NAM/ATAF/CUC Research Articles

Identification of NAC Transcription Factors in Suaeda glauca and Their Responses to Salt Stress.

NAC (NAM/ATAF1/2/CUC2) transcription factors regulate plant growth and development and stress responses. Because NAC transcription factors are known to play important roles in the regulation of salt tolerance in many plants, we aimed to explore their roles in the halophyte Suaeda glauca. Based on transcriptome sequencing data, we identified 25 NAC transcription factor gene family members. In a phylogenetic tree analysis with Arabidopsis thaliana NAC transcription factors, the SgNACs were divided into 10 groups. The physicochemical properties and conserved domains of the putative proteins, as well as the transcript profiles of their encoding genes, were determined for the 25 SgNAC genes using bioinformatic methods. Most of the S. glauca NAC genes were upregulated to some extent after 24 h of salt stress, suggesting that they play an important role in regulating the salt tolerance of S. glauca. These findings lay the foundation for further research on the functions and mechanisms of the NAC gene family in S. glauca.

Functional characterization of VrNAC15 for drought resistance in mung beans

Drought stress has become an important limiting factor in mung bean production, and NAC(NAM/ATAF/CUC) transcription factors are crucial for plant growth under stress conditions, so it is important to study the regulatory role of NAC transcription factors in mung bean under drought stress. In this investigation, VrNAC15, along with its promoter, was cloned, and its structure was meticulously analyzed. Using qPCR, we examined the tissue-specific expression patterns of VrNAC15, particularly under drought stress and ABA exposure. Additionally, We performed ectopic expression of VrNAC15 in Arabidopsis to assess its function.. Gene sequence analysis revealed that VrNAC15 has a total length of 1014 bp, encoding 337 amino acids. It contains a NAM domain, localizes within the nucleus, and exhibits transcriptional activation. Promoter analysis of VrNAC15 identified essential core promoter elements and cis-acting elements related to abscisic acid, methyl jasmonate, gibberellin, adversity stress, light, and metabolism. Expression analysis demonstrated the concentration of VrNAC15 in leaves, with significant alterations following ABA and drought treatments in mung beans. Cluster analysis revealed that VrNAC15 may enhanced drought tolerance in transgenic plants through its expression. Transgenic experiments supported these findings, showing that heterologous expression of VrNAC15 led to enhanced antioxidant and osmotic adjustment capabilities in Arabidopsis plants. This resulted in the maintenance of cell membrane structural integrity during drought stress and normal physiological and biochemical metabolic reactions within cells. This research provides valuable insights into the structural and functional characteristics of the VrNAC15, setting the stage for future endeavors in molecular breeding for improved drought resistance in mung beans.

Transcriptional Control of Seed Life: New Insights into the Role of the NAC Family.

Transcription factors (TFs) regulate gene expression by binding to specific sequences on DNA through their DNA-binding domain (DBD), a universal process. This update conveys information about the diverse roles of TFs, focusing on the NACs (NAM-ATAF-CUC), in regulating target-gene expression and influencing various aspects of plant biology. NAC TFs appeared before the emergence of land plants. The NAC family constitutes a diverse group of plant-specific TFs found in mosses, conifers, monocots, and eudicots. This update discusses the evolutionary origins of plant NAC genes/proteins from green algae to their crucial roles in plant development and stress response across various plant species. From mosses and lycophytes to various angiosperms, the number of NAC proteins increases significantly, suggesting a gradual evolution from basal streptophytic green algae. NAC TFs play a critical role in enhancing abiotic stress tolerance, with their function conserved in angiosperms. Furthermore, the modular organization of NACs, their dimeric function, and their localization within cellular compartments contribute to their functional versatility and complexity. While most NAC TFs are nuclear-localized and active, a subset is found in other cellular compartments, indicating inactive forms until specific cues trigger their translocation to the nucleus. Additionally, it highlights their involvement in endoplasmic reticulum (ER) stress-induced programmed cell death (PCD) by activating the vacuolar processing enzyme (VPE) gene. Moreover, this update provides a comprehensive overview of the diverse roles of NAC TFs in plants, including their participation in ER stress responses, leaf senescence (LS), and growth and development. Notably, NACs exhibit correlations with various phytohormones (i.e., ABA, GAs, CK, IAA, JA, and SA), and several NAC genes are inducible by them, influencing a broad spectrum of biological processes. The study of the spatiotemporal expression patterns provides insights into when and where specific NAC genes are active, shedding light on their metabolic contributions. Likewise, this review emphasizes the significance of NAC TFs in transcriptional modules, seed reserve accumulation, and regulation of seed dormancy and germination. Overall, it effectively communicates the intricate and essential functions of NAC TFs in plant biology. Finally, from an evolutionary standpoint, a phylogenetic analysis suggests that it is highly probable that the WRKY family is evolutionarily older than the NAC family.

