R2R3-MYB Transcription Factor Research Articles

FaMYB6-like negatively regulates FaMYB10-induced anthocyanin accumulation during strawberry fruit ripening

Anthocyanins are one of the essential compounds affecting the quality of strawberry fruit. Several R2R3-MYB transcription factors have been well characterized as the anthocyanin activators in strawberry. However, we still lack valuable information on how R2R3-MYB repressors regulate strawberry anthocyanin accumulation. Here, we cloned a novel R2R3-MYB gene, FaMYB6-like, and characterized its repressive role during anthocyanin accumulation. FaMYB6-like encodes a nuclear localization protein, which was clustered into the FaMYB1-like type repressor clade in phylogenetic analysis. FaMYB6-like showed high expressions in anthocyanin-accumulating fruit and low transcript abundance in flowers, vegetative tissues, and early development stages of fruit. Transient overexpression of FaMYB6L-like repressed the expression of anthocyanin biosynthetic and transport genes, and reduced anthocyanin content in postharvest strawberry fruit. Transgenic tobacco lines overexpressing FaMYB6-like displayed a reduction in petal pigmentation. Overexpression of FaMYB6L-like in white-fleshed strawberry inhibited anthocyanin accumulation induced by FaMYB10. Protein interaction and promoter activation assays revealed that FaMYB6-like repressed the activation of target gene promoters by FaMYB10-FabHLH3 activation complex via its interaction with FabHLH3. Additionally, FaMYB6-like promoter activity can be activated by FaMYB10-FabHLH3 complex. Taken together, our study revealed that FaMYB6-like acted as a transcriptional repressor involved in an ‘activator-and-repressor’ feedback regulation mechanism for maintaining the balance of anthocyanin concentrations.

The transcription factor RhMYB17 regulates the homeotic transformation of floral organs in rose (Rosa hybrida) under cold stress.

Low temperatures affect flower development in rose (Rosa hybrida), increasing petaloid stamen number and reducing normal stamen number. We identified the low-temperature-responsive R2R3-MYB transcription factor RhMYB17, which is homologous to Arabidopsis MYB17 by similarity of protein sequences. RhMYB17 was up-regulated at low temperatures, and RhMYB17 transcripts accumulated in floral buds. Transient silencing of RhMYB17 by virus-induced gene silencing decreased petaloid stamen number and increased normal stamen number. According to the ABCDE model of floral organ identity, class A genes APETALA 1 (AP1) and AP2 contribute to sepal and petal formation. Transcription factor binding analysis identified RhMYB17 binding sites in the promoters of rose APETALA 2 (RhAP2) and APETALA 2-LIKE (RhAP2L). Yeast one-hybrid assays, dual-luciferase reporter assays, and electrophoretic mobility shift assays confirmed that RhMYB17 directly binds to the promoters of RhAP2 and RhAP2L, thereby activating their expression. RNA sequencing further demonstrated that RhMYB17 plays a pivotal role in regulating the expression of class A genes, and indirectly influences the expression of the class C gene. This study reveals a novel mechanism for the homeotic transformation of floral organs in response to low temperatures.

Absence of long-term balancing selection on variation in EuMYB3, an R2R3-MYB gene responsible for the anther-color polymorphism in Erythronium umbilicatum

Balancing selection has been shown to be common in plants for several different types of traits, such as self-incompatibility and heterostyly. Generally, for these traits balancing selection is generated by interactions among individuals or between individuals and other species (e.g., pathogens or pollinators). However, there are phenotypic polymorphisms in plants that do not obviously involve types of interactions that generate balancing selection. Little is known about the extent to which balancing selection also acts to preserve these polymorphisms. Here we ask whether balancing selection preserves an anther-color polymorphism in Erythronium umbilicatum (Liliaceae). We identified a major gene underlying this polymorphism. We then attempted to detect signatures of balancing selection on that gene by developing a new coalescence test for balancing selection. We found that variation in anther color is in large part caused by variation in a paralog of EuMYB3, an anthocyanin-regulating R2R3-MYB transcription factor. However, we found little evidence for balancing selection having acted historically on EuMYB3. Our results thus suggest that plant polymorphisms, especially those not involved in interactions that are likely to generate negative frequency-dependent selection, may reflect a transient state in which one morph will eventually be fixed by either genetic drift or directional selection. Our results also suggest that regulation of the anthocyanin pathway is more evolutionarily labile than is generally believed.

