TRANSCRIPTIONAL REGULATION OF SPECIALIZED METABOLITES IN \u3cem\u3eARABIDOPSIS THALIANA\u3c/em\u3e AND \u3cem\u3eCATHARANTHUS ROSEUS\u3c/em\u3e

For millennia humans have utilized plant specialized metabolites for health benefits, fragrances, poisons, spices, and medicine. Valued metabolites are often produced in small quantities and may command high prices. Understanding when and how the plant synthesizes these compounds is important for improving their production. Phytohormone signaling cascades, such as jasmonate (JA) activate or repress transcription factors (TF) controlling expression of metabolite biosynthetic genes. TFs regulating specialized metabolite biosynthetic genes can be manipulated to engineer plants with increased metabolite production. WRKY transcription factor are known components of both JA signaling cascades and regulation of specialized metabolism. The presence of WRKY binding sites in promoters of several terpene indole alkaloids suggested their involvement in regulating biosynthesis of these compounds. A phylogenetic analysis was used to compare Arabidopsis and Catharanthus WRKY TFs families. Gene expression analysis identified WRKY TFs induced by JA in both Arabidopsis and Catharanthus, providing candidates for future characterization. WRKY TFs suggest a possible conserved regulatory network of TFs downstream of JA signaling cascades. The origin and conservation of JA signaling in plants remain ambiguous. Identification of the first algal TIFY factor helped determine when JA signaling appeared. The charophyte, Klebsormidium flaccidum does not possess genes encoding key green-plant JA signaling components, including CORONATINE INSENSTIVE1, JASMONATE-ZIM DOMAIN, NOVEL INTERACTOR OF JAZ, and the JAZ-interacting bHLH factors, yet their orthologs are present in the moss. A molecular clock analysis dated the evolution of JA signaling evolution to during the early Ediacaran to late Cambrian periods 628 to 491 million years ago – a time corresponding to rapid diversification of animal predators. The plant Mediator complex is a core component of gene expression. Conservation of the MED25 subunit in plants, and its known involvement in JA signaling implicates this factor in regulation of specialized metabolism. MED25 is involved in anthocyanin accumulation, but how it functions remains unknown. Characterization of MED25 in Arabidopsis revealed it interacts with the transcription factor GL3 as well as the JAZ1 repressor. Importantly, the interaction of JAZ1 with MED25 reveals a new mechanism by which JAZ proteins regulate gene expression, improving our understanding of JA signaling

Transcription Factors in Alkaloid Engineering

Plants produce a large variety of low-molecular-weight and specialized secondary compounds. Among them, nitrogen-containing alkaloids are the most biologically active and are often used in the pharmaceutical industry. Although alkaloid chemistry has been intensively investigated, characterization of alkaloid biosynthesis, including biosynthetic enzyme genes and their regulation, especially the transcription factors involved, has been relatively delayed, since only a limited number of plant species produce these specific types of alkaloids in a tissue/cell-specific or developmental-specific manner. Recent advances in molecular biology technologies, such as RNA sequencing, co-expression analysis of transcripts and metabolites, and functional characterization of genes using recombinant technology and cutting-edge technology for metabolite identification, have enabled a more detailed characterization of alkaloid pathways. Thus, transcriptional regulation of alkaloid biosynthesis by transcription factors, such as basic helix–loop–helix (bHLH), APETALA2/ethylene-responsive factor (AP2/ERF), and WRKY, is well elucidated. In addition, jasmonate signaling, an important cue in alkaloid biosynthesis, and its cascade, interaction of transcription factors, and post-transcriptional regulation are also characterized and show cell/tissue-specific or developmental regulation. Furthermore, current sequencing technology provides more information on the genome structure of alkaloid-producing plants with large and complex genomes, for genome-wide characterization. Based on the latest information, we discuss the application of transcription factors in alkaloid engineering

Analyses of Catharanthus Roseus and Arabidopsis Thaliana WRKY Transcription Factors Reveal Involvement in Jasmonate Signaling

BACKGROUND: To combat infection to biotic stress plants elicit the biosynthesis of numerous natural products, many of which are valuable pharmaceutical compounds. Jasmonate is a central regulator of defense response to pathogens and accumulation of specialized metabolites. Catharanthus roseus produces a large number of terpenoid indole alkaloids (TIAs) and is an excellent model for understanding the regulation of this class of valuable compounds. Recent work illustrates a possible role for the Catharanthus WRKY transcription factors (TFs) in regulating TIA biosynthesis. In Arabidopsis and other plants, the WRKY TF family is also shown to play important role in controlling tolerance to biotic and abiotic stresses, as well as secondary metabolism. RESULTS: Here, we describe the WRKY TF families in response to jasmonate in Arabidopsis and Catharanthus. Publically available Arabidopsis microarrays revealed at least 30% (22 of 72) of WRKY TFs respond to jasmonate treatments. Microarray analysis identified at least six jasmonate responsive Arabidopsis WRKY genes (AtWRKY7, AtWRKY20, AtWRKY26, AtWRKY45, AtWRKY48, and AtWRKY72) that have not been previously reported. The Catharanthus WRKY TF family is comprised of at least 48 members. Phylogenetic clustering reveals 11 group I, 32 group II, and 5 group III WRKY TFs. Furthermore, we found that at least 25% (12 of 48) were jasmonate responsive, and 75% (9 of 12) of the jasmonate responsive CrWRKYs are orthologs of AtWRKYs known to be regulated by jasmonate. CONCLUSION: Overall, the CrWRKY family, ascertained from transcriptome sequences, contains approximately 75% of the number of WRKYs found in other sequenced asterid species (pepper, tomato, potato, and bladderwort). Microarray and transcriptomic data indicate that expression of WRKY TFs in Arabidopsis and Catharanthus are under tight spatio-temporal and developmental control, and potentially have a significant role in jasmonate signaling. Profiling of CrWRKY expression in response to jasmonate treatment revealed potential associations with secondary metabolism. This study provides a foundation for further characterization of WRKY TFs in jasmonate responses and regulation of natural product biosynthesis

