and we found that the genes in the PAT subfamily were activated upon stress treatment, suggesting that these genes may help plants survive stressful environments. We found that PIF was downregulated and COR was upregulated after the transient overexpression of PATs, suggesting that PAT may be upstream regulators of cold stress. Discussion: Collectively, LcGRAS genes are conserved and play essential roles in plant development and adaptation to abiotic stress.

Phylogeny of Alisma (Alismataceae) revisited: implications for polyploid evolution and species delimitation.

Alisma L. is a genus of aquatic and wetland plants belonging to family Alismataceae. At present, it is thought to contain ten species. Variation in ploidy level is known in the genus, with diploids, tetraploids and hexaploids recorded. Previous molecular phylogenetic studies of Alisma have generated a robust backbone that reveals important aspects of the evolutionary history of this cosmopolitan genus, yet questions remain unresolved about the formation of the polyploid taxa and the taxonomy of one particularly challenging, widely distributed species complex. Here we directly sequenced, or cloned and sequenced, nuclear DNA (nrITS and phyA) and chloroplast DNA (matK, ndhF, psbA-trnH and rbcL) of multiple samples of six putative species and two varieties, and conducted molecular phylogenetic analyses. Alisma canaliculatum and its two varieties known in East Asia and A. rariflorum endemic to Japan possess closely related but heterogeneous genomes, strongly indicating that the two species were generated from two diploid progenitors, and are possibly siblings of one another. This evolutionary event may have occurred in Japan. Alisma canaliculatum var. canaliculatum is segregated into two types, each of which are geographically slightly differentiated in Japan. We reconstructed a single phylogeny based on the multi-locus data using Homologizer and then applied species delimitation analysis (STACEY). This allowed us to discern A. orientale as apparently endemic to the Southeast Asian Massif and distinct from the widespread A. plantago-aquatica. The former species was most likely formed through parapatric speciation at the southern edge of the distribution of the latter.

Two Distinct Molecular Types of Phytochrome A in Plants: Evidence of Existence and Implications for Functioning.

Phytochrome (phy) system in plants comprising a small number of phytochromes with phyA and phyB as major ones is responsible for acquiring light information in the red-far-red region of the solar spectrum. It provides optimal strategy for plant development under changing light conditions throughout all its life cycle beginning from seed germination and seedling establishment to fruiting and plant senescence. The phyA was shown to participate in the regulation of this cycle which is especially evident at its early stages. It mediates three modes of reactions-the very low and low fluence responses (VLFR and LFR) and the high irradiance responses (HIR). The phyA is the sole light receptor in the far-red spectral region responsible for plant's survival under a dense plant canopy where light is enriched with the far-red component. Its appearance is believed to be one of the main factors of plants' successful evolution. So far, it is widely accepted that one molecular phyA species is responsible for its complex functional manifestations. In this review, the evidence of the existence of two distinct phyA types-major, light-labile and soluble phyA' and minor, relatively light-stable and amphiphilic phyA″-is presented as what may account for the diverse modes of phyA action.

GmEID1 modulates light signaling through the Evening Complex to control flowering time and yield in soybean.

Soybean (Glycine max) morphogenesis and flowering time are accurately regulated by photoperiod, which determine the yield potential and limit soybean cultivars to a narrow latitudinal range. The E3 and E4 genes, which encode phytochrome A photoreceptors in soybean, promote the expression of the legume-specific flowering repressor E1 to delay floral transition under long-day (LD) conditions. However, the underlying molecular mechanism remains unclear. Here, we show that the diurnal expression pattern of GmEID1 is opposite to that of E1 and targeted mutations in the GmEID1 gene delay soybean flowering regardless of daylength. GmEID1 interacts with J, a key component of circadian Evening Complex (EC), to inhibit E1 transcription. Photoactivated E3/E4 interacts with GmEID1 to inhibit GmEID1-J interaction, promoting J degradation resulting in a negative correlation between daylength and the level of J protein. Notably, targeted mutations in GmEID1 improved soybean adaptability by enhancing yield per plant up to 55.3% compared to WT in field trials performed in a broad latitudinal span of more than 24°. Together, this study reveals a unique mechanism in which E3/E4-GmEID1-EC module controls flowering time and provides an effective strategy to improve soybean adaptability and production for molecular breeding.

Analysis of Genome Structure and Its Variations in Potato Cultivars Grown in Russia.

