Salicylic acid-activated BIN2 phosphorylation of TGA3 promotes Arabidopsis PR gene expression and disease resistance.

The plant defense hormone, salicylic acid (SA), plays essential roles in immunity and systemic acquired resistance. Salicylic acid induced by the pathogen is perceived by the receptor nonexpressor of pathogenesis-related genes 1 (NPR1), which is recruited by TGA transcription factors to induce the expression of pathogenesis-related (PR) genes. However, the mechanism by which post-translational modifications affect TGA's transcriptional activity by salicylic acid signaling/pathogen infection is not well-established. Here, we report that the loss-of-function mutant of brassinosteroid insensitive2 (BIN2) and its homologs, bin2-3 bil1 bil2, causes impaired pathogen resistance and insensitivity to SA-induced PR gene expression, whereas the gain-of-function mutant, bin2-1, exhibited enhanced SA signaling and immunity against the pathogen. Our results demonstrate that salicylic acid activates BIN2 kinase, which in turn phosphorylates TGA3 at Ser33 to enhance TGA3 DNA binding ability and NPR1-TGA3 complex formation, leading to the activation of PR gene expression. These findings implicate BIN2 as a new component of salicylic acid signaling, functioning as a key node in balancing brassinosteroid-mediated plant growth and SA-induced immunity.

The deubiquitinating enzymes UBP12 and UBP13 positively regulate recovery after carbon starvation by modulating BES1 stability in Arabidopsis thaliana.

BRI1-EMS-SUPPRESSOR1 (BES1), a core transcription factor in the brassinosteroid (BR) signaling pathway, primarily regulates plant growth and development by influencing BR-regulated gene expression. Several E3 ubiquitin (Ub) ligases regulate BES1 stability, but little is known about BES1 deubiquitination, which antagonizes E3 ligase-mediated ubiquitination to maintain BES1 homeostasis. Here, we report that two Arabidopsis thaliana deubiquitinating enzymes, Ub-SPECIFIC PROTEASE (UBP) 12 and UBP13, interact with BES1. UBP12 and UBP13 removed Ub from polyubiquitinated BES1 to stabilize both phosphorylated and dephosphorylated forms of BES1. A double mutant, ubp12-2w ubp13-3, lacking UBP12 and UBP13 function showed both BR-deficient and BR-insensitive phenotypes, whereas transgenic plants overexpressing UBP12 or UBP13 exhibited an increased BR response. Expression of UBP12 and UPB13 was induced during recovery after carbon starvation, which led to BES1 accumulation and quick recovery of stressed plants. Our work thus establishes a mechanism by which UBP12 and UBP13 regulate BES1 protein abundance to enhance BR-regulated growth during recovery after carbon starvation.

NAC transcription factor ATAF1 negatively modulates the PIF-regulated hypocotyl elongation under a short-day photoperiod.

Day length modulates hypocotyl elongation in seedlings to optimize their overall fitness. Variations in cell growth-associated genes are regulated by several transcription factors. However, the specific transcription factors through which the plant clock increases plant fitness are still being elucidated. In this study, we identified the no apical meristem, Arabidopsis thaliana-activating factor (ATAF-1/2), and cup-shaped cotyledon (NAC) family transcription factor ATAF1 as a novel repressor of hypocotyl elongation under a short-day (SD) photoperiod. Variations in day length profoundly affected the transcriptional and protein levels of ATAF1. ATAF1-deficient mutant exhibited increased hypocotyl length and cell growth-promoting gene expression under SD conditions. Moreover, ATAF1 directly targeted and repressed the expression of the cycling Dof factor 1/5 (CDF1/5), two key transcription factors involved in hypocotyl elongation under SD conditions. Additionally, ATAF1 interacted with and negatively modulated the effects of phytochrome-interacting factor (PIF), thus inhibiting PIF-promoted gene expression and hypocotyl elongation. Taken together, our results revealed ATAF1-PIF as a crucial pair modulating the expression of key transcription factors to facilitate plant growth during day/night cycles under fluctuating light conditions.

Nitrogen-inducible GLK1 modulates phosphate starvation response via the PHR1-dependent pathway.

