The PdeWRKY65-UGT75L28 gene module negatively regulates lignin biosynthesis in poplar petioles

Abstract

Lignin is made up of three main p-hydroxycinamyl alcohol precursors or monolignols. Lignin biosynthesis is not only important for plant growth, but also the key factor for human use of plants as biofuels. In past decades, there were numerous studies have focused on lignification in plant stems; however, little has been uncovered about lignification in petioles. In this study, a gene encoding a glycosyltransferase was identified in poplar and named UGT75L28 . A relatively higher expression of UGT75L28 could be observed in petioles. Transgenic lines overexpressing UGT75L28 showed reduced lignification in petioles, while reduced expression (RE) of this gene displayed opposite phenotype. The protein of UGT75L28 fused with six His tags was expressed in E. coli and purified. In vitro enzyme reactions were conducted for a number of substrates for this fused protein. The results revealed that UGT75L28 could only glycosylate coniferyl aldehyde, which is a substrate in lignin biosynthesis in plants. The identification of the upstream regulator of UGT75L28 found that the transcription factor PdeWRKY65 could bind to the promoter of UGT75L28 and activate its expression both in vivo and in vitro by the yeast one-hybrid (Y1H) assay, the coexpression of the promoters of UGT75L28 and PdeWRKY65 in tobacco, and chromatin immunoprecipitation PCR (ChIP-PCR). Overexpression (OE) and RE PdeWRKY65 transgenic lines in poplar were also generated. The phenotype assay also showed that OE and RE PdeWRKY65 could reduce and enhance lignification in poplar petioles, respectively. The expression examination of PdeWRKY65 also showed that it was mainly expressed in petioles. Collectively, these results suggest that the gene module PdeWRKY65 - UGT75L28 negatively regulates petiole lignification by modulating the glycosylation of coniferyl aldehyde in poplar. • UGT75L28 can glycosylate a substrate in lignin biosynthesis in poplar. • PdeWRKY65 TF directly activates the expression of UGT75L28 . • PdeWRKY65 - UGT75L28 negatively regulates lignin biosynthesis in poplar petioles.