Enhancer trapping in woody plants: Isolation of the ET304 gene encoding a putative AT-hook motif transcription factor and characterization of the expression patterns conferred by its promoter in transgenic Populus and Arabidopsis

Enhancer trapping is a useful tool in isolation of novel genes and functional characterization of promoters directing tissue-specific expression in trees. Using an enhancer trap approach we isolated a novel gene ET304 from Populus (Populus: aspens and cottonwoods). Both ET304 enhancer trap line and putative ET304 promoter fused to a GUS reporter gene conferred strongly root-predominant expression patterns and directed expression to specific root tissue and cell types. GUS activity was detected in lateral root primordia, root apical meristem, elongation zone and cortex. ET304 promoter sequence contained a canonic auxin response element (AuxRE) located upstream of the enhancer trap insertion site. In a synchronized lateral root induction system ET304 promoter conferred an auxin-responsive expression in newly emerging lateral roots of both poplar and Arabidopsis. A detailed histochemical examination of poplar transgenics showed that ET304 promoter was highly active in actively growing lateral roots, their primordia and to a lesser extent-in secondary meristems of aerial organs rich in free endogenous auxin. These results were consistent with the expression profiling of ET304 mRNA in various tissues of mature poplar trees. The analysis of Populus genomic sequence suggested that ET304 represents a large family of putative transcription factors containing a conserved AT-hook motif and DNA binding domain.