Identification of the function of a key gene NnHCT1 in lignin synthesis in petioles of Nelumbo nucifera

Abstract

Leaf petiole or stem strength is an important agronomic trait affecting the growth of underground organs as a channel for material exchange and plays a vital role in the quality and yield of crops and vegetables. There are two different types of petioles in lotus, floating leaf petioles and vertical leaf petioles; however, the internal difference mechanism between these petioles is unclear. In this study, we investigated the differences between the initial vertical leaf petioles and the initial floating leaf petioles based on RNA sequencing (RNA-seq), and >2858 differentially expressed genes were annotated. These genes were chiefly enriched in phenylpropanoid biosynthesis, which is the source of the lignin and cellulose in petioles and stems. Lignin biology-related gene NnHCT1 was identified, and subsequent biological function validation demonstrated that the transient overexpression of NnHCT1 significantly increased the lignin and cellulose contents in lotus petioles and tobacco leaves. In contrast, silencing NnHCT1 through virus-induced gene silencing significantly reduced petiole lignin synthesis. Additionally, differentially up-regulated MYB family transcription factors were identified using RNA-seq. Yeast-one-hybrid and dual-luciferase reporter assays demonstrated that MYB4 could bind to the NnHCT1 promoter and up-regulate NnHCT1 expression. These findings demonstrate the significant potential of NnHCT1 to enhance lignin synthesis, thereby improving stem or petiole resistance to stunting and explaining the need for the study of differential petiole relationships in plants.