Overexpression of a Fragaria vesca NAM, ATAF, and CUC (NAC) Transcription Factor Gene (FvNAC29) Increases Salt and Cold Tolerance in Arabidopsis thaliana.

The NAC (NAM, ATAF1/2, CUC2) family of transcription factors (TFs) is a vital transcription factor family of plants. It controls multiple parts of plant development, tissue formation, and abiotic stress response. We cloned the FvNAC29 gene from Fragaria vesca (a diploid strawberry) for this research. There is a conserved NAM structural domain in the FvNAC29 protein. The highest homology between FvNAC29 and PaNAC1 was found by phylogenetic tree analysis. Subcellular localization revealed that FvNAC29 is localized onto the nucleus. Compared to other tissues, the expression level of FvNAC29 was higher in young leaves and roots. In addition, Arabidopsis plants overexpressing FvNAC29 had higher cold and high-salinity tolerance than the wild type (WT) and unloaded line with empty vector (UL). The proline and chlorophyll contents of transgenic Arabidopsis plants, along with the activities of the antioxidant enzymes like catalase (CAT), peroxidase (POD), and superoxide dismutase (SOD) under 200 mM NaCl treatment or -8 °C treatment, were higher than those activities of the control. Meanwhile, malondialdehyde (MDA) and the reactive oxygen species (ROS) content were higher in the WT and UL lines. FvNAC29 improves transgenic plant resistance to cold and salt stress by regulating the expression levels of AtRD29a, AtCCA1, AtP5CS1, and AtSnRK2.4. It also improves the potential to tolerate cold stress by positively regulating the expression levels of AtCBF1, AtCBF4, AtCOR15a, and AtCOR47. These findings suggest that FvNAC29 may be related to the processes and the molecular mechanisms of F. vesca response to high-salinity stress and LT stress, providing a comprehensive understanding of the NAC TFs.

MiR164-MhNAC1 regulates apple root nitrogen uptake under low nitrogen stress.

Nitrogen is an essential nutrient for plant growth and serves as a signaling molecule to regulate gene expression inducing physiological, growth and developmental responses. An excess or deficiency of nitrogen may have adverse effects on plants. Studying nitrogen uptake will help us understand the molecular mechanisms of utilization for targeted molecular breeding. Here, we identified and functionally validated an NAC (NAM-ATAF1/2-CUC2) transcription factor based on the transcriptomes of two apple rootstocks with different nitrogen uptake efficiency. NAC1, a target gene of miR164, directly regulates the expression of the high-affinity nitrate transporter (MhNRT2.4) and citric acid transporter (MhMATE), affecting root nitrogen uptake. To examine the role of MhNAC1 in nitrogen uptake, we produced transgenic lines that overexpressed or silenced MhNAC1. Silencing MhNAC1 promoted nitrogen uptake and citric acid secretion in roots, and enhanced plant tolerance to low nitrogen conditions, while overexpression of MhNAC1 or silencing miR164 had the opposite effect. This study not only revealed the role of the miR164-MhNAC1 module in nitrogen uptake in apple rootstocks but also confirmed that citric acid secretion in roots affected nitrogen uptake, which provides a research basis for efficient nitrogen utilization and molecular breeding in apple.

Transcription factor NtNAC56 regulates jasmonic acid-induced leaf senescence in tobacco.