Phosphorus (P) is a limiting nutrient for plant growth and productivity. Thus, a deep understanding of the molecular mechanisms of plants' response to phosphate starvation is significant when breeding crops with higher phosphorus-use efficiency. Here, we found that GARP-type transcription factor GLK1 acted as a positive regulator for phosphate-starvation response (PSR) via the PHR1-dependent pathway in Arabidopsis thaliana. GLK1 increased the transcription activity of PHR1 through the direct physical interaction and regulated the multiple responses to inorganic orthophosphate (Pi) starvation. Nitrogen (N) is a key factor in the regulation of PSR. We also found that the N status controlled the function of the GLK1-PHR1 signaling module under Pi-deficient (LP) conditions by regulating the accumulation of GLK1 and PHR1. Ultimately, we showed that the presence of GLK1 effectively promoted the protein accumulation of PHR1 at low N concentrations, and this action was helpful to maintain the activation of PSR. According to these findings, we establish the working model for GLK1 in PSR and propose that GLK1 mediates the interaction between N and P by influencing the effect of N on PHR1 in Arabidopsis thaliana.

Brassinosteroids enhance BES1-required thermomemory in Arabidopsis thaliana.

Heat stress (HS) caused by ambient high temperature poses a threat to plants. In the natural and agricultural environment, plants often encounter repeated and changeable HS. Moderate HS primes plants to establish a molecular 'thermomemory' that enables plants to withstand a later-and possibly more extreme-HS attack. Recent years, brassinosteroids (BRs) have been implicated in HS response, whereas the information is lacking on whether BRs signal transduction modulates thermomemory. Here, we uncover the positive role of BRs signalling in thermomemory of Arabidopsis thaliana. Heat priming induces de novo synthesis and nuclear accumulation of BRI1-Ethyl methyl sulfon-SUPPRESSOR (BES1), which is the key regulator of BRs signalling. BRs promote the accumulation of dephosphorylated BES1 during memory phase, and stoppage of BRs synthesis impairs dephosphorylation. During HS memory, BES1 is required to maintain sustained induction of HS memory genes and directly targets APX2 and HSFA3 for activation. In summary, our results reveal a BES1-required, BRs-enhanced transcriptional control module of thermomemory in Arabidopsis thaliana.

NAP is involved in GA-mediated chlorophyll degradation and leaf senescence by interacting with DELLAs in Arabidopsis.

KEY MESSAGE: RGA/GAI and NAP interacted with each other, and NAP was involved in GA signaling as a role of regulating age-dependent and dark-induced leaf senescence in Arabidopsis. Leaf senescence is a significant biological process which is beneficial for plant growth, development, and generation alternation in Arabidopsis. Recent researches have shown gibberellins (GAs) could accelerate leaf senescence. Nevertheless, the GA signaling involved in leaf senescence process remains elusive. Here, we reported a new potential regulation mechanism of GA-mediated chlorophyll degradation and leaf senescence. In this study, we confirmed that NAP positively regulated age-dependent and dark-induced leaf senescence and NAP knockout mutant nap was hyposensitive to GA3 (an active form of GA) treatment. DELLA family proteins with highly conserved structural domain function as master growth repressors that integrated GA signaling and leaf senescence. We validated RGA and GAI could interact with NAP in vitro and in vivo, and subsequently impaired the transcriptional activities of NAP to induce SAG113 and AAO3 expression in nap protoplasts. Taken together, we suggest that NAP is a novel component of the regulatory network that modulates the progress of leaf senescence in GA signaling.

SHY2 as a node in the regulation of root meristem development by auxin, brassinosteroids, and cytokinin.

In multicellular organisms, the balance between cell division and differentiation determines organ size, and represents a central unknown in developmental biology. In Arabidopsis roots, this balance is mediated between cytokinin and auxin through a regulatory circuit converging on the IAA3/SHORT HYPOCOTYL 2 (SHY2) gene. Here, we show that crosstalk between brassinosteroids (BRs) and auxin occurs in the vascular transition zone to promote root meristem development. We found that BR increases root meristem size by up-regulating expression of the PINFORMED 7 (PIN7) gene and down-regulating expression of the SHY2 gene. In addition, BES1 could directly bind to the promoter regions of both PIN7 and SHY2, indicating that PIN7 and SHY2 mediate the BR-induced growth of the root meristem by serving as direct targets of BES1. Moreover, the PIN7 overexpression and loss-of-function SHY2 mutant were sensitive to the effects of BR and could partially suppress the short-root phenotypes associated with deficient BR signaling. Interestingly, BRs could inhibit the accumulation of SHY2 protein in response to cytokinin. Taken together, these findings suggest that a complex equilibrium model exists in which regulatory interactions among BRs, auxin, and cytokinin regulate optimal root growth.

