A Novel WRKY Transcription Factor HmoWRKY40 Associated with Betalain Biosynthesis in Pitaya (Hylocereus monacanthus) through Regulating HmoCYP76AD1.

Lulu Zhang,Jietang Zhao,Guibing Hu,Canbin Chen,Yonghua Qin,Zhike Zhang,Fangfang Xie,Qingzhu Hua,Jianye Chen,Rong Zhang

doi:10.3390/ijms22042171

Abstract

Betalains are water-soluble nitrogen-containing pigments with multiple bioactivities. Pitaya is the only large-scale commercially grown fruit containing abundant betalains for consumers. However, the upstream regulators in betalain biosynthesis are still not clear. In this study, HmoWRKY40, a novel WRKY transcription factor, was obtained from the transcriptome data of pitaya (Hylocereus monacanthus). HmoWRKY40 is a member of the Group IIa WRKY family, containing a conserved WRKY motif, and it is located in the nucleus. The betalain contents and expression levels of HmoWRKY40 increased rapidly during the coloration of pitaya and reached their maximums on the 23rd day after artificial pollination (DAAP). Yeast one-hybrid and transient expression assays showed that HmoWRKY40 could bind and activate the promoter of HmoCYP76AD1. Silencing the HmoWRKY40 gene resulted in a significant reduction of betacyanin contents. These results indicate that HmoWRKY40 transcriptionally activates HmoCYP76AD, which is involved in the regulation of pitaya betalain biosynthesis. The results of the present study provide new regulatory networks related to betalain biosynthesis in pitaya.

Highlights

Betalains are water-soluble and nitrogen-containing pigments with antioxidant properties, which were first extracted from the common beet (Beta vulgaris)
We investigated the possible association of HmoWRKY40, a Group IIa WRKY transcription factors (TFs), with the direct activation of HmoCYP76AD1, which is responsible for betalain biosynthesis in pitayas
These results indicate that the HmoWRKY40 protein has a DNA binding activity and could directly bind to the promoter of HmoCYP76AD1, which is possibly involved in the betalain biosynthesis of pitaya

Summary

Introduction

Betalains are water-soluble and nitrogen-containing pigments with antioxidant properties, which were first extracted from the common beet (Beta vulgaris). Understanding the biosynthetic and regulatory pathways for betalain biosynthesis is essential to the improvement of betalain production in plants through biotechnological approaches. CYP76AD1 foryellow yellow versus red and pigmentation) produces both yellow and pigmentation [9]. The expression of CYP76AD1 in yeast confirms itsmaps role to in the betalain biosynthesis [9,11]. Responsible for yellow versus red pigmentation) produces bothor yellow and red pigmenMoreover, heterologous expression of BvCYP76AD1. Yeast In confirms its rolestudy, in betalain biosynthesis HmoCYP76AD1), an important gene of cyt-P450 genes involved in the betalain results in biosynthetic an accumulation of betalains [11]. Expression levels of HmoCYP76AD1 increased betalain biosynthetic pathway, wasanalyzed [10].ofIncyt-P450 this study, the involved expression (named HmoCYP76AD1), an important candidate gene genes in and the promoter of HmoCYP76AD1 were

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Conclusion