Comprehensive genome-wide analyses of R2R3-MYB transcription factors and tolerance to drought stress in Populus davidiana × Populus bolleana

Comprehensive genome-wide analyses of R2R3-MYB transcription factors and tolerance to drought stress in Populus davidiana × Populus bolleana

Chinese cherry CpMYB44-CpSPDS2 module regulates spermidine content and florescence in tobacco.

The flower bud differentiation plays a crucial role in cherry yield and quality. In a preliminary study, we revealed the promotion of spermidine (Spd) in bud differentiation and quality. However, the molecular mechanism underlying Spd regulating cherry bud differentiation remains unclear. To address this research gap, we cloned CpSPDS2, a gene that encodes Spd synthase and is highly expressed in whole flowers and pistils of the Chinese cherry (cv. 'Manaohong'). Furthermore, an overexpression vector with this gene was constructed to transform tobacco plants. The findings demonstrated that transgenic lines exhibited higher Spd content, an earlier flowering time by 6 d, and more lateral buds and flowers than wild-type lines. Additionally, yeast one-hybrid assays and two-luciferase experiments confirmed that the R2R3-MYB transcription factor (CpMYB44) directly binds to and activates the CpSPDS2 promoter transcription. It is indicated that CpMYB44 promotes Spd accumulation via regulating CpSPDS2 expression, thus accelerating the flower growth. This research provides a basis for resolving the molecular mechanism of CpSPDS2 involved in cherry bud differentiation.

Soybean flower-specific R2R3-MYB transcription factor gene GmMYB108 induces anthocyanin production in Arabidopsis thaliana

R2R3-MYB transcription factors (TFs) are known to play a key role in regulating the expression of structural genes involved in plant flavonoid biosynthesis. However, the regulatory networks and related genes controlling isoflavonoid biosynthesis in soybean are poorly understood. We previously reported that ethephon application increases the production of isoflavonoids in soybean leaves. In this study, we attempted to identify a potential regulatory gene that positively controls isoflavonoid production in response to ethephon treatment in soybean (Glycine max L.). RNA sequencing (RNA-seq) revealed that ethephon application led to the upregulation of 22 genes, including the genes for R2R3-MYB TFs, related to isoflavonoid biosynthesis in soybean plants. Ethephon treatment highly induced the expression of GmMYB108, and its expression was exclusively enriched in flowers as determined using in silico and real-time quantitative PCR analyses. Furthermore, GmMYB108 overexpression resulted in an intense accumulation of anthocyanins as well as total flavonoid production in the leaf tissues of transgenic Arabidopsis plants. In addition, GmMYB108 overexpression increased the transcript levels of several genes involved in the biosynthesis of anthocyanins and their regulatory pathways in Arabidopsis. These results suggest that GmMYB108 is a potential positive regulator of the biosynthesis of flavonoids and anthocyanins in soybean flowers.

A novel transcription factor FnMYB4 regulates pigments metabolism of yellow leaf mutants in Fragaria nilgerrensis