Solanum tuberosum L. (common potato) is one of the most important crops produced almost all over the world. Genomic sequences of potato opens the way for studying the molecular variations related to diversification. We performed a reconstruction of genomic sequences for 15 tetraploid potato cultivars grown in Russia using short reads. Protein-coding genes were identified; conserved and variable parts of pan-genome and the repertoire of the NBS-LRR genes were characterized. For comparison, we used additional genomic sequences for twelve South American potato accessions, performed analysis of genetic diversity, and identified the copy number variations (CNVs) in two these groups of potato. Genomes of Russian potato cultivars were more homogeneous by CNV characteristics and have smaller maximum deletion size in comparison with South American ones. Genes with different CNV occurrences in two these groups of potato accessions were identified. We revealed genes of immune/abiotic stress response, transport and five genes related to tuberization and photoperiod control among them. Four genes related to tuberization and photoperiod were investigated in potatoes previously (phytochrome A among them). A novel gene, homologous to the poly(ADP-ribose) glycohydrolase (PARG) of Arabidopsis, was identified that may be involved in circadian rhythm control and contribute to the acclimatization processes of Russian potato cultivars.

Regulation of Plant Photoresponses by Protein Kinase Activity of Phytochrome A.

Extensive research has been conducted for decades to elucidate the molecular and regulatory mechanisms for phytochrome-mediated light signaling in plants. As a result, tens of downstream signaling components that physically interact with phytochromes are identified, among which negative transcription factors for photomorphogenesis, PHYTOCHROME-INTERACTING FACTORs (PIFs), are well known to be regulated by phytochromes. In addition, phytochromes are also shown to inactivate an important E3 ligase complex consisting of CONSTITUTIVELY PHOTOMORPHOGENIC 1 (COP1) and SUPPRESSORs OF phyA-105 (SPAs). This inactivation induces the accumulation of positive transcription factors for plant photomorphogenesis, such as ELONGATED HYPOCOTYL 5 (HY5). Although many downstream components of phytochrome signaling have been studied thus far, it is not fully elucidated which intrinsic activity of phytochromes is necessary for the regulation of these components. It should be noted that phytochromes are autophosphorylating protein kinases. Recently, the protein kinase activity of phytochrome A (phyA) has shown to be important for its function in plant light signaling using Avena sativa phyA mutants with reduced or increased kinase activity. In this review, we highlight the function of phyA as a protein kinase to explain the regulation of plant photoresponses by phyA.

Stimulation of Tomato Drought Tolerance by PHYTOCHROME A and B1B2 Mutations.

Drought stress is a severe environmental issue that threatens agriculture at a large scale. PHYTOCHROMES (PHYs) are important photoreceptors in plants that control plant growth and development and are involved in plant stress response. The aim of this study was to identify the role of PHYs in the tomato cv. 'Moneymaker' under drought conditions. The tomato genome contains five PHYs, among which mutant lines in tomato PHYA and PHYB (B1 and B2) were used. Compared to the WT, phyA and phyB1B2 mutants exhibited drought tolerance and showed inhibition of electrolyte leakage and malondialdehyde accumulation, indicating decreased membrane damage in the leaves. Both phy mutants also inhibited oxidative damage by enhancing the expression of reactive oxygen species (ROS) scavenger genes, inhibiting hydrogen peroxide (H2O2) accumulation, and enhancing the percentage of antioxidant activities via DPPH test. Moreover, expression levels of several aquaporins were significantly higher in phyA and phyB1B2, and the relative water content (RWC) in leaves was higher than the RWC in the WT under drought stress, suggesting the enhancement of hydration status in the phy mutants. Therefore, inhibition of oxidative damage in phyA and phyB1B2 mutants may mitigate the harmful effects of drought by preventing membrane damage and conserving the plant hydrostatus.

Novel and multifaceted regulations of photoperiodic flowering by phytochrome A in soybean.

Photoperiod is an important environmental cue. Plants can distinguish the seasons and flower at the right time through sensing the photoperiod. Soybean is a sensitive short-day crop, and the timing of flowering varies greatly at different latitudes, thus affecting yields. Soybean cultivars in high latitudes adapt to the long day by the impairment of two phytochrome genes, PHYA3 and PHYA2, and the legume-specific flowering suppressor, E1. However, the regulating mechanism underlying phyA and E1 in soybean remains largely unknown. Here, we classified the regulation of the E1 family by phyA2 and phyA3 at the transcriptional and posttranscriptional levels, revealing that phyA2 and phyA3 regulate E1 by directly binding to LUX proteins, the critical component of the evening complex, to regulate the stability of LUX proteins. In addition, phyA2 and phyA3 can also directly associate with E1 and its homologs to stabilize the E1 proteins. Therefore, phyA homologs control the core flowering suppressor E1 at both the transcriptional and posttranscriptional levels, to double ensure the E1 activity. Thus, our results disclose a photoperiod flowering mechanism in plants by which the phytochrome A regulates LUX and E1 activity.