Suberin Biosynthesis and Deposition in the Wound-Healing Potato (Solanum tuberosum L.) Tuber Model

Suberin is a heteropolymer comprising a cell wall-bound poly(phenolic) domain (SPPD) covalently linked to a poly(aliphatic) domain (SPAD) that is deposited between the cell wall and plasma membrane. Potato tuber skin contains suberin to protect against water loss and microbial infection. Wounding triggers suberin biosynthesis in usually non-suberized tuber parenchyma, providing a model system to study suberin production. Spatial and temporal coordination of SPPD and SPAD-related metabolism are required for suberization, as the former is produced soon after wounding, and the latter is synthesized later into wound-healing. Many steps involved in suberin biosynthesis remain uncharacterized, and the mechanism(s) that regulate and coordinate SPPD and SPAD production and assembly are not understood. To explore the role of abscisic acid (ABA) in the differential regulation of SPPD and SPAD biosynthesis, I subjected wounded tubers to exogenous treatments including additional ABA, or the ABA biosynthesis inhibitor fluridone. Quantitative reverse transcription polymerase chain reaction (RT-qPCR) expression analysis of SPPD and SPAD biosynthetic genes, coupled with metabolite analyses, revealed that ABA positively influenced SPAD-, but not SPPD-associated, transcript and metabolite accumulation, indicating a role for ABA in the differential induction of wound-induced phenolic and aliphatic metabolism. I took an RNA-seq approach to study broader transcriptional changes that occur during wound-healing. The wound-healing transcriptome time-course illustrated that wounding leads to a substantial reconfiguration of transcription, followed by fine-tuning of responses dominated by suberization. Transcriptome analysis revealed that primary metabolic pathways demonstrate similar temporal expression patterns during wound-healing, but suberin-specific steps display distinct patterns at entire pathway and sub-branch levels. The observed transcriptional changes support a model in which wounding initially alters primary metabolism required to fuel SPPD, and subsequent SPAD, production. This investigation also provided support for uncharacterized biosynthetic steps, and highlighted putative transcription factors and suberin polymer assembly genes (Casparian strip membrane domain proteins and GDSL lipase/esterases) that may play key roles in the regulation and coordination of SPPD and SPAD monomer biosynthesis, polymer assembly and deposition. Overall, my findings offer further insight into the coordination and timing of metabolic and regulatory events involved in wound-healing and associated suberization

Physiological responses and transcriptome analyses of upland rice following exposure to arsenite and arsenate

Acknowledgements This research was financially supported by the National Natural Science Foundation of China (No.41471274) and the Scottish Government’s Rural and Environment Science and Analytical Service Division (RESAS).Peer reviewedPostprin

Phosphate and zinc transport and signalling in plants: toward a better understanding of their homeostasis interaction

Zn and Pi are essential elements for plant growth. Current understanding of the regulation of their homeostasis interaction and signalling cross-talk is presente

The Strawberry FaWRKY1 Transcription Factor Negatively Regulates Resistance to Colletotrichum acutatum in Fruit Upon Infection

Strawberry (Fragaria ×ananassa) is a major food crop worldwide, due to the flavor, aroma and health benefits of the fruit, but its productivity and quality are seriously limited by a large variety of phytopathogens, including Colletotrichum spp. So far, key factors regulating strawberry immune response remain unknown. The FaWRKY1 gene has been previously proposed as an important element mediating defense responses in strawberry to Colletotrichum acutatum. To get further insight into the functional role that FaWRKY1 plays in the defense mechanism, Agrobacterium-mediated transient transformation was used both to silence and overexpress the FaWRKY1 gene in strawberry fruits (Fragaria ×ananassa cv. Primoris), which were later analyzed upon C. acutatum inoculation. Susceptibility tests were performed after pathogen infection comparing the severity of disease between the two agroinfiltrated opposite halves of the same fruit, one half bearing a construct either for FaWRKY1 overexpression or RNAi-mediated silencing and the other half bearing the empty vector, as control. The severity of tissue damage was monitored and found to be visibly reduced at five days after pathogen inoculation in the fruit half where FaWRKY1 was transiently silenced compared to that of the opposite control half and statistical analysis corroborated a significant reduction in disease susceptibility. Contrarily, a similar level of susceptibility was found when FaWRKY1 overexpression and control fruit samples, was compared. These results unravel a negative regulatory role of FaWRKY1 in resistance to the phytopathogenic fungus C. acutatum in strawberry fruit and contrast with the previous role described for this gene in Arabidopsis as positive regulator of resistance against the bacteria Pseudomonas syringae. Based on previous results, a tentative working model for WRKY75 like genes after pathogen infection is proposed and the expression pattern of potential downstream FaWRKY1 target genes was also analyzed in strawberry fruit upon C. acutatum infection. Our results highlight that FaWRKY1 might display different function according to species, plant tissue and/or type of pathogen and underline the intricate FaWRKY1 responsive defense regulatory mechanism taking place in strawberry against this important crop pathogen