Growth and Development of higher plants

The linear organization of a root tip, with cells organised in discrete files radiating from the quiescent centre and with successive regions of meristem, elongation zone and mature cells make it an ideal system to investigate the cellular and molecular basis of plant growth regulation. Moreover, the minimal number of cell layers in the roots of Arabidopsis limits its thickness to only 150 microns, which facilitates microscopical analysis by means of light (using DIC optics) and confocal microscopy (using fluorescent dyes or GFP tagged lines). To investigate the relationships between the cellular processes cell division and cell expansion and growth of the root as a whole, kinematic analyses have been developed based on fluid dynamics. P1C has successfully pioneered the practical use of these methods for the studies of cell division and cell expansion rates in Arabidopsis roots analyzing the effect of stress conditions (salt stress), natural (ecotypes) and engineered genetic variation (mutants and transgenic lines). This state-of-the-art kinematic platform has been extensively used during the previous phase of this network by various partners and will be again crucial for understanding root growth phenotypes obtained in this project.
These analyses unequivocally established the importance of cell division parameters (cell division rate and size of meristem) in determining root growth rates. Therefore P1C conducted a gene discovery experiment using genome-wide Affymetrics microarrays, analyzed gene expression in three zones of the root tip (meristem, elongation zone and mature tissue) and compared with corresponding stages of developing leaves. This way P1C was able to identify 430 genes that were specifically expressed in proliferating tissues. A large number of those genes are already known and almost without exception have a direct link to cell division. Therefore the unknown genes in this selection are strong candidates for new functions in cell cycle and plant growth regulation and they will be functionally analyzed in the framework of this project.
In this workpackage, P1C will generate overexpressing lines, knock-outs or knock-downs using publicly available T-DNA mutant collections of a selection of these putative new 430 cell division genes. Preferentially unknown genes will be selected based on criteria such as fold-change, number of homologs in Arabidopsis, presence of homologs in the human genome and type of the protein that is encoded (e.g. transcription factors, kinases and completely unknowns). These genes will be functionally analyzed. The phenotype of such lines will be studied using an array of tools available, among others including kinematic analysis, transcript profiling and flow-cytometry that will facilitate the identification of the function of the gene involved.
This analysis will no doubt increase further the already large number of genes involved in cell cycle regulation. Together these gene products form a regulatory network that is complex and often behaves in unexpected ways. To start addressing this complexity in the context of a growing multi-cellular organ, it is necessary to develop computer models that simulate experimentally determined and hypothetical interaction mechanisms (see WP 6, P1A, P1B, P4C, EU1).
Furthermore, P1C will continue performing kinematic analysis of genetic material and environmental or physiological treatments that are relevant to the project (P5, EU1).