CmNAC73 Mediates the Formation of Green Color in Chrysanthemum Flowers by Directly Activating the Expression of Chlorophyll Biosynthesis Genes HEMA1 and CRD1

Abstract

Chrysanthemum is one of the most beautiful and popular flowers in the world, and the flower color is an important ornamental trait of chrysanthemum. Compared with other flower colors, green flowers are relatively rare. The formation of green flower color is attributed to the accumulation of chlorophyll; however, the regulatory mechanism of chlorophyll metabolism in chrysanthemum with green flowers remains largely unknown. In this study, we performed Illumina RNA sequencing on three chrysanthemum materials, Chrysanthemum vestitum and Chrysanthemum morifolium cultivars ‘Chunxiao’ and ‘Green anna,’ which produce white, light green and dark green flowers, respectively. Based on the results of comparative transcriptome analysis, a gene encoding a novel NAC family transcription factor, CmNAC73, was found to be highly correlated to chlorophyll accumulation in the outer whorl of ray florets in chrysanthemum. The results of transient overexpression in chrysanthemum leaves showed that CmNAC73 acts as a positive regulator of chlorophyll biosynthesis. Furthermore, transactivation and yeast one-hybrid assays indicated that CmNAC73 directly binds to the promoters of chlorophyll synthesis-related genes HEMA1 and CRD1. Thus, this study uncovers the transcriptional regulation of chlorophyll synthesis-related genes HEMA1 and CRD1 by CmNAC73 and provides new insights into the development of green flower color in chrysanthemum and chlorophyll metabolism in plants.

TcMYB8, a R3-MYB Transcription Factor, Positively Regulates Pyrethrin Biosynthesis in Tanacetum cinerariifolium

“…The TcMYB8 CDS was amplified by PCR using sequence-specific primers containing Swa I and Kpn I restriction sites at the end. Using a ClonExpress ® II One Step Cloning Kit (Vazyme Biotech, Nanjing, China), the TcMYB8 CDS (without the stop codon) was cloned into the pSuper1300-GFP vector (derived from pCambia130075) [ 49 ]. The obtained plasmid or empty vector (control) was introduced into Agrobacterium tumefaciens strain GV3101, which was then co-infiltrated into N. benthamiana leaves with the RFP-NLS plasmid (nuclear marker) [ 49 ].…”

“…Using a ClonExpress ® II One Step Cloning Kit (Vazyme Biotech, Nanjing, China), the TcMYB8 CDS (without the stop codon) was cloned into the pSuper1300-GFP vector (derived from pCambia130075) [ 49 ]. The obtained plasmid or empty vector (control) was introduced into Agrobacterium tumefaciens strain GV3101, which was then co-infiltrated into N. benthamiana leaves with the RFP-NLS plasmid (nuclear marker) [ 49 ]. After 72 h of shade adaptation, fluorescence signals were detected with a confocal laser scanning microscope (Leica TCS-SP8; Leica, Wetzlar, Germany).…”

“…The transient gene expression assay was performed as previously described in chrysanthemum [ 49 ]. Briefly, Agrobacterium GV3101 (pSoup19) cells containing the pGreen-62-SK-TcMYB8 vector were inoculated into the YEB liquid medium containing 100 mg/L kanamycin, and cultured on a rotatory shaker at 28 °C for 12 h. The cells were harvested by centrifugation.…”

SMRT and Illumina RNA Sequencing and Characterization of a Key NAC Gene LoNAC29 during the Flower Senescence in Lilium oriental ‘Siberia’

“…For example, in chrysanthemum (Chrysanthemum morifolium Ramat. ), a NAC transcription factor was found to mediate chlorophyll synthesis partly through direct activation of HEMA1 and CRD1 genes [51]. In L. oriental 'Siberia', most of the genes involved in chlorophyll synthesis and photosynthesis were clustered into subcluster 1, subcluster 2, and subcluster 11.…”

Important Roles of Key Genes and Transcription Factors in Flower Color Differences of Nicotiana alata

“…Chlorophylls are present in many flowering plants during the early development stage of petals, and their contents gradually decrease as the petals develop [ 4 ]. The chlorophyll metabolism pathway has been studied in petals in many species, such as carnation [ 5 ], chrysanthemum [ 4 , 6 , 7 ] and petunia [ 8 ]. 5-Aminolevulinic acid (ALA) biosynthesis is the rate-limiting step of chlorophyll synthesis, and ALA is formed from glutamyl-tRNA by two enzymatic steps.…”