Characterisation of Pt MYB1, an R2R3-MYB from pine xylem.

A cDNA encoding a member of the R2R3-MYB family of transcription factors was cloned from a library constructed from differentiating Pinus taeda (loblolly pine) xylem RNA. This MYB family member, Pinus taeda MYB1 (PtMYB1), was most abundantly expressed in differentiating xylem, as assessed by both ribonuclease protection assays, and by northern blot analysis with poly(A)-enriched RNA. Similar to other plant R2R3-MYB family members, recombinant Pt MYB1 protein was able to bind to AC elements in electrophoretic mobility shift assays (EMSAs). AC elements are DNA motifs rich in adenosine and cytosine that have been implicated in the xylem-localised regulation of genes encoding lignin biosynthetic enzymes. Pt MYB1 not only bound to AC elements, but was also able to induce AC-element-dependent shifts in the electrophoretic mobility of a plant promoter that contains three AC elements, the minimal PHENYLALANINE AMMONIA-LYASE 2 (PAL2) promoter from Phaseolus vulgaris. Transcriptional activation assays conducted using yeast showed that Pt MYB1 also activated transcription, and that it did so in an AC-element-dependent fashion. Pt MYB1 also activated transcription from the minimal PAL2 promoter in plant cells in an AC-element-dependent fashion, as revealed by transient transcriptional activation assays with microprojectile-bombarded tobacco NT-1 cells. Taken together, these finding are consistent with the hypothesis that Pt MYB1 may regulate transcription from cis -acting AC elements in pine xylem.