Leaf senescence is a vital aspect of plant physiology and stress responses and is induced by endogenous factors and environmental cues. The plant-specific NAC (NAM, ATAF1/2, CUC2) transcription factor family influences growth, development, and stress responses in Arabidopsis (Arabidopsis thaliana) and other species. However, the roles of NACs in tobacco (Nicotiana tabacum) leaf senescence are still unclear. Here, we report that NtNAC56 regulates leaf senescence in tobacco. Transgenic plants overexpressing NtNAC56 (NtNAC56-OE) showed induction of senescence-related genes and exhibited early senescence and lower chlorophyll content compared to wild-type (WT) plants and the Ntnac56-19 mutant. In addition, root development and seed germination were inhibited in the NtNAC56-OE lines. Transmission electron microscopy observations accompanied by physiological and biochemical assays revealed that NtNAC56 overexpression triggers chloroplast degradation and reactive oxygen species accumulation in tobacco leaves. Transcriptome analysis demonstrated that NtNAC56 activates leaf senescence-related genes and jasmonic acid (JA) biosynthesis pathway genes. In addition, the JA content of NtNAC56-OE plants was higher than in WT plants, and JA treatment induced NtNAC56 expression. We performed DNA affinity purification sequencing to identify direct targets of NtNAC56, among which we focused on LIPOXYGENASE 5 (NtLOX5), a key gene in JA biosynthesis. A dual-luciferase reporter assay and a yeast one-hybrid assay confirmed that NtNAC56 directly binds to the TTTCTT motif in the NtLOX5 promoter. Our results reveal a mechanism whereby NtNAC56 regulates JA-induced leaf senescence in tobacco and provide a strategy for genetically manipulating leaf senescence and plant growth.

Rice transcription factor OsNAC2 maintains the homeostasis of immune responses to bacterial blight.

Rice (Oryza sativa) bacterial blight, caused by Xanthomonas oryzae pv. Oryzae (Xoo), threatens plant growth and yield. However, the molecular mechanisms underlying rice immunity against Xoo remain elusive. Here, we identified a NAC (NAM-ATAF-CUC) transcription factor OsNAC2 as a negative regulator in the resistance to bacterial blight disease in rice. Constitutive overexpression of OsNAC2 inhibited the expression of salicylic acid (SA) biosynthesis-related genes (i.e. isochorismate synthase 1 (OsICS1), phenylalanine ammonia lyase 3 (OsPAL3), etc.) with adverse impacts on the pathogenesis-related proteins (PRs) responses and compromised blight resistance. Moreover, OsNAC2 interacted with APETALA2/ethylene-responsive element binding protein (AP2/EREBP) transcription factor OsEREBP1 and possibly threatened its protein stability, destroying the favorable interaction of OsEREBP1-Xa21-binding protein OsXb22a in the cytoplasm during Xoo-induced infection. On the contrary, downregulation of OsNAC2 resulted in enhanced resistance to bacterial blight in rice without any growth or yield penalties. Our results demonstrated that OsNAC2 inhibits SA signaling and stably interacted with OsEREBP1 to impair disease resistance. This OsNAC2-OsEREBP1-based homeostatic mechanism provided insights into the competition between rice and bacterial pathogens, and it will be useful to improve the disease resistance of important crops through breeding.

The NAC transcription factor LuNAC61 negatively regulates fiber development in flax (Linum usitatissimum L.)

Flax is a crucial fiber crop that exhibits excellent textile properties and serves as a model plant for investigating phloem fiber development. The regulation of multiple genes significantly influences fiber development, notably involving NAC (NAM, ATAF1/2, CUC2) transcription factors in forming the fiber secondary cell wall (SCW). Overexpression of LuNAC61 in flax resulted in sparse top meristematic zone leaves and significantly reduced stem cellulose content. Scanning electron microscopy and staining observations revealed a significant reduction in fiber bundles. β-Glucuronidase (GUS) staining analysis demonstrated high activity of the LuNAC61 promoter in the bast fibers of the flax stem. Additionally, several members of the LuPLATZ and LuCesA families exhibited significant coexpression with LuNAC61. Subcellular localization indicated the presence of LuPLATZ24 protein in the nucleus and cytoplasm, LuNAC61 protein exclusively in the nucleus, and LuCesA10 in the nucleus and endoplasmic reticulum. LuPLATZ24 positively regulates LuNAC61, whereas LuNAC61 negatively affects LuCesA10, suggesting the involvement of a metabolic network in regulating flax fiber development. In conclusion, this study provides a critical opportunity for a comprehensive and in-depth analysis of the mechanisms governing flax fiber development and the potential use of biotechnology to enhance flax fiber yield.