Phytochrome A in plants comprises two structurally and functionally distinct populations - water-soluble phyA' and amphiphilic phyA″.

Photoreceptor phytochrome A (phyA) plays a key role both in the individual development and in the evolution of higher plants. It acts in three distinct modes - far-red light-induced very low fluence responses (VLFRs), high irradiance responses (HIRs), and red/far-red-reversible low fluence responses (LFRs). Signal transduction from phyA includes its transportation from the cytoplasm into the nucleus and activation of light-responsive genes there. It is also active in the cytoplasm. Two types of phyA speckles were detected upon its light-induced nucleocytoplasmic partitioning and a fraction remained in the cytoplasm. In this review, we present a concept that this complex picture of the phyA action is due, at least partially, to the existence of two phyA types in the cell differing by the structure of the N-terminus, probably, by its serine phosphorylation. These are phosphorylated water-soluble phyA' and underphosphorylated ambiquitous phyA″ represented by two fractions - water-soluble and membrane-associated. From the analysis of the phyA pools' activity in the regulation of phyA synthesis, seed germination, seedling establishment, and (proto)chlorophyll biosynthesis it is concluded that phyA″ is responsible for the regulation of seed germination, whereas in seedlings phyA' mediates the VLFRs, and the water-soluble phyA″ fraction, the HIRs. The membrane-associated phyA″ is likely to be active in cytoplasmic photoregulatory events. Functional interaction between phyA and the defense-related hormone jasmonic acid is also considered.

Arabidopsis SPA2 represses seedling de-etiolation under multiple light conditions.

In Arabidopsis, phytochrome (phy) A, phyB, and cryptochrome 1 (cry1) are representative far-red, red, and blue light photoreceptors, respectively. Members of the SUPPRESSOR OF PHYA-105 (SPA) protein family (SPA1-SPA4) form E3 ubiquitin ligase complexes with CONSTITUTIVE PHOTOMORPHOGENIC1 (COP1), which mediates the degradation of photomorphogenesis-promoting factors to desensitize light signaling. SPA2 has been reported to promote seedling etiolation in the dark. However, the unique roles of SPA2 and its three functional domains in suppressing photomorphogenesis under different light conditions are largely unknown. Here, we demonstrate that overexpression of the full-length or the central coiled-coil and C-terminal WD-repeat domains of SPA2 cause hyper-etiolation phenotypes under several light conditions. The SPA2 central coiled-coil and C-terminal WD-repeat domains are necessary and sufficient for repressing seedling de-etiolation, cotyledon unfolding, and promoting hypocotyl negative gravitropism under several light conditions. Furthermore, phyA, phyB, cry1, and COP1 repress protein accumulation or nuclear translocation of SPA2 through direct interactions with its kinase-like and coiled-coil domains located in the N-terminus in response to far-red, red, and blue light treatments, respectively. Taken together, our results demonstrate that SPA2 functions under multiple light conditions; moreover, light-activated photoreceptors rapidly suppress SPA2 activity via direct interactions in response to different light treatments.

Phytochromes A and B Mediate Light Stabilization of BIN2 to Regulate Brassinosteroid Signaling and Photomorphogenesis in Arabidopsis.

Phytochromes A and B (phyA and phyB) are the far-red and red lights photoreceptors mediating many light responses in Arabidopsis thaliana. Brassinosteroid (BR) is a pivotal phytohormone regulating a variety of plant developmental processes including photomorphogenesis. It is known that phyB interacts with BES1 to inhibit its DNA-binding activity and repress BR signaling. Here, we show that far-red and red lights modulate BR signaling through phyA and phyB regulation of the stability of BIN2, a glycogen synthase kinase 3 (GSK3)-like kinase that phosphorylates BES1/BZR1 to inhibit BR signaling. The BIN2 gain-of-function mutant bin2-1 displays an enhanced photomorphogenic phenotype in both far-red and red lights. phyA-enhanced accumulation of BIN2 promotes the phosphorylation of BES1 in far-red light. BIN2 acts genetically downstream from PHYA to regulate photomorphogenesis under far-red light. Both phyA and phyB interact directly with BIN2, which may promote the interaction of BIN2 with BES1 and induce the phosphorylation of BES1. Our results suggest that far-red and red lights inhibit BR signaling through phyA and phyB stabilization of BIN2 and promotion of BES1 phosphorylation, which defines a new layer of the regulatory mechanism that allows plants to coordinate light and BR signaling pathways to optimize photomorphogenesis.

