IDENTIFICATION OF GENES CONTROLLING XYLEM CELL DIFFERENTIATION IN ARABIDOPSIS

Development of the vascular system in higher plants is a fundamental process that affects plant growth and yield. In addition, xylem production per se has an enormous economic impact. In fact, efficient water utilization, wood and fiber industry, biofuel production, food and feed digestibility largely depend on differentiation of lignified elements in the vascular apparatus specialized for water transport and mechanical support. The HD-ZIP III family of transcription factors is one of the most important and well characterized player that control vascular development (Myashima et al., 2012). ATHB8, in particular, is a marker of preprocambial cell state that accurately predicts sites of leaf vein formation and promotes proliferation and differentiation of xylem cells (Baima et al., 2001; Donner et al., 2009). Recently, the ACAULIS5 (ACL5) gene, encoding a polyamine synthase, has been involved in some aspects of xylem differentiation; it has been proposed that its product thermospermine, an isomer of spermine, is important to prevent premature cell death of tracheary elements (Knott et al., 2007; Muniz et al., 2008). In addition, it has been previously demonstrated that the acl5 mutant, also known as thickvein, causes the formation of an increased number of veins and vascular elements in leaves and stems (Hanzawa et al., 1997, 2000; Clay et al., 2005), likely by altering auxin signaling (Yoshimoto et al., 2012). More recently, taking advantage of several complementary experimental approaches, including ChIP, we demonstrated that ACL5 is directly regulated in a positive manner by ATHB8. We also proposed a model in which ATHB8 and other HD-ZIPIII transcription factors and ACL5 establish a negative feedback loop, likely connected to auxin, necessary for proper regulation of vascular cell differentiation (Baima et al., submitted). In the course of these studies, we found that transgenic plants expressing high levels of ACL5 in xylem precursor cells significantly delay or completely inhibit the differentiation of these cells into tracheary elements in leaf as well as in primary and secondary vasculature of stem and hypocotyl. Consistent with the proposed regulatory loop, gene expression analysis revealed that ATHB8 expression is down-regulated in these transgenic plants. To determine which genes involved in xylem cell differentiation are regulated by the ATHB8/ACL5 module, the transcriptome of transgenic lines enriched in undifferentiated xylem cells was compared to that of plants displaying a wild-type vasculature. The results of this analysis will be presented.