CtACO1 Overexpression Resulted in the Alteration of the Flavonoids Profile of Safflower

Abstract

Background: Flavonoids with various structures play a vital role in plant acclimatization to varying environments as well as in plant growth, development, and reproduction. Exogenous applications of ethylene and 1-aminocyclopropane carboxylic acid (ACC), could affect the accumulation of flavonoids. Very few attempts have been made to investigate the effect of 1-aminocyclopropane carboxylic acid oxidase (ACO), a unique enzyme that catalyzes ACC to ethylene, on genes and metabolites in the flavonoid biosynthetic pathway. In this study, two ACOs in safflower (CtACOs) were cloned, and then transgenic safflower with overexpressed CtACO1 was generated through the Agrobacterium-mediated floral dipping method. Results: CtACO1 and CtACO2 were both characterized by the 2-oxoglutarate binding domain RxS and the ferrous iron binding site HxDxnH as ACOs from other plants. However, the transcript levels of CtACO1 in flowers at stages I, II, III, and IV were all higher than those of CtACO2. At the cellular level, by using electroporation transformation, CtACO1 was found to be localized at the cytomembrane in onion epidermal cells. CtACO1 overexpression had varying effects on genes involved in the ethylene and flavonoid biosynthetic pathways. The metabolites analysis showed that CtACO1 overexpression lines had a higher accumulation of quercetin and its glycosylated derivatives (quercetin 3-β-d-glucoside and rutin). In contrast, the accumulation of quinochalcones (hydroxysafflor yellow A and carthamin), kaempferol glycosylated derivatives (kaempferol-3-O-β-rutinoside and kaempferol-3-O-β-d-glucoside), apigenin, and luteolin in CtACO1 overexpression lines were decreased. Conclusion: This study confirmed the feasibility of applying the floral dipping method to safflower and showed a novel regulatory effect of CtACO1 in the flavonoid biosynthetic pathway. It provides hypothetical and practical groundwork for further research on regulating the overall metabolic flux of flavonoids in safflower, particularly hydroxysafflor yellow A and other quinochalcones, by using appropriate genetic engineering strategies.

1-Aminocyclopropane-1-Carboxylic Acid Oxidase (ACO): The Enzyme That Makes the Plant Hormone Ethylene

“…A recent study tagged a safflower ( Carthamus tinctorius ) ACO with a GFP (green fluorescent protein) and performed an ectopic localization in onion epidermis cells. Their results showed that Ct ACO1 localizes in the cytosol (potentially linked with membranes) and in the nucleus, but their images lacked markers for these organelles (Tu et al, 2019). All studies combined are not conclusive about the exact ACO localization, and thus the actual subcellular site of ethylene production.…”

“…Besides silencing ACO expression and reducing ethylene production, it can sometimes be desirable to boost ethylene production, and then an ACO overexpression construct is most suitable. In safflower, ACO1 overexpression was shown to stimulate the flavonoid biosynthesis pathway, which could be interesting for oilseed production (Tu et al, 2019). Overexpression of ACO1 in poplar ( Populus tremula × tremuloides ) caused a stimulation of cambial cell division, which in turn resulted in an increased xylem development and an inhibition of elongation growth, which are desirable traits for the wood industry (Love et al, 2009).…”

The Comprehensive Evaluation of Safflowers in Different Producing Areas by Combined Analysis of Color, Chemical Compounds, and Biological Activity

“…From the many previous reports, it has been confirmed that HSYA has a strong correlation with the color in the safflower [11,12]. Additionally, recent researches have shown that quinochalcones and flavonoids have a certain correlation with safflower color [12,25,26]. Therefore, three quinochalcones (HSYA, SC, and ASYB) and 13 flavonoids were selected, including their chromaticity-related characteristic components, for quantitative analysis in the present research.…”

Modulation of Organogenesis and Somatic Embryogenesis by Ethylene: An Overview

“…This modulation can be achieved using sense-or antisense-mediated gene silencing, co-suppression, or by using an RNA interference (RNAi) approach, with several studies carried out in several plant species [30,49,58,59]. Regarding an ethylene overproduction strategy, the overexpression of the ACO gene was achieved in poplar [60] and in safflower [61], by transforming these plants with an overexpression vector containing the open reading frame of the specific ACO. Note that the ACO enzyme is the most studied and suitable target regarding genetic alterations of ethylene biosynthesis due to its reduced risk of interfering with other metabolic pathways, such as the methionine cycle and ACC metabolism [30].…”