The strawberry species Fragaria nilgerrensis Schlechtendal ex J. Gay, renowned for its distinctive white, fragrant peach-like fruits and strong disease resistance, is an exceptional research material. In a previous study, an ethyl methane sulfonate (EMS) mutant library was established for this species, resulting in various yellow leaf mutants. Leaf yellowing materials are not only the ideal materials for basic studies on photosynthesis mechanism, chloroplast development, and molecular regulation of various pigments, but also have important utilization value in ornamental plants breeding. The present study focused on four distinct yellow leaf mutants: mottled yellow leaf (MO), yellow green leaf (YG), light green leaf (LG), and buddha light leaf (BU). The results revealed that the flavonoid content and carotenoid-to-chlorophyll ratio exhibited a significant increase among these mutants, while experiencing a significant decrease in chlorophyll and carotenoid contents compared to the wild type (WT). To clarify the regulatory mechanisms and network relationships underlying these mutants, the RNA-seq and weighted gene co-expression network (WGCNA) analyses were employed. The results showed flavonoid metabolism pathway was enriched both in MO and YG mutants, while the chlorophyll biosynthesis pathway and carotenoid degradation pathway were only enriched in MO and YG mutants, respectively. Subsequently, key structural genes and transcription factors were identified on metabolic pathways of three pigments through correlation analyses and quantitative experiments. Furthermore, a R2R3-MYB transcription factor, FnMYB4, was confirmed to be positively correlated with flavonoid synthesis through transient overexpression, virus-induced gene silencing (VIGS), and RNA interference (RNAi), accompanying by reoccurrence and attenuation of mutant phenotype. Finally, dual-luciferase (LUC) and yeast one-hybrid assays confirmed the binding of FnMYB4 to the FnFLS and FnF3H promoters, indicating that FnMYB4 positively regulates flavonoid synthesis. In addition, correlation analyses suggested that FnMYB4 also might be involved in chlorophyll and carotenoid metabolisms. These findings demonstrated the pivotal regulatory role of FnMYB4 in strawberry leaf coloration.

Transcriptome-Based Identification of the SaR2R3-MYB Gene Family in Sophora alopecuroides and Function Analysis of SaR2R3-MYB15 in Salt Stress Tolerance.

As one of the most prominent gene families, R2R3-MYB transcription factors significantly regulate biochemical and physiological processes under salt stress. However, in Sophora alopecuroides, a perennial herb known for its exceptional saline alkali resistance, the comprehensive identification and characterization of SaR2R3-MYB genes and their potential functions in response to salt stress have yet to be determined. We investigated the expression profiles and biological functions of SaR2R3-MYB transcription factors in response to salt stress, utilizing a transcriptome-wide mining method. Our analysis identified 28 SaR2R3-MYB transcription factors, all sharing a highly conserved R2R3 domain, which were further divided into 28 subgroups through phylogenetic analysis. Some SaR2R3-MYB transcription factors showed induction under salt stress, with SaR2R3-MYB15 emerging as a potential regulator based on analysis of the protein-protein interaction network. Validation revealed the transcriptional activity and nuclear localization of SaR2R3-MYB15. Remarkably, overexpression of SaR2R3-MYB15 in transgenic plants could increase the activity of antioxidant enzymes and the accumulation of proline but decrease the content of malondialdehyde (MDA), compared with wild-type plants. Moreover, several salt stress-related genes showed higher expression levels in transgenic plants, implying their potential to enhance salt tolerance. Our findings shed light on the role of SaR2R3-MYB genes in salt tolerance in S. alopecuroides.

OsMYB84, a transcriptional regulator of OsCOPT2 and OsHMA5, modulates copper uptake and transport and yield production in rice

Transcription factors regulating crop uptake and translocation of the micronutrient Cu have not been identified. We isolated a novel R2R3-MYB transcription factor, OsMYB84, and showed that it was a positive regulator involved in uptake and transport of Cu via activation of OsCOPT2 and OsHMA expression. OsMYB84 was highly expressed in roots and anthers and induced by Cu. Overexpression of OsMYB84 promoted uptake and root-to-shoot translocation of Cu in rice, facilitated Cu distribution into grain and increased grain yield. In contrast, mutation of OsMYB84 reduced Cu concentration in xylem sap. OsMYB84 bound to the promoter region of OsCOPT2 and OsHMA5 and upregulated their expression. OsCOPT2 mutants showed reduced uptake of Cu and OsHMA5 overexpression lines showed increased root-to-shoot translocation of Cu.

Ectopic Expression of a Wheat R2R3-Type MYB Gene in Transgenic Tobacco Enhances Osmotic Stress Tolerance via Maintaining ROS Balance and Improving Root System Architecture.

