Subject: UV-B radiation and low temperature promoted hypericin biosynthesis in adventitious root culture of Hypericum perforatum

Abstract

The hypericin is assumed as a highly demanded and key bioactive compound, which has antiviral, antimicrobial, antioxidant, and antitumor properties isolated from Hypericum perforatum. Nowadays, increasing bioactive molecules' contents through generating novel compounds is one of the major research objectives of H. perforatum biotechnology; however, this plant remains recalcitrant and unmanageable to Agrobacterium mediated transformation and genetic improvement programs. In order to overcome these challenges, many researchers have focused on this unruly herb using biotic and abiotic eliciting strategies. Therefore, two experiments were separately designed for the evaluation of two types of abiotic elicitors, aiming at increasing the productivity of hypericin in the adventitious root suspension culture of H. perforatum. The first one was accomplished to evaluate the effect of UV-B light elicitors (the exposure time of 30, 60, and 90 min) and the recovery treatment (with or without) on hypericin content while the second one was assessed the effect of various temperatures (4°C, 8°C, 16°C, and 25°C) in three different exposure times (24 h, 72 h, and 7 d). Based on the results, UV-B (60 min) treatment followed by the recovery produced 0.430 µg/g DW hypericin and was distinguished as the most effective UV-B elicitation treatment. In addition, a temperature of 4°C for a period of 72 hours is required to get the highest amount of hypericin content. These findings indicate the fact that hypericin biosynthesis is notably affected by UV-B exposure time and Low-temperature. The data also clearly elucidate further mechanisms of hypericin production in H. perforatum adventitious root culture.

Physiological and Transcriptomic Analysis Provide Insight into Low Temperature Enhancing Hypericin Biosynthesis in Hypericum perforatum

“…While low temperatures improving the plant growth and Hyp accumulation by stimulating related gene expressions has been observed in H. perforatum [20,22,27,28], the mechanism responsible for temperature-dependent growth and Hyp biosynthesis has not been dissected. Here, it is shown that there is a greater plant biomass and Hyp accumulation, more leaf dark glands, stomatal density and rounded stomatal shape at 15 compared to 22 • C. By treating seedlings at 15 and 22 • C, ca.…”

“…Experimental evidence shows that H. perforatum plants can adapt to low temperatures by increasing the biomass and morphogenesis (e.g., leaf area, number of stem and its internode and root), and low temperatures can improve Hyp biosynthesis by increasing the number of dark glands and related gene expressions [10,12,20,22,27,28]. The current study exhibited greater biomass by increasing the stomatal density and opening percentages, as well as a more rounded stomatal shape with the ratio of length/width and greater Hyp content via the overexpression of 61 genes involved in Hyp biosynthesis (Figures 2 and 3 and Table 1).…”

“…Based on biological functions, the 749 genes were divided into 11 categories: photosynthesis/energy (25), primary metabolism (85), secondary metabolism (62), cell morphogenesis (60), bio-signaling (93), hormone biosynthesis (23), polynucleotide biosynthesis (33), transcription factor (112), translation (58), transport (104) and stress response (94) (Figure 4D). Based on the pathways for the control of Hyp biosynthesis, plant growth and stress response [10,20], finally, 150 DEGs (90 UR and 60 DR) were dug out as master genes involved in photosynthesis (12), carbohydrate metabolism (13), fatty acid metabolism (15), cytochrome P450 (21), morphophysiological traits (15), heat shock protein (15), coldresponsive protein (3) and transcription factors (56). The differential expression levels of the 150 genes based on Reads Per kb per Million (RPKM) values are shown in a heat map (Figure 5 and Tables S4-S10).…”

“…Currently, Hyp is mainly extracted from H. perforatum [8]. In order to improve the Hyp production, many approaches that have been investigated include dissecting the Hyp biosynthetic pathway [9][10][11], screening germplasm resources [12][13][14], evaluating harvest stages [8,12,15], finding out the optimization of environmental factors such as nutrients [16], drought stress [17], UV-B radiation [18][19][20], light [21] and temperature [12,20,22,23].…”

“…Specifically, Brunáková et al [27] reported that a −4 • C treatment improved the Hyp content on a dry weight (DW) basis. Tavakoli et al [20] found that the Hyp content on a DW basis in a root suspension culture reached the highest amount at 4 • C than at 8, 16 and 25 • C. Velada et al [28] determined that the genes (phenolic oxidative coupling protein (Hyp-1), PKS1 and PKS2) involved in Hyp synthesis were differentially expressed at different temperatures. Our previous studies found that 15 • C could enhance aerial parts of a biomass and Hyp contents of dry weight (DW) on a per plant basis, as well as mRNA expressions of the hyp-1 and PKS genes, compared to 22 • C [22].…”