Molecular mechanisms of vascular pattern formation

Vascular plants have developed a complex network of vascular systems through the plant body, allowing efficient transport of water, nutrients and signals. To understand molecular mechanisms of vascular pattern formation, we have made two approaches. First we have isolated Arabidopsis mutants with defects in vascular pattern formation. Microscopic and genetic examination of the cotyledonary venation of 3400 M3 lines led to the identification of 8 mutant lines whose abnormality was caused by mutations in 7 genetic loci designated VAN1-VAN7. Morphological analysis of van mutants indicated that vanl-van6 mutations caused fragmentation of lateral veins in cotyledons and of tertiary veins in rosette leaves, but did less injurious effects on the formation of their main veins or of vasculatures in hypocotyls and roots, van mutants were further characterized using pAthb8::GUS and pTED3::GUS as molecular markers of provascular cells and tracheary element precursor cells, respectively. As a result, it was revealed that most of van mutants lacked provascular cells at the disconnection points of the vascular network even at walking stick stage of embryogenesis. These results suggest that VAN genes are involved in the spatial control of provascular tissue differentiation, which realizes a continuous network of the vascular system. Second, we have analyzed regulation of cell-cell communication that may be involved in continuous formation of the vascular system using Zinnia cell culture. For this purpose, we developed two culture methods, thin-sheet culture and microbead culture. These culture methods indicated the presence of a high-molecular weight proteinaceous substance that promotes tracheary element differentiation. An improved microbead culture method brought about the partial purification of the substance, revealing that it is an arabinogalactan protein. Based on these results, we will discuss molecular mechanism of vascular pattern formation.