Water scarcity is a critical cause of plant yield loss and decreased quality. Manipulation of root system architecture to minimize the impact of water scarcity stresses may greatly contribute towards an improved distribution of roots in the soil and enhanced water and nutrient uptake abilities. In this study, we explored the potential of TaMYB20 gene, a wheat gene belonging to the R2R3-MYB transcription factor family, to improve root system architecture in transgenic tobacco plants. The full-length TaMYB20 gene was isolated from Triticum aestivum.cv. Sakha94 and used to produce genetically engineered tobacco plants. The transgenic plants exhibited enhanced tolerance to extended osmotic stress and were able to maintain their root system architecture traits, including total root length (TRL), lateral root number (LRN), root surface area (RSa), and root volume (RV), while the wild-type plants failed to maintain the same traits. The transgenic lines presented greater relative water content in their roots associated with decreased ion leakage. The oxidative stress resulted in the loss of mitochondrial membrane integrity in the wild-type (WT) plants due to the overproduction of reactive oxygen species (ROS) in the root cells, while the transgenic lines were able to scavenge the excess ROS under stressful conditions through the activation of the redox system. Finally, we found that the steady-state levels of three PIN gene transcripts were greater in the TaMYB20-transgenic lines compared to the wild-type tobacco. Taken together, these findings confirm that TaMYB20 is a potentially useful gene candidate for engineering drought tolerance in cultivated plants.

OsMYB58 Negatively Regulates Plant Growth and Development by Regulating Phosphate Homeostasis.

Phosphate (Pi) starvation is a critical factor limiting crop growth, development, and productivity. Rice (Oryza sativa) R2R3-MYB transcription factors function in the transcriptional regulation of plant responses to various abiotic stresses and micronutrient deprivation, but little is known about their roles in Pi starvation signaling and Pi homeostasis. Here, we identified the R2R3-MYB transcription factor gene OsMYB58, which shares high sequence similarity with AtMYB58. OsMYB58 expression was induced more strongly by Pi starvation than by other micronutrient deficiencies. Overexpressing OsMYB58 in Arabidopsis thaliana and rice inhibited plant growth and development under Pi-deficient conditions. In addition, the overexpression of OsMYB58 in plants exposed to Pi deficiency strongly affected root development, including seminal root, lateral root, and root hair formation. Overexpressing OsMYB58 strongly decreased the expression of the rice microRNAs OsmiR399a and OsmiR399j. By contrast, overexpressing OsMYB58 strongly increased the expression of rice PHOSPHATE 2 (OsPHO2), whose expression is repressed by miR399 during Pi starvation signaling. OsMYB58 functions as a transcriptional repressor of the expression of its target genes, as determined by a transcriptional activity assay. These results demonstrate that OsMYB58 negatively regulates OsmiR399-dependent Pi starvation signaling by enhancing OsmiR399s expression.

PtoMYB031, the R2R3 MYB transcription factor involved in secondary cell wall biosynthesis in poplar.

The biosynthesis of the secondary cell wall (SCW) is orchestrated by an intricate hierarchical transcriptional regulatory network. This network is initiated by first-layer master switches, SCW-NAC transcription factors, which in turn activate the second-layer master switches MYBs. These switches play a crucial role in regulating xylem specification and differentiation during SCW formation. However, the roles of most MYBs in woody plants are yet to be fully understood. In this study, we identified and isolated the R2R3-MYB transcription factor, PtoMYB031, from Populus tomentosa. We explored its expression, mainly in xylem tissues, and its role as a transcriptional repressor in the nucleus. We used overexpression and RNA interference techniques in poplar, along with Yeast two-hybrid (Y2H) and bimolecular fluorescence complementation (BiFC) assays, to analyze the regulatory effects of PtoMYB031. Overexpression of PtoMYB031 in poplar significantly reduced lignin, cellulose, and hemicellulose content, and inhibited vascular development in stems, resulting in decreased SCW thickness in xylem tissues. Gene expression analysis showed that structural genes involved in SCW biosynthesis were downregulated in PtoMYB031-OE lines. Conversely, RNA interference of PtoMYB031 increased these compounds. Additionally, PtoMYB031 was found to recruit the repressor PtoZAT11, forming a transcriptional inhibition complex. Our findings provide new insights into how PtoMYB031, through its interaction with PtoZAT11, forms a complex that can suppress the expression of key regulatory genes, PtoWND1A and PtoWND2B, in SCW biosynthesis. This study enhances our understanding of the transcriptional regulation involved in SCW formation in poplar, highlighting the significant role of PtoMYB031.

