Generation of reiterative growth patterns in plants

Every flowering plant is made up of repeating units: leaves, stems, branches, flowers and root axes. Repetition may also occur within organs, as with the multiple lobes of an oak leaf. Such reiterations depend on growing regions, meristems, producing microscopic growth modules, which have stereotypic patterns of gene activity. Understanding how the initiation and spacing of growth modules is controlled is fundamental for our ability to predict the number, arrangement and shape of organs on a plant, key traits of agronomic importance. Two elements are at play in growth module control. One is restriction of growth module production to defined regions of the plant, termed competence zones. Secondly, growth modules interact with each other such that new modules are produced far away from previous ones, creating a spacing pattern. Hypotheses have been proposed for module-module interaction but we know much less about how the competence zone is controlled. This interdisciplinary project builds on preliminary results with a gene in Arabidopsis, CUC2. Normally CUC2 is active at the base of the leaf margin, where serration growth modules are produced. However, if CUC2 is activated throughout the leaf, serrations are produced all along the leaf margin, creating a fractal pattern (serrations upon serrations). These findings indicate that localised CUC2 activity normally restricts the zone of competence for module formation, and provide an opportunity to study the principles of competence control and how it interacts with module-module restriction.

We hypothesise that regulation of CUC2 restricts modules to basal regions of the leaf, and that another gene, KANADI, restricts them to the leaf margin. If CUC2 is overactive in a plant with reduced KANADI activity, we expect growth modules to be produced throughout the leaf surface and margin. Our first objective is to test this prediction in two species: Arabidopsis and barley. By comparing these two systems we aim to establish how generally principles apply across different taxonomic groups, and their relevance to crops. The second objective is to exploit the fractal behaviour exhibited by leaves with overactive CUC2 to test hypotheses for module-module interactions in a simplified context. By following the cells of these plants as the leaf develops, we will be able to establish rules for module-module spacing, how markers that distinguish different ends of the cell (polarity markers) change over time, how growth influences serration shape, how cell divisions are modulated, and how these properties vary between leaves with different extents of serration. The third objective is to induce over-active CUC2 at different times and places, to test ideas for which genes are activated by CUC2 and whether they are activated directly or via signalling between cells. We will also determine whether CUC2 activity in the leaf margin is critical for effects on growth, and test candidate signalling molecules. The fourth objective is to determine how CUC2 interacts with the plant growth hormone, auxin, which is known to influence growth module formation. This will be achieved by determining the effect of overactive CUC2 in plants where auxin transport is compromised by drugs or genetic mutations. The fifth objective is to develop computational models based on our experimental observations to clarify predictions of different hypotheses and thus discriminate between them.

Meeting these objectives will provide a deeper mechanistic understanding of how competence is controlled and interacts with module-module restrictions to generate reiterative growth patterns, underpinning our ability to predict and modify plant growth and architecture.

Plant growth depends on repeated production of growth modules from meristematic regions. This process is regulated by two mechanisms: restriction of module formation to a zone of competence and module-module restrictions that generate a spacing pattern. In this interdisciplinary project we study how these mechanisms interact, using induced ectopic expression of a micoRNA-resistant form of Arabidopsis CUC2 (CUC2-m4). Inducing CUC2-m4 throughout the leaf leads to growth modules forming along the entire leaf margin, indicating regulated CUC2 expression normally restricts competence to proximal regions in combination with margin identity factors.

Our first objective is to test the broad applicability of this hypothesis through ectopic expression of CUC2-m4 in a mutant background with ectopic margin identity in Arabidopsis and barley. The second objective is to exploit the fractal pattern generated by ectopic CUC2-m4 to study the mechanisms of module-module interaction in a simplifying context. By live imaging we will test rules for how modules are spaced, how orientations of growth and cell polarity change over time, how cell divisions are arrested, and how these features vary between leaves with different serration numbers. The third objective is to determine which target genes are activated by ectopic CUC2-m4 and whether activation is cell-autonomous or involves cell-cell signalling. We will also determine how CUC2 expression in the leaf margin influences growth and test the role of candidate signalling factors. The fourth objective is to determine how ectopic CUC2-m4 interacts with auxin to control module restriction. The fifth objective is to use computational modelling to integrate experimental findings and establish hypotheses that best explain them. The project will therefore provide a deeper understanding of how competence and module-module restrictions interact, underpinning our ability to predict and modify plant growth and architecture.