mRNA and miRNA Expression Analysis Reveal the Regulation for Flower Spot Patterning in Phalaenopsis 'Panda'.

Phalaenopsis cultivar 'Panda' is a beautiful and valuable ornamental for its big flower and unique big spots on the petals and sepals. Although anthocyanins are known as the main pigments responsible for flower colors in Phalaenopsis, and the anthocyanins biosynthetic pathway in Phalaenopsis is generally well known, the detailed knowledge of anthocynins regulation within the spot and non-spot parts in 'Panda' flower is limited. In this study, transcriptome and small RNA libraries analysis from spot and non-spot sepal tissues of 'Panda' were performed, and we found PeMYB7, PeMYB11, and miR156g, miR858 is associated with the purple spot patterning in its sepals. Transcriptome analyses showed a total 674 differentially expressed genes (DEGs), with 424 downregulated and 250 upregulated (Non-spot-VS-Spot), and 10 candidate DEGs involved in anthocyanin biosynthetic pathway. The qPCR analysis confirmed that seven candidate structure genes (PeANS, PeF3'H, PeC4H, PeF3H, PeF3H1, Pe4CL2, and PeCHI) have significantly higher expressing levels in spot tissues than non-spot tissues. A total 1552 differentially expressed miRNAs (DEMs) were detected with 676 downregulated and 876 upregulated. However, microRNA data showed no DEMs targeting on anthocyanin biosynthesis structure gene, while a total 40 DEMs target transcription factor (TF) genes, which expressed significantly different level in spot via non-spot sepal, including 2 key MYB regulator genes. These results indicated that the lack of anthocyanidins in non-spot sepal may not directly be caused by microRNA suppressing anthocyanidin synthesis genes rather than the MYB genes. Our findings will help in understanding the role of miRNA molecular mechanisms in the spot formation pattern of Phalaenopsis, and would be useful to provide a reference to similar research in other species.

Tyramine and tyrosine decarboxylase gene contributes to the formation of cyanic blotches in the petals of pansy (Viola × wittrockiana).

Tyrosine decarboxylase (TYDC) can catalyze tyrosine into tyramine. Several studies demonstrated its roles in the acidity, salidroside and defense response. Here we found that TYDC from Viola × wittrockiana Gam (VwTYDC) may contribute to the formation of cyaninc blotches in the petal. VwTYDC gene were cloned from Viola × wittrockiana and the cDNA full-length sequences were 1634 bp encoding 494 amino acids. Gene expression of VwTYDC in different tissues and developmental stages showed that they were significantly higher expressed in flowers than stems, leaves and roots. In addition, VwTYDC expression were higher in cyanic blotches than those observed in acyanic blotches of petal. Metabolites analysis showed the contents of tyramine in cyanic blotches were also higher than that in acyanic areas. Furthermore, in vitro assay revealed the absorption peak of anthocyanins had a red shift and an increase when fed tyramine. We speculated that tyramine might contribute to flower color expression of pansy as co-pigment. Our study demonstrated for the first time that the contents of tyramine led to flower blotches formation in cyanic blotches of the petals in plant flowers, and this may due to the higher expression of VwTYDC gene.

Exogenous Hydrogen Peroxide Contributes to Heme Oxygenase-1 Delaying Programmed Cell Death in Isolated Aleurone Layers of Rice Subjected to Drought Stress in a cGMP-Dependent Manner.

Hydrogen peroxide (H2O2) is a reactive oxygen species (ROS) that plays a dual role in plant cells. Here, we discovered that drought (20% polyethylene glycol-6000, PEG)-triggered decreases of HO-1 transcript expression and HO activity. However, exogenous H2O2 contributed toward the increase in HO-1 gene expression and activity of the enzyme under drought stress. Meanwhile, the HO-1 inducer hematin could mimic the effects of the H2O2 scavengers ascorbic acid (AsA) and dimethylthiourea (DMTU) and the H2O2 synthesis inhibitor diphenyleneiodonium (DPI) for scavenging or diminishing drought-induced endogenous H2O2. Conversely, the zinc protoporphyrin IX (ZnPPIX), an HO-1-specific inhibitor, reversed the effects of hematin. We further analyzed the endogenous H2O2 levels and HO-1 transcript expression levels of aleurone layers treated with AsA, DMTU, and DPI in the presence of exogenous H2O2 under drought stress, respectively. The results showed that in aleurone layers subjected to drought stress, when the endogenous H2O2 level was inhibited, the effect of exogenous H2O2 on the induction of HO-1 was enhanced. Furthermore, exogenous H2O2-activated HO-1 effectively enhanced amylase activity. Application of 8-bromoguanosine 3',5'-cyclic guanosine monophosphate (8-Br-cGMP) (the membrane permeable cGMP analog) promoted the effect of exogenous H2O2-delayed PCD of aleurone layers in response to drought stress. More importantly, HO-1 delayed the programmed cell death (PCD) of aleurone layers by cooperating with nitric oxide (NO), and the delayed effect of NO on PCD was achieved via mediation by cGMP under drought stress. In short, in rice aleurone layers, exogenous H2O2 (as a signaling molecule) triggered HO-1 and delayed PCD via cGMP which possibly induced amylase activity under drought stress. In contrast, as a toxic by-product of cellular metabolism, the drought-generated H2O2 promoted cell death.