A novel R2R3-MYB transcription factor PbMYB1L of Pyrus bretschneideri regulates cold tolerance and anthocyanin accumulation.

PbMYB1L enhances the cold tolerance and anthocyanin accumulation of transgenic Arabidopsis by regulating the expression of genes related to the cold-responsive genes pathway and anthocyanin synthesis pathway. MYB transcription factors (TFs) have been demonstrated to play diverse roles in plant growth and development. In the present study, we identified a novel R2R3-MYB transcription factor, PbMYB1L, from the peel of 'Red Zaosu' pear (Pyrus bretschneideri), which was induced by cold stress and acted as a positive regulator in anthocyanin biosynthesis. Notably, the transgenic Arabidopsis lines exhibited enhanced tolerance to cold stress. Compared to the Arabidopsis wild-type plants, the transgenic lines displayed longer primary roots and reduced reactive oxygen species (ROS) levels including O2-, hydrogen peroxide (H2O2), and malondialdehyde (MDA). Furthermore, significant upregulation of key cold-responsive genes AtCBF1, AtCBF2, AtCBF3, AtCBF4, and AtKIN1 was observed in the transgenic plants under cold stress conditions compared to wild type. Arabidopsis plants overexpressing PbMYB1L had significant anthocyanin accumulation in leaves after cold treatment with quantitative results indicating higher expression of anthocyanin structural genes compared to wild type. These findings suggest that PbMYB1L not only plays a vital role in conferring cold tolerance but also acts as a crucial regulator of anthocyanin biosynthesis.

LsMybW-encoding R2R3-MYB transcription factor is responsible for a shift from black to white in lettuce seed

Key messageWe identified LsMybW as the allele responsible for the shift in color from black to white seeds in wild ancestors of lettuce to modern cultivars.Successfully selected white seeds are a key agronomic trait for lettuce cultivation and breeding; however, the mechanism underlying the shift from black—in its wild ancestor—to white seeds remains uncertain. We aimed to identify the gene/s responsible for white seed trait in lettuce. White seeds accumulated less proanthocyanidins than black seeds, similar to the phenotype observed in Arabidopsis TT2 mutants. Genetic mapping of a candidate gene was performed with double-digest RAD sequencing using an F2 population derived from a cross between “ShinanoPower” (white) and “Escort” (black). The white seed trait was controlled by a single recessive locus (48.055–50.197 Mbp) in linkage group 7. Using five PCR-based markers and numerous cultivars, eight candidate genes were mapped in the locus. Only the LG7_v8_49.251Mbp_HinfI marker, employing a single-nucleotide mutation in the stop codon of Lsat_1_v5_gn_7_35020.1, was completely linked to seed color phenotype. In addition, the coding region sequences for other candidate genes were identical in the resequence analysis of “ShinanoPower” and “Escort.” Therefore, we proposed Lsat_1_v5_gn_7_35020.1 as the candidate gene and designated it as LsMybW (Lactuca sativaMyb White seeds), an ortholog encoding the R2R3-MYB transcription factor in Arabidopsis. When we validated the role of LsMybW through genome editing, LsMybW knockout mutants harboring an early termination codon showed a change in seed color from black to white. Therefore, LsMybW was the allele responsible for the shift in seed color. The development of a robust marker for marker-assisted selection and identification of the gene responsible for white seeds have implications for future breeding technology and physiological analysis.