OsNAC120 balances plant growth and drought tolerance by integrating GA and ABA signaling in rice

The crosstalk between gibberellin (GA) and abscisic acid (ABA) signaling is crucial for balancing plant growth and adaption to environmental stress. Nevertheless, the molecular mechanism of their mutual antagonism still remains to be fully clarified. In this study, we found that knockout of the rice NAC (NAM, ATAF1/2, CUC2) transcription factor gene OsNAC120 inhibits plant growth but enhances drought tolerance, whereas OsNAC120 overexpression produces the opposite results. Exogenous GA can rescue the semi-dwarf phenotype of osnac120 mutants, and further study showed that OsNAC120 promotes GA biosynthesis by transcriptionally activating the GA biosynthetic genes OsGA20ox1 and OsGA20ox3. The DELLA protein SLENDER RICE1 (SLR1) interacts with OsNAC120 and impedes its transactivation ability, and GA treatment can remove the inhibition of transactivation activity caused by SLR1. On the other hand, OsNAC120 negatively regulates rice drought tolerance by repressing ABA-induced stomatal closure. Mechanistic investigation revealed that OsNAC120 inhibits ABA biosynthesis via transcriptional repression of the ABA biosynthetic genes OsNCED3 and OsNCED4. Rice OSMOTIC STRESS/ABA-ACTIVATED PROTEIN KINASE 9 (OsSAPK9) physically interacts with OsNAC120 and mediates its phosphorylation, which results in OsNAC120 degradation. ABA treatment accelerates OsNAC120 degradation and reduces its transactivation activity. Together, our findings provide evidence that OsNAC120 plays critical roles in balancing GA-mediated growth and ABA-induced drought tolerance in rice. This research will help us to understand the mechanisms underlying the trade-off between plant growth and stress tolerance and to engineer stress-resistant, high-yielding crops.

Rice Basic Helix-Loop-Helix 079 (OsbHLH079) Delays Leaf Senescence by Attenuating ABA Signaling

Leaf senescence represents the final phase of leaf development and is characterized by a highly organized degenerative process involving the active translocation of nutrients from senescing leaves to growing tissues or storage organs. To date, a large number of senescence-associated transcription factors (sen-TFs) have been identified that regulate the initiation and progression of leaf senescence. Many of these TFs, including NAC (NAM/ATAF1/2/CUC2), WRKY, and MYB TFs, have been implicated in modulating the expression of downstream senescence-associated genes (SAGs) and chlorophyll degradation genes (CDGs) under the control of phytohormones. However, the involvement of basic helix-loop-helix (bHLH) TFs in leaf senescence has been less investigated. Here, we show that OsbHLH079 delays both natural senescence and dark-induced senescence: Overexpression of OsbHLH079 led to a stay-green phenotype, whereas osbhlh079 knockout mutation displayed accelerated leaf senescence. Similar to other sen-TFs, OsbHLH079 showed a gradual escalation in expression as leaves underwent senescence. During this process, the mRNA levels of SAGs and CDGs remained relatively low in OsbHLH079 overexpressors, but increased sharply in osbhlh079 mutants, suggesting that OsbHLH079 negatively regulates the transcription of SAGs and CDGs under senescence conditions. Additionally, we found that OsbHLH079 delays ABA-induced senescence. Subsequent RT-qPCR and dual-luciferase reporter assays revealed that OsbHLH079 downregulates the expression of ABA signaling genes, such as OsABF2, OsABF4, OsABI5, and OsNAP. Taken together, these results demonstrate that OsbHLH079 functions in delaying leaf yellowing by attenuating the ABA responses.

