Wheat TaMYB1A Negatively Regulates the Phenylpropanoid Pathway and Plant Height / 中国生物化学与分子生物学报

The phenylpropanoid pathway is one of the important pathways for synthesizing plant secondary metabolites, which can produce lignin, flavonoid, and sinapoylmalate. These compounds can not only affect the plant growth, development, and stress response, but also be used to produce perfume, pesticide, dye, medicine, feed, and biomass energy. R2R3-MYBs play important roles in regulating plant secondary metabolism, organ development, and in responding to environmental stresses. Wheat (Triticum aestivum L.) is an important food crop, but lots of straw will be produced accompanied by grain yields. Therefore, elucidating the function and regulatory mechanism of R2R3 MYBs of wheat is crucial for the effective utilization of the wheat straw. RT-PCR results showed that TaMYB1A was highly expressed in the wheat stems, and the GFP-TaMYB1A fusion protein was mainly localized in the nucleus of the N. benthamiana epidermal cells. TaMYB1A has transcriptional repressive activity in yeast cells. In this study, TaMYB1A-overexpressed transgenic Arabidopsis lines were generated to elucidate the effect of overexpression of TaMYB1A on the biosynthesis of lignin and flavonoid. Our results suggested that overexpression of TaMYB1A inhibited the plant height (P < 0. 05) and decreased the lignin (P < 0. 05) and flavonoid (P < 0. 05) biosynthesis of the transgenic Arabidopsis plants significantly. TaMYB1A could bind to the promoters of the Arabidopsis At4CL1, AtC4H, AtC3H, and AtCHS as well as the wheat Ta4CL1 and TaC4H1 revealed by yeast one-hybrid (Y1H) assasy, the transcriptional repressive effect of TaMYB1A on At4CL1, AtC4H, AtC3H, and AtCHS was confirmed by dual-luciferase reporter systems and also on Ta4CL1 and TaC4H1 by a genetic approach. Gene chip and quantitative RT-PCR (qRT-PCR) results showed that overexpression of TaMYB1A down-regulated the expression of most of the key genes involved in the phenylpropanoid metabolism and decreased the 4CL activity (P < 0. 05) of the transgenic Arabidopsis plants significantly. As suggested above, the wheat TaMYB1A belongs to the subgroup 4 R2R3 MYB transcription factors. TaMYB1A could bind to the promoters of the key genes involved in phenylpropanoid metabolism, repress their expression and negatively regulate the phenylpropanoid metabolism pathway and plant height.

Cloning and expression of AeMYB4 gene related to stolon development of Amana edulis / 中国中药杂志

Stolon is an important organ for reproduction and regeneration of Amana edulis. Previous analysis of transcriptome showed that MYB was one of the most active transcription factor families during the development of A. edulis stolon. In order to study the possible role of MYB transcription factors in stolon development, the authors screened out an up-regulated MYB gene named AeMYB4 was by analyzing the expression profile of MYB transcription factors. In the present study, sequence analysis demonstrated that AeMYB4 contained an open reading frame of 756 bp encoding 251 amino acids, and domain analysis revealed that the predicted amino acids sequence contained two highly conserved SANT domains and binding sites for cold stress factor CBF. By multiple sequence alignment and phylogenetic analysis, it is indicated that AeMYB4 clustered with AtMYB15 from Arabidopsis thaliana, belonging to subgroup S2 of R2 R3-MYB. And most of the transcription factors in this subfamily are related to low temperature stress. The GFP-AeMYB4 fusion protein expression vector for subcellular localization was constructed and transferred into Agrobacterium tumefaciens to infect the leaves of Nicotiana benthamiana, and the results showed the protein was located in the nucleus. To investigate the transcriptional activation, the constructed pGBKT7-AeMYB4 fusion expression vector was transferred into Y2 H Gold yeast cells, which proved that AeMYB4 was a transcription activator with strong transcriptional activity. Real-time quantitative PCR was used to detect the expression of AeMYB4 gene in three different development stages of stolon and in leaves, flowers, and bulbs of A. edulis, which indicated that AeMYB4 transcription factor was tissue-specific in expression, mainly in the stolon development stage, and that the expression was the most active in the middle stage of stolon development, suggesting that AeMYB4 gene may play an important role in stolon development. This study contributes to the further research on the function of AeMYB4 transcription factor in stolon development of A. edulis.

