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

Anjin Zhao,Tingge Li,Yang Zhou,Wenjun Huang,Huiqin Pei,Jian Wang,Ting Peng,Xiqiang Song,Zheng Cui,Ying Zhao,Xueqing Li,Yuhui Sheng

doi:10.3390/ijms20174250

Abstract

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.

Highlights

Flower spots are heterochromatic dots or streaks with a specific texture and pattern appearing on the corollas of plants, which can affect the behavior of pollinators and the ornamental value of flowers [1,2,3,4]
Peonidin-3-O-glucoside, malvidin-3-O-glucoside, delphinidin-3-O-diglucoside, and cyanidin-3-O-glucoside are the main anthocyanins found in petal and sepal spots in Oncidium [5], and cyanidin and delphinidin are the main anthocyanidins in the spot of pansy (Viola × wittrockiana Gams.) petals [6]
Molecular mechanism of anthocyanins accumulation has been clearly studied in some plants, such as Dianthus hybrida [7], Antirrhinum majus [8], Petunia hibrida [9], Platycodonis Radix [10], Gerbera jamesonii [11], and Phalaenopsis equestris [12], and the anthocyanin pathway, which is a branch of the flavonoid pathway, has been elucidated as well [13,14]

Summary

Introduction

Flower spots are heterochromatic dots or streaks with a specific texture and pattern appearing on the corollas of plants, which can affect the behavior of pollinators and the ornamental value of flowers [1,2,3,4]. Previous studies have confirmed the flower spot is caused by the accumulation of anthocyanins in a specific area of corollas. The large purple spots in Phalaenopsis ‘Everspring Fairy’ was mainly caused by the expression of MYB [12], and the LhMYB6 and LhMYB12 positively regulate anthocyanin biosynthesis and determine organ- and tissue-specific accumulation of anthocyanin in Asiatic hybrid lily ‘Montreux’ [20]. Hsu conducted a detailed study on three MYBs in Phalaenopsis, and found that the color patterning of flower sepals, petals and lips is regulated by different MYB genes combinations, and the pigmented veins and spots on the petals are regulated by these three MYB genes [21]

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Results

Conclusion