Control of stem cell behavior by CLE-JINGASA signaling in the shoot apical meristem in Marchantia polymorpha

Land plants undergo indeterminate growth by the activity of meristems in both gametophyte (haploid) and sporophyte (diploid) generations. In the sporophyte of the flowering plant Arabidopsis thaliana, the apical meristems are located at the shoot and root tips in which a number of regulatory gene homologs are shared for their development, implying deep evolutionary origins. However, little is known about their functional conservation with gametophytic meristems in distantly related land plants such as bryophytes, even thoughgenomic studies have revealed that the subfamily-level diversity of regulatory genes is mostly conserved throughout land plants. Here, we show that a NAM/ATAF/CUC (NAC) domain transcription factor, JINGASA (MpJIN), acts downstream of CLAVATA3 (CLV3)/ESR-related (CLE) peptide signaling and controls stem cell behavior in the gametophytic shoot apical meristem of the liverwort Marchantia polymorpha. In the meristem, strong MpJIN expression was associated with the periclinal cell division at the periphery of the stem cell zone (SCZ), whereas faint MpJIN expression was found at the center of the SCZ. Time course observation indicates that the MpJIN-negative cells are lost from the SCZ and respecified de novo at two separate positions during the dichotomous branching event. Consistently, the induction of MpJIN results in ectopic periclinal cell division in the SCZ and meristem termination. Based on the comparative expression data, we speculate that the function of JIN/FEZ subfamily genes was shared among the shoot apical meristems in the gametophyte and sporophyte generations in early land plants but was lost in certain lineages, including the flowering plant A.thaliana.

Two adjacent NAC transcription factors regulate fruit maturity date and flavor in peach.

Although maturity date (MD) is an essential factor affecting fresh fruit marketing and has a pleiotropic effect on fruit taste qualities, the underlying mechanisms remain largely unclear. In this study, we functionally characterized two adjacent NAM-ATAF1/2-CUC2 (NAC) transcription factors (TFs), PpNAC1 and PpNAC5, both of which were associated with fruit MD in peach. PpNAC1 and PpNAC5 were found capable of activating transcription of genes associated with cell elongation, cell wall degradation and ethylene biosynthesis, suggesting their regulatory roles in fruit enlargement and ripening. Furthermore, PpNAC1 and PpNAC5 had pleiotropic effects on fruit taste due to their ability to activate transcription of genes for sugar accumulation and organic acid degradation. Interestingly, both PpNAC1 and PpNAC5 orthologues were found in fruit-producing angiosperms and adjacently arranged in all 91 tested dicots but absent in fruitless gymnosperms, suggesting their important roles in fruit development. Our results provide insight into the regulatory roles of NAC TFs in MD and fruit taste.

Comparative Physiological and Transcriptome Analyses Reveal Mechanisms of Salicylic-Acid-Reduced Postharvest Ripening in 'Hosui' Pears (Pyrus pyrifolia Nakai).

Postharvest ripening of sand pear fruit leads to quality deterioration, including changes in texture, flavor, and fruit color. Salicylic acid (SA), an important defense-related hormone, delays fruit ripening and maintains fruit quality, but the underling mechanism remains unclear. Herein, we evaluated the efficacy of SA in delaying the ripening process of Pyrus pyrifolia cv. 'Hosui' pear fruit, as evidenced by the reduction in fruit weight loss, inhibition of firmness loss, cell wall degradation and soluble sugars, and retention of total phenols. Based on comparative transcriptomic data, a total of 3837 and 1387 differentially expressed genes (DEGs) were identified during room-temperature storage of control fruit and between SA-treated and control fruit, respectively. Further KEGG analysis revealed that the DEGs were mainly implicated in plant hormone signal transduction, starch and sugar metabolism, and cell wall modification. Moreover, exogenous SA treatment also altered the expression of many transcription factor (TF) families, including those in the ethylene-responsive factor (ERF), NAM, ATAF, CUC (NAC), basic helix-loop-helix (bHLH), basic leucine zipper (bZIP), and v-myb avian myeloblastosis viral oncogene homolog (MYB) families. Together, the results offer important insights into the role of SA-responsive genes in controlling fruit ripening in sand pears.

MaMADS1-MaNAC083 transcriptional regulatory cascade regulates ethylene biosynthesis during banana fruit ripening.

