The RAV transcription factor TEMPRANILLO1 involved in ethylene-mediated delay of chrysanthemum flowering.

TEMPRANILLO1 (TEM1) is a transcription factor belonging to related to ABI3 and VP1 family, which is also known as ethylene response DNA-binding factor 1 and functions as a repressor of flowering in Arabidopsis. Here, a putative homolog of AtTEM1 was isolated and characterized from chrysanthemum, designated as CmTEM1. Exogenous application of ethephon leads to an upregulation in the expression of CmTEM1. Knockdown of CmTEM1 promotes floral initiation, while overexpression of CmTEM1 retards floral transition. Further phenotypic observations suggested that CmTEM1 involves in the ethylene-mediated inhibition of flowering. Transcriptomic analysis established that expression of the flowering integrator CmAFL1, a member of the APETALA1/FRUITFULL subfamily, was downregulated significantly in CmTEM1-overexpressing transgenic plants compared with wild-type plants but was verified to be upregulated in amiR-CmTEM1 lines by quantitative RT-PCR. In addition, CmTEM1 is capable of binding to the promoter of the CmAFL1 gene to inhibit its transcription. Moreover, the genetic evidence supported the notion that CmTEM1 partially inhibits floral transition by targeting CmAFL1. In conclusion, these findings demonstrate that CmTEM1 acts as a regulator of ethylene-mediated delayed flowering in chrysanthemum, partly through its interaction with CmAFL1.

Comparative transcriptome analysis reveals whole-genome duplications and gene selection patterns in cultivated and wild Chrysanthemum species.

KEY MESSAGE: Comparative transcriptome analysis of wild and cultivated chrysanthemums provides valuable genomic resources and helps uncover common and divergent patterns of genome and gene evolution in these species. Plants are unique in that they employ polyploidy (or whole-genome duplication, WGD) as a key process for speciation and evolution. The Chrysanthemum genus is closely associated with hybridization and polyploidization, with Chrysanthemum species exhibiting diverse ploidy levels. The commercially important species, C. morifolium is an allohexaploid plant that is thought to have originated via the hybridization of several Chrysanthemum species, but the genomic and molecular evolutionary mechanisms remain poorly understood. In the present study, we sequenced and compared the transcriptomes of C. morifolium and the wild Korean diploid species, C. boreale. De novo transcriptome assembly revealed 11,318 genes in C. morifolium and 10,961 genes in C. boreale, whose functions were annotated by homology searches. An analysis of synonymous substitution rates (Ks) of paralogous and orthologous genes suggested that the two Chrysanthemum species commonly experienced the Asteraceae paleopolyploidization and recent genome duplication or triplication before the divergence of these species. Intriguingly, C. boreale probably underwent rapid diploidization, with a reduction in chromosome number, whereas C. morifolium maintained the original chromosome number. Analysis of the ratios of non-synonymous to synonymous nucleotide substitutions (Ka/Ks) between orthologous gene pairs indicated that 107 genes experienced positive selection, which may have been crucial for the adaptation, domestication, and speciation of Chrysanthemum.

Identification and characterization of a novel NAC-like gene in chrysanthemum (Dendranthema lavandulifolium).

KEY MESSAGE: A NAC -like gene named DlNAC1 was identified in chrysanthemum and characterized; it may be involved in regulation of response to abiotic stressors, especially in tolerance to drought and salinity. NAC transcription factors in plants play crucial roles in tolerance to abiotic stressors, and overexpression of the NAC gene in Arabidopsis has been demonstrated to lead to improved drought tolerance. Functions of the NAC genes in chrysanthemum, however, remain poorly understood. In this study, a NAC-like gene named DlNAC1 was identified in chrysanthemum (Dendranthema lavandulifolium) and characterized. Phylogenetic analysis indicated that DlNAC1 contains a typical NAC domain and belongs to the ONAC022 subgroup. According to the subcellular localization and yeast one-hybrid assay, the DlNAC1 protein is localized to nuclei and has a transcription activation ability. Moreover, quantitative real-time PCR analyses showed that DlNAC1 was induced by low-temperature, high-salinity, and drought conditions (separately), but not by abscisic acid (ABA) and heat shock. In these experiments, the downstream genes of NAC transcription factors were found to be up-regulated, including stress-responsive genes KIN1 and AMY1. To further explore the effects of DlNAC1 in response to abiotic stressors, DlNAC1 was overexpressed in tobacco, and these transgenic plants showed significantly enhanced tolerance to drought and salinity. This study suggests that in chrysanthemum, the DlNAC1 gene is involved in regulation of the response to abiotic stressors, especially in tolerance to drought and salinity.

Overexpression of a novel chrysanthemum Cys2/His2-type zinc finger protein gene DgZFP3 confers drought tolerance in tobacco.

A drought stress-responsive Cys2/His2-type zinc finger protein gene DgZFP3 was previously isolated (Liu et al., Afr J Biotechnol 11:7781-7788, 2012b) from chrysanthemum. To assess roles of DgZFP3 in plant drought stress responses, we performed gain-of-function experiment. The DgZFP3-overexpression tobacco plants showed significant drought tolerance over the wild type (WT). The transgenic lines exhibited less accumulation of H2O2 under drought stress, more accumulation of proline and greater activities of peroxidase (POD) and superoxide dismutase than the WT under both control conditions and drought stress. In addition, there was greater up-regulation of the ROS-related enzyme genes (NtSOD and NtPOD) and stress-related genes (NtLEA5 and NtDREB) in transgenic lines under normal or drought conditons. Thus DgZFP3 probably plays a positive regulatory role in drought stress response and has the potential to be utilized in transgenic breeding to improve drought stress tolerance in plants.

Involvement of the ethylene response pathway in dormancy induction in chrysanthemum.

Temperature plays a significant role in the annual cycling between growth and dormancy of the herbaceous perennial chrysanthemum (Chrysanthemum morifolium Ramat.). After exposure to high summer temperatures, cool temperature triggers dormancy. The cessation of flowering and rosette formation by the cessation of elongation are characteristic of dormant plants, and can be stimulated by exogenous ethylene. Thus, the ethylene response pathway may be involved in temperature-induced dormancy of chrysanthemum. Transgenic chrysanthemums expressing a mutated ethylene receptor gene were used to assess this involvement. The transgenic lines showed reduced ethylene sensitivity: ethylene causes leaf yellowing in wild-type chrysanthemums, but leaves remained green in the transgenic lines. Extension growth and flowering of wild-type and transgenic lines varied between temperatures: at 20 degrees C, the transgenic lines showed the same stem elongation and flowering as the wild type; at cooler temperatures, the wild type formed rosettes with an inability to flower and entered dormancy, but some transgenic lines continued to elongate and flower. This supports the involvement of the ethylene response pathway in the temperature-induced dormancy of chrysanthemum. At the highest dosage of ethephon, an ethylene-releasing agent, wild-type plants formed rosettes with an inability to flower and became dormant, but one transgenic line did not. This confirms that dormancy is induced via the ethylene response pathway.