Auxin-Responsive R2R3-MYB Transcription Factors HcMYB1 and HcMYB2 Activate Volatile Biosynthesis in Hedychium coronarium Flowers

Auxin, an important plant hormone, induces the biosynthesis of various secondary metabolites by modulating the expression of auxin-responsive genes. In the ornamental plant Hedychium coronarium, linalool and methyl benzoate are biosynthesized by the terpene synthase (TPS) HcTPS5 and the benzoic/salicylic acid methyltransferase (BSMT) HcBSMT2, respectively. However, the transcriptional regulation of this process remains unclear. Here, we identified and functionally characterized the R2R3-MYB transcription factors HcMYB1 and HcMYB2 in regulating the biosynthesis of these floral aroma compounds. HcMYB1 and HcMYB2 are specifically expressed in flowers, their expression is correlated with the emission of volatile compounds in flowers, and is induced by auxin. Moreover, HcMYB1 and HcMYB2 interact with the HcBSMT2 promoter region. HcMYB2 activates the expression of the linalool synthase gene HcTPS5. In flowers with HcMYB1 or HcMYB2 silenced, the levels of floral scent compounds were significantly reduced, and HcBSMT2 and HcTPS5 were downregulated compared with the wild type. Moreover, HcMYB1 form protein-protein interaction with key scent-related HcIAA4 protein to regulate floral aroma production. Taken together, these results indicate that HcMYB1 and HcMYB2 play crucial roles in regulating the formation of scent compounds in Hedychium coronarium (H. coronarium) flowers in response to auxin signaling.

The Seed Development Factors TT2 and MYB5 Regulate Heat Stress Response in Arabidopsis

HEAT SHOCK FACTOR A2 (HSFA2) is a regulator of multiple environmental stress responses required for stress acclimation. We analyzed HSFA2 co-regulated genes and identified 43 genes strongly co-regulated with HSFA2 during multiple stresses. Motif enrichment analysis revealed an over-representation of the site II element (SIIE) in the promoters of these genes. In a yeast 1-hybrid screen with the SIIE, we identified the closely related R2R3-MYB transcription factors TT2 and MYB5. We found overexpression of MYB5 or TT2 rendered plants heat stress tolerant. In contrast, tt2, myb5, and tt2/myb5 loss of function mutants showed heat stress hypersensitivity. Transient expression assays confirmed that MYB5 and TT2 can regulate the HSFA2 promoter together with the other members of the MBW complex, TT8 and TRANSPARENT TESTA GLABRA 1 (TTG1) and that the SIIE was involved in this regulation. Transcriptomic analysis revealed that TT2/MYB5 target promoters were enriched in SIIE. Overall, we report a new function of TT2 and MYB5 in stress resistance and a role in SIIE-mediated HSFA2 regulation.

R2R3-MYB transcription factors, StmiR858 and sucrose mediate potato flavonol biosynthesis

Flavonols and other phenylpropanoids protect plants from biotic and abiotic stress and are dietarily desirable because of their health-promoting properties. The ability to develop new potatoes (Solanum tuberosum) with optimal types and amounts of phenylpropanoids is limited by lack of knowledge about the regulatory mechanisms. Exogenous sucrose increased flavonols, whereas overexpression of the MYB StAN1 induced sucrolytic gene expression. Heterologous StAN1 protein bound promoter fragments from sucrolytic genes (SUSY1 and INV1). Two additional MYBs and one microRNA were identified that regulated potato flavonols. Overexpression analysis showed MYB12A and C increased amounts of flavonols and other phenylpropanoids. Endogenous flavonol amounts in light-exposed organs were much higher those in the dark. Expression levels of StMYB12A and C were high in flowers but low in tubers. Transient overexpression of miR858 altered potato flavonol metabolism. Endogenous StmiR858 expression was much lower in flowers than leaves and correlated with flavonol amounts in these organs. Collectively, these findings support the hypothesis that sucrose, MYBs, and miRNA control potato phenylpropanoid metabolism in a finely tuned manner that includes a feedback loop between sucrose and StAN1. These findings will aid in the development of potatoes with phenylpropanoid profiles optimized for crop performance and human health.