A stress-responsive R2R3-MYB transcription factor, EpMYB1 is involved in the regulation of anthocyanin biosynthesis in purple coneflower

Purple coneflower (Echinacea purpurea (L.) Menoch), a North American native and world-popular medicinal and horticultural plant, has been named after its beautiful purple petals due to the accumulation of anthocyanins. However, so far the biosynthesis of anthocyanin and its regulation in purple coneflower remain elusive. In this study, the total anthocyanin contents of eight different purple coneflower tissues were measured, and anthocyanin biosynthetic genes were identified from the multi-tissue transcriptome. Using co-expression, phylogenetic screening, and transient functional verification, EpMYB1 was identified as a potential anthocyanin biosynthesis regulator. EpMYB1 is an R2R3-MYB TF and co-expressed with anthocyanin biosynthetic genes. When EpMYB1 was over-expressed in purple coneflower calli, it positively promoted anthocyanin biosynthesis by upregulating the anthocyanin-specific biosynthetic genes. Dual-luciferase reporter assay indicated that EpMYB1 could significantly activate anthocyanin biosynthetic gene expression by directly binding to the promoters. Moreover, EpMYB1 is a stress-responsive TF involved in the ultraviolet-induced anthocyanin biosynthesis in purple coneflower. The present study identifies an MYB TF that participated in anthocyanin biosynthesis and bridged the gap between environmental stress and anthocyanin biosynthesis in purple coneflower. It provides valuable insights into the anthocyanin biosynthesis in purple coneflower and will significantly improve the development of purple coneflower as an ornament plant.

MdBT2 regulates nitrogen-mediated cuticular wax biosynthesis via a MdMYB106-MdCER2L1 signalling pathway in apple.

Cuticular waxes play important roles in plant development and the interaction between plants and their environment. Researches on wax biosynthetic pathways have been reported in several plant species. Also, wax formation is closely related to environmental condition. However, the regulatory mechanism between wax and environmental factors, especially essential mineral elements, is less studied. Here we found that nitrogen (N) played a negative role in the regulation of wax synthesis in apple. We therefore analysed wax content, composition and crystals in BTB-TAZ domain protein 2 (MdBT2) overexpressing and antisense transgenic apple seedlings and found that MdBT2 could downregulate wax biosynthesis. Furthermore, R2R3-MYB transcription factor 16-like protein (MdMYB106) interacted with MdBT2, and MdBT2 mediated its ubiquitination and degradation through the 26S proteasome pathway. Finally, HXXXD-type acyl-transferase ECERIFERUM 2-like1 (MdCER2L1) was confirmed as a downstream target gene of MdMYB106. Our findings reveal an N-mediated apple wax biosynthesis pathway and lay a foundation for further study of the environmental factors associated with wax regulatory networks in apple.

Does an R2R3-MYB transcription factor affect anthurium spathe colour variation via regulation of vacuolar pH?

The anthurium is a popular cut flower worldwide having red, pink, coral, white, green, or brown spathes. There is a constant demand for new flower colours in the commercial market. Flower colour in plants is mainly determined by anthocyanins. Understanding anthocyanin variation and other factors affecting anthurium spathe colour is important for genetic engineering approaches. Therefore, our objectives were to assess the factors affecting colour variation of selected commercially available cut flower anthuriums and to determine the associated regulatory networks and transcription factors (TFs). Nineteen commercial cut flower anthurium cultivars were selected for this purpose. The colour of the spathe surface, anthocyanin location, anthocyanidin type and vacuolar pH were recorded. Anthocyanin associated Gene Network Model generation and analysis were carried out. The CIELAB colourimeter procedure indicated the colour variation among the selected 19 cultivars in terms of colour type, colour intensity, chroma, and hue angle. The location of anthocyanin was limited to mesophyll and epidermal cells. Cyanidin was detected in tested anthurium cultivars as the main anthocyanidin. The pH gradient in pigment extracts indicated a variation with a range of 4.6 to 4.94. The gene pathways of anthocyanin biosynthesis and transport were associated with that of the vacuolar pH/H+ pump according to Gene Network Model. Three pathways were regulated by an R2R3-MYB transcription factor. Although, cyanidin was the only pigment in all the tested cultivars, different pH levels by R2R3-MYB regulated V-H+ synthase was suggested to be the cause of the high colour variation in addition to the anthocyanidin type and location. Our results indicate the application of R2R3-MYB transcription factor genes for desirable vacuolar pH maintenance in genetic engineering of the blue anthurium in the future.

A metabolomics study in citrus provides insight into bioactive phenylpropanoid metabolism.