The hormone ethylene is crucial in the regulation of ripening in climacteric fruit, such as bananas. The transcriptional regulation of ethylene biosynthesis throughout banana fruit ripening has received much study, but the cascaded transcriptional machinery of upstream transcriptional regulators implicated in the ethylene biosynthesis pathway is still poorly understood. Here we report that ethylene biosynthesis genes, including MaACS1, MaACO1, MaACO4, MaACO5, and MaACO8, were upregulated in ripening bananas. NAC (NAM, ATAF, CUC) transcription factor, MaNAC083, a ripening and ethylene-inhibited gene, was discovered as a potential binding protein to the MaACS1 promoter by yeast one-hybrid screening. Further in vitro and in vivo experiments indicated that MaNAC083 bound directly to promoters of the five ethylene biosynthesis genes, thereby transcriptionally repressing their expression, which was further verified by transient overexpression experiments, where ethylene production was inhibited through MaNAC083-modulated transcriptional repression of ethylene biosynthesis genes in banana fruits. Strikingly, MaMADS1, a ripening-induced MADS (MCM1, AGAMOUS, DEFICIENS, SRF4) transcription factor, was found to directly repress the expression of MaNAC083, inhibiting trans-repression of MaNAC083 to ethylene biosynthesis genes, thereby attenuating MaNAC083-repressed ethylene production in bananas. These findings collectively illustrated the mechanistic basis of a MaMADS1-MaNAC083-MaACS1/MaACOs regulatory cascade controlling ethylene biosynthesis during banana fruit ripening. These findings increase our knowledge of the transcriptional regulatory mechanisms of ethylene biosynthesis at the transcriptional level and are expected to help develop molecular approaches to control ripening and improve fruit storability.

Expressing a Short Tandem Target Mimic (STTM) of miR164b/e-3p enhances poplar leaf serration by co-regulating the miR164–NAC module

MicroRNAs (miRNAs) are short non-coding RNAs (21–24 nt) that play important roles in plant growth and development. The miR164 family is highly conserved in plants and the miR164–NAM/ATAF/CUC (NAC) module is validated to regulate leaf and flower development, lateral root initiation and stress response. However, our knowledge of its role in Populus remains limited. In this study, two mature miRNA species, miR164e-5p and miR164e-3p, were identified in Populus deltoides. Their nucleotide sequences were identical to those of miR164a/b/c/d/e-5p and miR164b/e-3p in P. tremula × P. alba clone 717-1B4 (hereinafter poplar 717), respectively. Transgenic plants of poplar 717, including overexpression lines (35S::pri-miR164e) and Short Tandem Target Mimic lines (STTM-miR164a-d,e-5p and STTM-miR164b/e-3p), were generated to study the roles of miR164e-5p and miR164e-3p in poplar. Compared with poplar 717, the leaf margins of 35S::pri-miR164e lines were smoother, the leaves of STTM-miR164b/e-3p line were more serrated, while the leaf morphology of STTM-miR164a-d,e-5p lines had no obvious change. In addition, both 35S::pri-miR164e and STTM-miR164b/e-3p plants had a dwarf phenotype. Expressions of miR164a-d,e-5p target genes, including PtaCUC2a, PtaCUC2b and PtaORE1, was significantly reduced in the apex of 35S::pri-miR164e lines. Green fluorescent protein (GFP) reporter assay showed that PtaCUC2a/2b and PtaORE1 were cleaved by miR164a-d,e-5p, and the cleavage was inhibited by STTM-miR164b/e-3p. Therefore, miR164b/e-3p may cooperate with miR164a-d,e-5p to regulate certain NAC members, such as PtaCUC2a/2b and PtaORE1, thereby regulating leaf development and plant growth in poplar. Our findings add new insights into the mechanisms by which the miR164–NAC module regulates plant development.

In silico analysis of NAC gene family in the mangrove plant Avicennia marina provides clues for adaptation to intertidal habitats.