Mitochondrial alternative oxidase-dependent autophagy involved in ethylene-mediated drought tolerance in Solanum lycopersicum.

Mitochondrial alternative oxidase (AOX) is involved in a large number of plant physiological processes, such as growth, development and stress responses; however, the exact role of AOX in response to drought remains unclear. In our study, we provide solid evidences that the activated AOX capacity positively involved in ethylene-induced drought tolerance, in tomato (Solanum lycopersicum), accompanied by the changing level of hydrogen peroxide (H2 O2 ) and autophagy. In AOX1a-RNAi plants, the ethylene-induced drought tolerance was aggravated and associated with decreasing level of autophagy. The H2 O2 level was relatively higher in AOX1a-RNAi plants, whereas it was lower in AOX1a-overexpressing (35S-AOX1a-OE) plants after 1-(aminocarbonyl)-1-cyclopropanecarboxylic acid (ACC) pretreatment in the 14th day under drought stress. Interestingly, the accumulation of autophagosome was accompanied by the changing level of reactive oxygen species (ROS) in AOX transgenic tomato under drought stress whether or not pretreated with ACC. Pharmacological scavenging of H2 O2 accumulation in AOX1a-RNAi (aox19) stimulated autophagy acceleration under drought stress, and it seems that AOX-dependent ROS signalling is critical in triggering autophagy. Lower levels of ROS signalling positively induce autophagy activity, whereas higher ROS level would lead to rapid programmed cell death (PCD), especially in ethylene-mediated drought tolerance. Moreover, ethylene-induced autophagy during drought stress also can be through ERF5 binding to the promoters of ATG8d and ATG18h. These results demonstrated that AOX plays an essential role in ethylene-induced drought tolerance and also played important roles in mediating autophagy generation via balancing ROS level.

Cooperative transcriptional regulation by ATAF1 and HY5 promotes light-induced cotyledon opening in Arabidopsis thaliana.

The opening of the embryonic leaves (cotyledons) as seedlings emerge from the dark soil into the light is crucial to ensure the survival of the plant. Seedlings that sprout in the dark elongate rapidly to reach light but keep their cotyledons closed. During de-etiolation, the transition from dark to light growth, elongation slows and the cotyledons open. Here, we report that the transcription factor ACTIVATING FACTOR1 (ATAF1) participates in de-etiolation and facilitates light-induced cotyledon opening. The transition from dark to light rapidly induced ATAF1 expression and ATAF1 accumulation in cotyledons. Seedlings lacking or overexpressing ATAF1 exhibited reduced or enhanced cotyledon opening, respectively, and transcriptomic analysis indicated that ATAF1 repressed the expression of genes associated with growth and cotyledon closure. The activation of the photoreceptor phytochrome A (phyA) by far-red light induced its association with the ATAF1 promoter and stimulation of ATAF1 expression. The transcription factor ELONGATED HYPOCOTYL5 (HY5), which is also activated in response far-red light, cooperated with phyA to induce ATAF1 expression. ATAF1 and HY5 interacted with one another and cooperatively repressed the expression of growth-promoting and cotyledon closure genes. Together, our study reveals a mechanism through which far-red light promotes cotyledon opening.

Mitochondrial alternative oxidase enhanced ABA-mediated drought tolerance in Solanum lycopersicum.

The phytohormone abscisic acid (ABA) plays essential roles in modulating drought stress responses. Mitochondrial alternative oxidase (AOX) is critical for reactive oxygen species (ROS) scavenging in drought stress responses. However, whether ABA signal in concert with AOX to moderate drought stress response remains largely unclear. In our study, we uncover the positive role of AOX in ABA-mediated drought tolerance in tomato (Solanum lycopersicum). Here, we report that ABA participates in the regulation of alternative respiration, and the increased AOX was found to improve drought tolerance by reducing total ROS accumulation. We also found that transcription factor ABA response element-binding factor 1 (SlAREB1) can directly bind to the promoter of AOX1a to activate its transcription. Virus-induced gene silencing (VIGS) of SlAREB1 compromised the ABA-induced alternative respiratory pathway, disrupted redox homeostasis and decreased plant resistance to drought stress, while overexpression of AOX1a in TRV2-SlAREB1 plants partially rescued the severe drought phenotype. Taken together, our results indicated that AOX1a plays an essential role in ABA-mediated drought tolerance partially in a SlAREB1-dependent manner, providing new insights into how ABA modulates ROS levels to cope with drought stress by AOX.