ABSTRACT
Stress induced by ultraviolet-B (UV-B) irradiation stimulates the accumulation of various secondary metabolites in plants. Nitric oxide (NO) serves as an important secondary messenger in UV-B stress-induced signal transduction pathways. NO can be synthesized in plants by either enzymatic catalysis or an inorganic nitrogen pathway. The effects of UV-B irradiation on the production of baicalin and the associated molecular pathways in plant cells are poorly understood. In this study, nitric oxide synthase (NOS) activity, NO release and the generation of baicalin were investigated in cell suspension cultures of Scutellaria baicalensis exposed to UV-B irradiation. UV-B irradiation significantly increased NOS activity, NO release and baicalin biosynthesis in S. baicalensis cells. Additionally, exogenous NO supplied by the NO donor, sodium nitroprusside (SNP), led to a similar increase in the baicalin content as the UV-B treatment. The NOS inhibitor, Nω-nitro-l-arginine (LNNA), and NO scavenger, 2-(4-carboxyphenyl)-4,4,5,5-tetramethylimidazoline-1-oxyl-3-oxide (cPTIO) partially inhibited UV-B-induced NO release and baicalin accumulation. These results suggest that NO is generated by NOS or NOS-like enzymes and plays an important role in baicalin biosynthesis as part of the defense response of S. baicalensis cells to UV-B irradiation.
Subject(s)
Flavonoids/metabolism , Nitric Oxide/metabolism , Signal Transduction/radiation effects , Ultraviolet Rays , Benzoates/pharmacology , Cells, Cultured , Enzyme Inhibitors/pharmacology , Flavonoids/chemistry , Imidazoles/pharmacology , Molecular Structure , Nitric Oxide Donors/pharmacology , Nitric Oxide Synthase/antagonists & inhibitors , Nitric Oxide Synthase/metabolism , Nitroarginine/pharmacology , Nitroprusside/pharmacology , Plant Cells/drug effects , Plant Cells/metabolism , Plant Cells/radiation effects , Scutellaria baicalensis/cytology , Signal Transduction/drug effects , Time FactorsABSTRACT
R2R3-MYB proteins play role in plant development, response to biotic and abiotic stress, and regulation of primary and secondary metabolism. Little is known about the R2R3-MYB proteins in Scutellaria baicalensis which is an important Chinese medical plant. In this paper, nineteen putative SbMYB genes were identified from a S. baicalensis cDNA library, and eleven R2R3-MYBs were clustered into 5 subgroups according to phylogenetic reconstruction. In the S. baicalensis leaves which were sprayed with GA3, SbMYB2 and SbMYB7 had similar expression pattern with SbPALs, indicating that SbMYB2 and SbMYB7 might be involved in the flavonoid metabolism. Transactivation assay results showed that SbMYB2 and SbMYB7 can function as transcriptional activator. The expression of several flavonoid biosynthesis-related genes were induced or suppressed by overexpression of SbMYB2 or SbMYB7 in transgenic tobacco plants. Consistent with the change of the expression of NtDH29 and NtCHI, the contents of dicaffeoylspermidine and quercetin-3,7-O-diglucoside in SbMYB2-overexpressing or SbMYB7-overexpressing transgenic tobacco plants were decreased. The transcriptional level of NtUFGT in transgenic tobacco overexpressing SbMYB7 and the transcriptional level of NtHCT in SbMYB2-overexpressing tobacco plants were increased; however the application of GA3 inhibited the transcriptional level of these two genes. These results suggest that SbMYB2 and SbMYB7 might regulate the flavonoid biosynthesis through GA metabolism.
Subject(s)
Flavonoids/biosynthesis , Nicotiana/genetics , Nicotiana/metabolism , Plant Proteins/metabolism , Scutellaria baicalensis/genetics , Scutellaria baicalensis/metabolism , Transcription Factors/metabolism , Cloning, Molecular , Gene Expression Regulation, Plant/drug effects , Gibberellins/metabolism , Gibberellins/pharmacology , Intracellular Space/metabolism , Phenylalanine Ammonia-Lyase/genetics , Phylogeny , Plant Proteins/genetics , Plants, Genetically Modified , Promoter Regions, Genetic/genetics , Protein Transport , Scutellaria baicalensis/cytology , Spermidine/biosynthesis , Transcription Factors/genetics , Transcriptional Activation/drug effectsABSTRACT
An approach of combining flow cytometry (FCM) analysis with morphological and chemical profiling was used to assess the genetic stability and bioactive compound diversity in a Scutellaria baicalensis Georgi (Huang-qin) germplasm collection that was clonally maintained in in vitro for a period of over 6 years. Based on the FCM analysis of nuclei samples from young shoots, the nuclear DNA content of S. baicalensis was calculated as 0.84 pg/2C. FCM analysis showed no significant variation in the nuclear DNA contents and ploidy levels in the long-term in vitro maintained germplasm lines. Germplasm lines, acclimatized to ex vitro conditions, exhibited distinctive plant growth and bioactive compound production capacities. The high level of genetic stability observed in in vitro maintained S. baicalensis lines opens up a variety of opportunities such as allowing long-term aseptic preservation and easy distribution of well-characterized germplasm lines of this medicinal plant species. This study represents a novel approach for continuous maintenance, monitoring, and production of medicinal plant tissues with specific chemistry.