The role of phytochrome-mediated gibberellic acid signaling in the modulation of seed germination under low light stress in rice (O. sativa L.).

Seed germination plays cardinal roles in seedling establishment and their successive growth. However, seed germination is retarded by far-red (FR) enrichment under low light stress, and the inhibitory signalling mechanism remains ambiguous. Our results indicated that low light treatment, both in the open and growth chamber conditions, inhibits rice seed germination by decreasing the gibberellin (GA) contents. To explore the mechanism of GA-deficiency under low light stress, differential expression profiling of GA-anabolic, -catabolic, ABA -anabolic, -catabolic, and SLR1 was investigated, revealing that expression of ABA- anabolic, GA-catabolic genes and SLR1 was upregulated with a simultaneous downregulation of ABA-catabolic and GA-anabolic genes under low light treatment. These results suggested that FR-induced GA inadequacy is resulted by upregulation of SLR1 and GA-catabolism genes consequently increase DELLA that further subsided GA-responses in the germinating rice seeds. Moreover, we provided evidence that FR-induced GA inadequacy demotes rice seed germination by decreasing amylase activity, eventually decreasing the carbohydrate solubilization in the germinating seeds. Finally, we suggest that under low light stress, due to a retarded conversion of phytochrome A to their bioactive form, the ABA-catabolic genes were eventually upregulated with a simultaneous downregulation of GA-anabolic genes. Consequently, a lower GA pool fails to leverage the GA-dependent DELLA degradation, further shutting down the expected GA responses that reduce germination efficiency under FR-enriched light. Supplementary Information: The online version contains supplementary material available at 10.1007/s12298-022-01167-7.

Feedback loop promotes sucrose accumulation in cotyledons to facilitate sugar-ethylene signaling-mediated, etiolated-seedling greening.

De-etiolation is indispensable for seedling survival and development. However, how sugars regulate de-etiolation and how sugars induce ethylene (ET) for seedlings to grow out of soil remain elusive. Here, we reveal how a sucrose (Suc) feedback loop promotes de-etiolation by inducing ET biosynthesis. Under darkness, Suc in germinating seeds preferentially induces 1-amino-cyclopropane-1-carboxylate synthase (ACS7; encoding a key ET biosynthesis enzyme) and associated ET biosynthesis, thereby activating ET core component ETHYLENE-INSENSITIVE3 (EIN3). Activated EIN3 directly inhibits the function of Suc transporter 2 (SUC2; a major Suc transporter) to block Suc export from cotyledons and thereby elevate Suc accumulation of cotyledons to induce ET. Under light, ET-activated EIN3 directly inhibits the function of phytochrome A (phyA; a de-etiolation inhibitor) to promote de-etiolation. We therefore propose that under darkness, the Suc feedback loop (Suc-ACS7-EIN3-|SUC2-Suc) promotes Suc accumulation in cotyledons to guarantee ET biosynthesis, facilitate de-etiolation, and enable seedlings to grow out of soil.

Functional Characterization of Tomato Phytochrome A and B1B2 Mutants in Response to Heat Stress.

Heat stress (HS) is a prevalent negative factor affecting plant growth and development, as it is predominant worldwide and threatens agriculture on a large scale. PHYTOCHROMES (PHYs) are photoreceptors that control plant growth and development, and the stress signaling response partially interferes with their activity. PHYA, B1, and B2 are the most well-known PHY types in tomatoes. Our study aimed to identify the role of tomato 'Money Maker' phyA and phyB1B2 mutants in stable and fluctuating high temperatures at different growth stages. In the seed germination and vegetative growth stages, the phy mutants were HS tolerant, while during the flowering stage the phy mutants revealed two opposing roles depending on the HS exposure period. The response of the phy mutants to HS during the fruiting stage showed similarity to WT. The most obvious stage that demonstrated phy mutants' tolerance was the vegetative growth stage, in which a high degree of membrane stability and enhanced water preservation were achieved by the regulation of stomatal closure. In addition, both mutants upregulated the expression of heat-responsive genes related to heat tolerance. In addition to lower malondialdehyde accumulation, the phyA mutant enhanced proline levels. These results clarified the response of tomato phyA and phyB1B2 mutants to HS.