Studies of cell-cell communication during vascular formation

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1. Analysis with Arabidopsis vascular mutants. Van1-van7: We performed cloning of causal genes of Arabidqpsis vascular mutants, vanl-van6 showing fragmented vascular patterns that we have screened. VAN3 encoded a protein regulating the activation of small G-protein. VAN4 encoded a unknown protein, but which may function in membrane traffic. This gene was expressed preferentially hi vascular tissues.2. Analysis of vascular cell differentiation with an in vitro Zinnia system in which mesophyll cells differentiate into xylem cells.(1)We isolated ZePSKl, a homologue of phytosulfokine (PSK) from Zinnia. ZePSKl mRNA accumulated in early and late stages of xylem cell differentiation. Pulse treatment with an inhibitor of PSK biosynthesis and PSK revealed that PSK is necessary for progress of both early and late processes.(2) To identify factors controlling cell-cell communication during vascular cell development, we tried to isolate antibodies recognizing such factors. For such purpose, we used phage display subtraction method that we have developed. Finally we succeeded hi the isolation of about 8000 antibody clones. Tissue staining with randomly selected 90 antibodies revealed that these antibodies recognize apoplastic factors in vascular tissues differently. Thus we could get tools to find out factors involved of cell-cell communication.(3) We succeeded in the isolation encoding "xylogen" which we have identified as a tracheary element-inducing factor.2. Isolation and characterization of a ommature phloem-specific HD-Zip homeobox gene. We isolated a Zinnia HD- Zip class I gene whose mRNA accumulates specifically in immature phloem. Using this gene as a phloem marker, phloem regeneration was analyzed in wounded stems.