Identification and characterization of novel homeodomain leucine zipper (HD-Zip) transcription factors associated with heat tolerance in perennial ryegrass

The homeodomain leucine zipper (HD-Zip) family transcription factors play roles in regulating plant development and responses to abiotic stress; however, how HD-Zip genes are associated with plant heat tolerance is unclear. The objectives of this study were to characterize HD-Zip I genes in perennial ryegrass (Lolium perenne), analyze the expression pattern under different abiotic stresses and identify novel HD-Zip I members associated with genetic variations in heat tolerance. A total of 13 HD-Zip I genes were identified in the transcriptome of perennial ryegrass. A comprehensive phylogenetic tree of HD-Zip I proteins in perennial ryegrass, Arabidopsis and rice was constructed. Motif analysis between LpHD-Zip I and OsHD-Zip I by MEME was characterized. LpHD-Zip I had close evolutionary relationship to OsHD-Zip I, and HD-Zip I may be subject to functional differentiation between monocotyledons and dicotyledons. qPCR analysis of 13 HD-Zip I genes showed these genes expressed at different levels in roots and leaves. These genes also showed various expression levels under different abiotic stresses (cold, heat, salt and osmotic). Physiological responses of heat-sensitive lines (321681 and 418714) and heat-tolerant lines (231565 and 275660) of perennial ryegrass to heat stress were evaluated. Differential expression levels of four genes (LpHOX6, -8, -21 and -24) were detected between heat-sensitive and heat-tolerant lines. LpHOX6, -8 and -24 expression levels were negatively associated with heat tolerance while LpHOX21 was positively associated with heat tolerance of perennial ryegrass. This is the first study demonstrating the association of four members of the HD-Zip I family (LpHOX6, -8, -21, and -24) with genetic variations in heat tolerance.