Regulation of plant MYB transcription factors in anther development / 生物工程学报

MYB transcription factor is one of the largest transcription families and involved in plant growth and development, stress response, product metabolism and other processes. It regulates the development of plant flowers, especially anther development, a key role in the reproduction of plant progeny. Here, we discuss the regulatory effects of MYB transcription factors on the development of anther, including tapetum development, anther dehiscence, pollen development, carbohydrates and hormone pathways. We provide a reference for the further study of the regulation mechanism and network of plant anther development.

Molecular characterization of PgUFGT gene and R2R3-PgMYB transcription factor involved in flavonoid biosynthesis in four tissues of wild pomegranate (Punica granatum L.)

The diversity on fruit colouration in plants directly depends on the flavonoids that explain the development of different pigmentation patterns. Anthocyanins are the major class of flavonoid pigments that are synthesized through flavonoid biosynthetic pathway. In the present study, two genes: PgUFGT gene and R2R3-PgMYB gene, involved in anthocyanin biosynthesis were analysed in four tissues of wild pomegranate. The structural genes, UDP-glucose: flavonoid-3-O-glucosyl transferase (PgUFGT; GenBank accession number: MK058491) and its myeloblastosis transcription factor (R2R3-PgMYB; GenBank accession number: MK092063) were isolated and their expression pattern were studied. Molecular modelling indicated that the main secondary structures of PgUFGT and R2R3-PgMYB genes are α-helix and random coil. In addition, expression profiling of PgUFGTand R2R3-PgMYB by quantitative-real time PCR indicated a positive correlation between anthocyanin content and their expression in leaves, flowers, green and red fruits of wild pomegranate. Among all the tissues, the red fruit exhibited high transcripts levels of PgUFGT as well as R2R3-PgMYB transcription factor. An extensive homology with UFGTs from other plants was revealed on comparative and bioinformatic analyses. Present study reveals that PgUFGT plays a predominant role in anthocyanin content in wild pomegranate fruits. Further, it is strongly suggested that R2R3-PgMYB transcription factor regulates the anthocyanin biosynthesis in wild pomegranate via expression of PgUFGT gene. This is the first study which provides an insight on expression profile of PgUFGT and R2R3-PgMYB that are involved in colour development and fruit ripening in wild pomegranate.

Cloning and expression analysis of MYB transcription factor genes in safflower / 药学学报

Safflower is a dried flower of the annual herbaceous plant safflower (Carthamus tinctorius L.). As a traditional Chinese medicine, it was widely used in the regulation of blood circulation. Flavonoids are the main active ingredients in safflower. MYB transcription factors are involved in the regulation of flavonoids. The cloning and expression analysis of MYB transcription factor genes in safflower is of great significance, not only for clarifying the regulation mechanism of flavonoids biosynthesis in safflower, but also for the artificial regulation of flavonoid biosynthesis in safflower. Based on the transcriptome data, we used iTAK to annotate the MYB transcription factors in safflower. The MYB transcription factors were cloned and their sequences were analyzed. Besides, their expressions were analyzed by a Real-time PCR. In the experiment, eight long fragment MYB transcription factors were screened and six MYB transcription factors was successfully cloned, named CtMYB-TF1, CtMYB-TF2, CtMYB-TF4, CtMYB-TF5, CtMYB-TF6 and CtMYB-TF7, respectively. The six MYB transcription factors had the core domain of MYB transcription factor family, and evolutionary analysis showed that the CtMYB-TF7 transcription factor was closely related to the factors AtMYBL2 and AtMYB12. Expression analysis showed that the expression of CtMYB-TF5, CtMYB-TF6 and CtMYB-TF7 was low in roots, stems and leaves, and was high in the flower. The results provide a foundation for study of mechanism of molecular regulation of safflower flavonoids.

Cloning and expression analysis of R1-MYB transcription factor in Lycium ruthenicum / 中草药