Expression pattern analysis of three R2R3-MYB transcription factors for the production of anthocyanin in different vegetative stages of Arabidopsis leaves

Anthocyanin is a type of flavonoid that appears purple in plants. PAP1, PAP2, and MYB113 are the three major R2R3-MYB transcription factors that regulate flavonoid biosynthesis in Arabidopsis thaliana. In this study, we found that the three MYB genes regulate anthocyanin accumulation in different leaf stages. Under limited nutrient conditions, PAP1 and PAP2 genes were highly induced in juvenile leaves. Conversely, MYB113 was expressed mainly in adult leaves. In addition, we investigated the role of trans-acting siRNA4 (TAS4) in the post-transcriptional regulation of anthocyanin expression in Arabidopsis leaves. In plant growth, the inhibition of PAP1 and PAP2 gene expression by TAS4 was observed only in juvenile leaves, and MYB113 inhibition was observed in adult leaves. In conclusion, we found that transcription and transcript repression of the three MYB genes is differentially regulated by TAS4 in leaf developmental stages. Our results improve the understanding of the regulation of plant anthocyanin production under stress conditions.

Origins and Stepwise Expansion of R2R3-MYB Transcription Factors for the Terrestrial Adaptation of Plants

The R2R3-MYB transcription factors play critical roles in various processes in embryophytes (land plants). Here, we identified genes encoding R2R3-MYB proteins from rhodophytes, glaucophytes, Chromista, chlorophytes, charophytes, and embryophytes. We classified the R2R3-MYB genes into three subgroups (I, II, and III) based on their evolutionary history and gene structure. The subgroup I is the most ancient group that includes members from all plant lineages. The subgroup II was formed before the divergence of charophytes and embryophytes. The subgroup III genes form a monophyletic group and only comprise members from land plants with conserved exon–intron structure. Each subgroup was further divided into multiple clades. The subgroup I can be divided into I-A, I-B, I-C, and I-D. The I-A, I-B, and I-C are the most basal clades that have originated before the divergence of Archaeplastida. The I-D with the II and III subgroups form a monophyletic group, containing only green plants. The II and III subgroups form another monophyletic group with Streptophyta only. Once on land, the subgroup III genes have experienced two rounds of major expansions. The first round occurred before the origin of land plants, and the second round occurred after the divergence of land plants. Due to significant gene expansion, the subgroup III genes have become the predominant group of R2R3-MYBs in land plants. The highly unbalanced pattern of birth and death evolution of R2R3-MYB genes indicates their important roles in the successful adaptation and massive radiation of land plants to occupy a multitude of terrestrial environments.

Phylogenomic Analysis of R2R3 MYB Transcription Factors in Sorghum and their Role in Conditioning Biofuel Syndrome

Background : Large scale cultivation of sorghum for food, feed, and biofuel requires concerted efforts for engineering multipurpose cultivars with optimised agronomic traits. Due to their vital role in regulating the biosynthesis of phenylpropanoid-derived compounds, biomass composition, biotic, and abiotic stress response, R2R3-MYB family transcription factors are ideal targets for improving environmental resilience and economic value of sorghum. Methods: We used diverse computational biology tools to survey the sorghum genome to identify R2R3-MYB transcription factors followed by their structural and phylogenomic analysis. We used inhouse generated as well as publicly available high throughput expression data to analyse the R2R3 expression patterns in various sorghum tissue types. Results: We have identified a total of 134 R2R3-MYB genes from sorghum and developed a framework to predict gene functions. Collating information from the physical location, duplication, structural analysis, orthologous sequences, phylogeny, and expression patterns revealed the role of duplications in clade-wise expansion of the R2R3-MYB family as well as intra-clade functional diversification. Using publicly available and in-house generated RNA sequencing data, we provide MYB candidates for conditioning biofuel syndrome by engineering phenylpropanoid biosynthesis and sugar signalling pathways in sorghum. Conclusion: The results presented here are pivotal to prioritize MYB genes for functional validation and optimize agronomic traits in sorghum.