| Summary: | Transcript profiling has the potential to reveal transcriptional networks operating during development and provides expression data for genes of unknown function. Based on previous studies employing global transcription profiling of Arabidopsis thaliana inflorescence stem development and A. thaliana root cell type-specific expression, ten candidate transcription factor (TF) genes potentially associated with secondary wall formation and lignification were identified. Similar transcript profiling experiments had revealed gene expression patterns associated with secondary wall formation during secondary xylem formation in Populus species. I verified gene expression patterns of the poplar putative orthologs of the A. thaliana candidates, and the combination of the A. thaliana and poplar data identified a subset of conserved MYB and homeodomain TFs that behaved similarly in both plants.
I analyzed the expression patterns of promoter-GUS fusions of all candidate genes in A. thaliana, most of which showed expression in xylem and cortex cells adjacent to interfascicular fibers in the stem, and in the root stele. T-DNA insertion mutants for most candidate genes were characterized, but only homeodomain transcription factor KNAT7 (At1g62990) T-DNA insertion mutants exhibited an obvious phenotype in inflorescence stems. The knat7 phenotype is characterized by irregular xylem (irx), interfascicular fibers with thicker cell walls, and defects in secreted seed mucilage. I also assayed for changes in gene expression in the knat7 background, and the results suggested that KNAT7 directly or indirectly regulates lignin biosynthesis genes.
KNAT7 is known to interact with members of the Ovate Family Protein (OFP) transcription co-regulators. I confirmed the KNAT7-OFP1 and KNAT7-OFP4 interactions in planta, and showed that the interaction enhances KNAT7 transcriptional repression activity. Furthermore, an ofp4 mutant exhibits similar phenotypes as knat7, and the pleiotropic effect of OFP1 and OFP4 overexpression depends upon KNAT7 function. A knat7/ofp4 double mutant showed a very similar phenotype to the single mutants, supporting the hypothesis that the two proteins work in the same pathway. KNAT7 thus appears to form a functional complex with OFP proteins, and may directly or indirectly regulate lignin biosynthesis through interaction with OFP family members. These investigations provide new insights into the regulatory network(s) governing secondary wall biosynthesis in A. thaliana and poplar.
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