Objective To clone the R1-MYB transcription factor participated in the anthocyanidin metabolism, and to analyze by bioinformatics analysis. Different expression of different varieties, different organs of the same species and salt stress conditions in Lycium were analyzed. To clone the full-length cDNA encoding R1-MYB, to perform bioinformatic analysis, and to study its expression in different cultivators and different developmental stage and in response to NaCl stress in Lycium ruthenicum and L. barbarum. Methods The full-length cDNA encoding R1-MYB was cloned using homology-based cloning and rapid amplification of cDNA ends (RACE) technique in L. ruthenicum, and the homologous gene was obtained by transcriptome in L. barbarum. The bioinformatics analysis was carried out by using Prot, Param, Smart, PSORT, and SOPMA methods. And the phylogenetic tree was constructed based on software MEGA5.0. Gene expression analysis was done by method of Real-time PCR. Results We the MYB transcription factor in L. ruthenicum was cloned and named as LrMYB1R1 (GenBank accession number KY568981), and LbMYB1R1 (GenBank accession number KY568982) in L. barbarum. Bioinformatics analysis showed that the length of LrMYB1R1 was 1 496 bp and the CDS was 927 bp. The coding products contained 308 amino acids, the molecular weight of the protein was 33 400 and 33 490, the theoretical isoelectric point was 7.80 and 7.78, belonging to the R1-MYB transcription factor, and the encoded protein is predicted to be located in the nucleus. The results of phylogenetic tree analysis showed that LrMYB1R1 and LbMYB1R1 were highly similar to MYB1R1-like protein in Solanum lycopersicum, Solanum tuberosum, and Nicotiana tabacum. Real-time PCR analysis showed that LrMYB1R1 had higher expression level in leaves and young fruits in L. ruthenicum, followed by stems, young leaves, flowers, purple fruits and black fruits, only slightly expressed in roots. In addition, the relative expression levels of LrMYB1R1 decreased in response to salt stress. Conclusion The study of R1 MYB transcription factor has been enriched, which has laid the foundation for the subsequent research on gene function and for the high-yielding anthocyanin by genetic engineering method in L. ruthenicum.

Cloning and expression analysis of transcription factor AsMYB1 and AsMYB2 from Aquilaria sinensis / 中国中药杂志

The MYB gene family comprises one of the richest groups of transcription factors in plants. The full length of two MYB genes were isolated through heterologous screening of Aquilaria sinensis calli transcriptome data, and the reverse transcription PCR was performed to obstain the corrected MYB clones, named AsMYB1, AsMYB2. The MYB transmembrane domain and phylogenetic analysis were predicted by different software to analyze the bioinformatics of MYB proteins. The transcript level of AsMYB1, AsMYB2 was performed by real-time quantitative RT-PCR in different tissues and in responds to abiotic stresses including salt, cold, metal and drought stress, and hormone treatments including abscisic acid (ABA), salicylic acid (SA), gibberellins (GA3) and methyl jasmonate (MeJA) treatment. The AsMYB1 cDNA sequence had an ORF of 1 063 nucleotides, encoding a protein of 353 amino acids. The largest AsMYB2 ORF was 1 081 nucleotides, and its predicted translation products consisted of 359 amino acids. Two MYB genes had a tissues-specific pattern in A. sinensis. Moreover, the expression level of AsMYB1 and AsMYB2 was regulated by different abiotic stresses and hormone treatments, suggesting the transcription factors AsMYB1 and AsMYB2 play an important role in plant defense and hormone signal transduction in A. sinensis.

Cloning and expression analysis of MYB transcription factor gene involved in regulation of flavonoids biosynthesis in Carthamus tinctorius / 中草药

Objective To clone the MYB transcription factor gene in safflower, the sequence information and gene expression were analyzed to preliminarily identify the MYB transcription factor gene involved in regulation of flavonoid biosynthesis. Methods All the sequences of MYB transcription factor gene that was reported to involved in regulation of flavonoid biosynthesis were analyzed. The degenerate primers were designed to clone the core sequence. The full length of MYB transcription factor genes were cloned by RACE. The bioinformatics were used to analyze the sequences. The expression of MYB transcription factor genes in different tissues and different developmental stages of flower were analyzed by semi-quantitative PCR. Results Three candidate MYB transcription factors were cloned, named as CtFRMYB1, CtFRMYB2 and CtFRMYB3. The full lengths of the sequence were 1 223 bp, 1 080 bp and 1 348 bp, respectively. And the molecular weight were 17 878.15, 28 766.45 and 27 987.89, respectively. All three transcription factors have DNA binding domain and belong to the MYB family. Homologous analysis showed that CtFRMYB1 and CtFRMYB2 were closely related to AtMYB12. Expression analysis showed that CtFRMYB1 and CtFRMYB2 were only expressed in flowers, and the expression level was higher in flowering stage 3. Conclusion Three candidate MYB transcription factor genes involved in regulation of flavonoid biosynthesis (CtFRMYB1, CtFRMYB2 and CtFRMYB30) were successfully cloned. Two MYB transcription factor genes (CtFRMYB1 and CtFRMYB2) were identified by bioinformatics and expression analysis. These result laid a foundation for the molecular mechanism analysis of flavonoid biosynthesis.