Abstract Citrus fruits are widely consumed worldwide in juices or as fresh and provide a broad range of phytonutrients that are important for human health. Here, a citrus multi-omics resource is presented: comprehensive metabolic profiling of various citrus species was performed and metabolic profiles were compared among species, with a focus on the phenylpropanoid metabolic pathway. A metabolite-based genome-wide association analysis (mGWAS) of 154 pummelo accessions was performed using factored spectrally transformed linear mixed models (FaST-LMM) and efficient mixed-model association eXpedited (EMMAX), and the genetic and biochemical basis of metabolomic variation was comprehensively analysed. A metabolite-single nucleotide polymorphism-gene (metabolite-SNP-gene) interaction network was constructed for pummelo, and many candidate loci controlling the synthesis and regulation of bioactive compounds were identified; among these loci, three BAHD malonyltransferases were involved in the malonylation of flavonoid glycosides. Further investigation revealed that an R2R3-MYB transcription factor CgMYB1 positively controls the metabolism of phenylpropanoid molecules, particularly the flavonoid derivatives. This study provides valuable data resources on the metabolic regulatory networks of bioactive components in citrus, in addition to demonstrating an efficient method for metabolic pathway dissection and providing targets for future breeding work with the aim of improving nutritional value.

MdMYB44-like positively regulates salt and drought tolerance via the MdPYL8-MdPP2CA module in apple.

Abscisic acid (ABA) is involved in salt and drought stress responses, but the underlying molecular mechanism remains unclear. Here, we demonstrated that the overexpression of MdMYB44-like, an R2R3-MYB transcription factor, significantly increases the salt and drought tolerance of transgenic apples and Arabidopsis. MdMYB44-like inhibits the transcription of MdPP2CA, which encodes a type 2C protein phosphatase that acts as a negative regulator in the ABA response, thereby enhancing ABA signaling-mediated salt and drought tolerance. Furthermore, we found that MdMYB44-like and MdPYL8, an ABA receptor, form a protein complex that further enhances the transcriptional inhibition of the MdPP2CA promoter by MdMYB44-like. Significantly, we discovered that MdPP2CA can interfere with the physical association between MdMYB44-like and MdPYL8 in the presence of ABA, partially blocking the inhibitory effect of the MdMYB44-like-MdPYL8 complex on the MdPP2CA promoter. Thus, MdMYB44-like, MdPYL8, and MdPP2CA form a regulatory loop that tightly modulates ABA signaling homeostasis under salt and drought stress. Our data reveal that MdMYB44-like precisely modulates ABA-mediated salt and drought tolerance in apples through the MdPYL8-MdPP2CA module.

PagMYB73 enhances salt stress tolerance by regulating reactive oxygen species scavenging and osmotic maintenance in poplar

The MYB transcription factor (TF) family, widely recognized for its involvement in multiple biological processes, holds a significant position among plant TF families due to its substantial size. The R2R3-MYB TF has been extensively studied and known for its pivotal role in regulating the synthesis of anthocyanin, lignin, and secondary wall components in plants. However, there is currently limited information available regarding the function of R2R3-MYB genes in response to salt stress specifically in poplar. Here, we focused on a specific R2R3-MYB gene, PagMYB73, which belongs to the 84K poplar (Populus alba × P. glandulosa) species. We aimed to investigate the role of PagMYB73 in the regulatory mechanisms involved in salt stress response. PagMYB73 showed strong induction in leaves under salt stress conditions. Transgenic poplar plants overexpressing PagMYB73 exhibited enhanced salt tolerance, while suppression of the gene resulted in increased sensitivity to high salt levels. PagMYB73 can regulate the expression of several stress-related genes, including PagGSTU19, PagLEA51, PagSODs, PagPODs, and PagP5CS2. It achieves this by binding to the promoter regions of these genes that orchestrate adaptive responses to salt stress by modulating the osmotic balance and reducing the levels of reactive oxygen species (ROS). Indeed, overexpression of PagGSTU19 and PagLEA51 has been shown to enhance the salt tolerance of poplar. In conclusion, PagMYB73 regulates the expression of stress-related genes to improve ROS scavenging ability and osmotic maintenance, thereby improving the salt tolerance of the transgenic poplar.