Identification of Isoflavonoid Biosynthesis-Related R2R3-MYB Transcription Factors in Callerya speciosa (Champ. ex Benth.) Schot Using Transcriptome-Based Gene Coexpression Analysis

Abstract

The R2R3-MYB family is one of the largest plant transcription factor (TF) families playing vital roles in defense, plant growth, and secondary metabolism biosynthesis. Although this gene family has been studied in many species, isoflavonoid biosynthesis-related R2R3-MYB TFs in Callerya speciosa (Champ. ex Benth.) Schot, a traditional Chinese medicinal herb, are poorly understood. Here, a total of 101 R2R3-MYB TFs were identified from C. speciosa transcriptome dataset. 25 clades divided into five functional groups were clustered based on the sequence similarity and phylogenetic tree. Conserved motifs and domain distribution, expression patterns, and coexpression networks were also employed to identify the potential R2R3-MYB TFs in the regulation of isoflavonoid biosynthesis. In silico evaluation showed that the deduced R2R3-CsMYB proteins contain highly conserved R2R3 repeat domain at the N-terminal region, that is the signature motif of R2R3-type MYB TFs. Eight potential TFs (CsMYB17, CsMYB36, CsMYB41, CsMYB44, CsMYB45, CsMYB46, CsMYB72, and CsMYB81) had high degrees of coexpression with four key isoflavonoid biosynthetic genes (CsIFS, CsCHS7, CsHID-1, and CsCHI3), in which CsMYB36 as a potential regulator possessed the highest degree. HPLC analysis showed that formononetin and maackiain contents were significantly increased during the development of tuberous roots, which might be controlled by both related R2R3-CsMYBs and structural genes involved in the isoflavonoid biosynthesis pathway. The transcriptome data were further validated by reverse transcription real-time PCR (RT-qPCR) analysis, and similar expression profiles between TFs and key structural genes were identified. This study was the first step toward the understanding of the R2R3-MYB TFs regulating isoflavonoid biosynthesis in C. speciosa. The results will provide information for further functional analysis and quality improvement through genetic manipulation of these potential R2R3-CsMYB genes in C. speciosa.

Comprehensive physiological, transcriptomic, and metabolomic analyses reveal the synergistic mechanism of Bacillus pumilus G5 combined with silicon alleviate oxidative stress in drought-stressed Glycyrrhiza uralensis Fisch.

“…p -Coumaroyl-CoA, a key intermediate in the flavonoid biosynthesis pathway, provides the C6–C3–C6 carbon skeleton to the flavonoid compounds ( Yu et al., 2021 ). After the production of p -coumaroyl-CoA, the biosynthetic pathway of flavonoids begins to branch under the activity of CHS, CHR, and CHI ( Wang et al., 2015 ).…”

Selection and Validation of Suitable Reference Genes for Gene Expression Studies in Callerya speciosa (Champ. ex Benth.) Schot under Different Experimental Conditions

“…Under ABA treatment, the ideal reference genes of Nitraria tangutorum were EF1-α and His [20]. Our previous studies revealed that several R2R3-MYB TFs and structural genes might be involved regulating iso avonoid biosynthesis in C. speciosa, and CsMYB36 has the highest expression in R2R3-MYB TFs [21]. To understand these related gene expression changes will be facilitated to explain their functions in C. speciosa under different experimental conditions.…”

“…Our previous studies revealed that several R2R3-MYB TFs and structural genes might be involved regulating isoflavonoid biosynthesis in C. speciosa, in which one of the R2R3-MYB TFs CsMYB36 has a high expression level. 24 Understanding the expression patterns of these genes involved in the pathway of isoflavonoid biosynthesis will be facilitated to understand the accumulation and dynamic trends of (iso)flavonoids in C. speciosa. Furthermore, explaining the molecular mechanism associated with hormone stimuli to exogenous regulators may also help us to reveal what facilitates the quality of C. speciosa.…”

“…In order to further verify the reliability of the reference genes, an iso avone synthesis gene CsMYB36 was chosen to be the target gene. CsMYB36 has a high-level expression level in the C. speciosa R2R3-MYB transcription factor family[21]. The expression pattern of CsMYB36 in different samples was investigated using the two most stable reference genes and the least stable reference gene ranked by RefFinder program.…”

Transcriptome and metabolome reveal the accumulation of secondary metabolites in different varieties of Cinnamomum longepaniculatum

“…Many studies have shown that flavonoids are synthesized in plants via the phenylpropanoid and acetate–malonate metabolic pathways [ 5 ]. Many flavonoid biosynthesis-related genes have been studied in depth, including those encoding phenylalanine ammonia lyase (PAL), cinnamate 4-hydroxylase (C4H), 4-coumaroyl CoA ligase (4CL) and MYB transcription factors [ 6 ]. In addition, flavonoids can negatively regulate auxin transport [ 7 ].…”