NAC (NAM, ATAF1/2, CUC2) transcription factors (TFs) constitute a plant-specific gene family. It is reported that NAC TFs play important roles in plant growth and developmental processes and in response to biotic/abiotic stresses. Nevertheless, little information is known about the functional and evolutionary characteristics of NAC TFs in mangrove plants, a group of species adapting coastal intertidal habitats. Thus, we conducted a comprehensive investigation for NAC TFs in Avicennia marina, one pioneer species of mangrove plants. We totally identified 142 NAC TFs from the genome of A. marina. Combined with NAC proteins having been functionally characterized in other organisms, we built a phylogenetic tree to infer the function of NAC TFs in A. marina. Gene structure and motif sequence analyses suggest the sequence conservation and transcription regulatory regions-mediated functional diversity. Whole-genome duplication serves as the driver force to the evolution of NAC gene family. Moreover, two pairs of NAC genes were identified as positively selected genes of which AmNAC010/040 may be imposed on less constraint toward neofunctionalization. Quite a few stress/hormone-related responsive elements were found in promoter regions indicating potential response to various external factors. Transcriptome data revealed some NAC TFs were involved in pneumatophore and leaf salt gland development and response to salt, flooding and Cd stresses. Gene co-expression analysis found a few NAC TFs participates in the special biological processes concerned with adaptation to intertidal environment. In summary, this study provides detailed functional and evolutionary information about NAC gene family in mangrove plant A. marina and new perspective for adaptation to intertidal habitats.

Comprehensive Analysis of NAC Genes Reveals Differential Expression Patterns in Response to Pst DC3000 and Their Overlapping Expression Pattern during PTI and ETI in Tomato.

NAC (NAM/ATAF/CUC) transcription factors belong to a unique gene family in plants, which play vital roles in regulating diverse biological processes, including growth, development, senescence, and in response to biotic and abiotic stresses. Tomato (Solanum lycopersicum), as the most highly valued vegetable and fruit crop worldwide, is constantly attacked by Pseudomonas syringae pv. tomato DC3000 (Pst DC3000), causing huge losses in production. Thus, it is essential to conduct a comprehensive identification of the SlNAC genes involved in response to Pst DC3000 in tomato. In this study, a complete overview of this gene family in tomato is presented, including genome localization, protein domain architectures, physical and chemical features, and nuclear location score. Phylogenetic analysis identified 20 SlNAC genes as putative stress-responsive genes, named SSlNAC 1-20. Expression profiles analysis revealed that 18 of these 20 SSlNAC genes were significantly induced in defense response to Pst DC3000 stress. Furthermore, the RNA-seq data were mined and analyzed, and the results revealed the expression pattern of the 20 SSlNAC genes in response to Pst DC3000 during the PTI and ETI. Among them, SSlNAC3, SSlNAC4, SSlNAC7, SSlNAC8, SSlNAC12, SSlNAC17, and SSlNAC19 were up-regulated against Pst DC3000 during PTI and ETI, which suggested that these genes may participate in both the PTI and ETI pathway during the interaction between tomato and Pst DC3000. In addition, SSlNAC genes induced by exogenous hormones, including indole-3-acetic acid (IAA), abscisic acid (ABA), salicylic acid (SA), and methyl jasmonic acid (MeJA), were also recovered. These results implied that SSlNAC genes may participate in the Pst DC3000 stress response by multiple regulatory pathways of the phytohormones. In all, this study provides important clues for further functional analysis and of the regulatory mechanism of SSlNAC genes under Pst DC3000 stress.

Cowpea NAC1/NAC2 transcription factors improve growth and tolerance to drought and heat in transgenic cowpea through combined activation of photosynthetic and antioxidant mechanisms.

NAC (NAM/ATAF1/2/CUC2) transcription factors are central switches of growth and stress responses in plants. However, unpredictable interspecies conservation of function and regulatory targets makes the well-studied NAC orthologs inapt for pulse engineering. The knowledge of suitable NAC candidates in hardy pulses like cowpea (Vigna unguiculata (L.) Walp.) is still in infancy, hence warrants immediate biotechnological intervention. Here, we showed that overexpression of two native NAC genes (VuNAC1 and VuNAC2) promoted germinative, vegetative, and reproductive growth and conferred multiple abiotic stress tolerance in a commercial cowpea variety. The transgenic lines displayed increased leaf area, thicker stem, nodule-rich denser root system, early flowering, higher pod production (∼3.2-fold and ∼2.1-fold), and greater seed weight (10.3% and 6.0%). In contrast, transient suppression of VuNAC1/2 caused severe growth retardation and flower inhibition. The overexpressor lines showed remarkable tolerance to major yield-declining terminal stresses, such as drought, salinity, heat, and cold, and recovered growth and seed production by boosting photosynthetic activity, water use efficiency, membrane integrity, Na+ /K+ homeostasis, and antioxidant activity. The comparative transcriptome study indicated consolidated activation of genes involved in chloroplast development, photosynthetic complexes, cell division and expansion, cell wall biogenesis, nutrient uptake and metabolism, stress response, abscisic acid, and auxin signaling. Unlike their orthologs, VuNAC1/2 direct synergistic transcriptional tuning of stress and developmental signaling to avoid unwanted trade-offs. Their overexpression governs the favorable interplay of photosynthesis and reactive oxygen species regulation to improve stress recovery, nutritional sufficiency, biomass, and production. This unconventional balance of strong stress tolerance and agronomic quality is useful for translational crop research and molecular breeding of pulses.

Overexpression of ONAC054 Improves Drought Stress Tolerance and Grain Yield in Rice

Drought stress negatively affects plant growth and development, thus reducing plant productivity. Therefore, understanding the molecular mechanisms underlying drought stress responses is essential for crop improvement. The plant-specific NAM/ATAF1,2/CUC2 (NAC) transcription factors play important roles in the drought stress response. Here, we show that rice (Oryza sativa) ONAC054, a membrane-bound NAC transcription factor, is involved in the drought stress response. We found that onac054 mutants were sensitive, whereas ONAC054-overexpressing (ONAC054-OX) plants were tolerant to drought stress. Under drought stress conditions, several genes associated with abscisic acid (ABA) synthesis and signaling were downregulated in onac054 mutants but upregulated in ONAC054-OX plants. Among these genes, the TRANSCRIPTION FACTOR RESPONSIBLE FOR ABA REGULATION 1 (TRAB1), which encodes an ABA-inducible bZIP transcription factor, was directly activated by ONAC054. On the other hand, the expression of ONAC054 was directly activated by several ABA-responsive elements (ABRE)-binding factors (ABFs) in an ABA-dependent manner, indicating that ONAC054 acts as an enhancer of ABA-induced drought stress tolerance. Additionally, the overexpression of ONAC054 in rice greatly improved grain yield under drought stress conditions, indicating that the overexpression of ONAC054 could facilitate the improvement of drought stress tolerance in rice and other crops.

Bolstered plant tolerance to low temperatures by overexpressing NAC transcription factors: identification of critical variables by meta-analysis.

The potential biotechnological application of NAC overexpression has been challenged by meta-analysis, establishing a correlation between the magnitudes of several physiological and biochemical parameters and the enhanced tolerance to cold. Overexpression of various NAC (NAM/ATAF/CUC) transcription factors in different plant systems was shown to confer enhanced tolerance to low temperatures by inducing both common and distinctive stress response pathways. However, lack of consensus on the type of parameters evaluated, their magnitudes, and direction of the responses complicates drawing general conclusions on the effects of NAC expression in plant physiology. We report herein a meta-analysis summarizing the most critical response variables used to study the effect of overexpressing NAC regulators on cold stress tolerance. We found that NAC overexpression affected all of the outcome parameters in stressed plants, and one response in control conditions. Transformed plants displayed an increase of at least 40% in positive responses, while negative outcomes were reduced by at least 30%. The most reported parameters included survival, electrolyte leakage, and malondialdehyde contents, whereas the most sensitive to the treatments were the Fv/Fm parameter, survival, and the activity of catalases. We also explored how different experimental arrangements affected the magnitudes of the responses. NAC-mediated improvements were best observed after severe stress episodes and during brief treatments (ranging from 5 to 24h), especially in terms of antioxidant activities, accumulation of free proline, and parameters related to membrane integrity. Use of heterologous expression also favored several indicators of plant fitness. Our findings should help both basic and applied research on the influence of NAC expression on enhanced tolerance to cold.