My laboratory's goal is to understand the mechanisms of plant development. To do this, we use both experimental and computational methods to test hypotheses. We concentrate on the shoot apical meristem (SAM) and its derivative structures (primarily flowers), because this meristem is responsible for the development of the entire aboveground part of the mature plant and utilizes a number of different pattern-forming processes that are poorly understood. Our experimental organism is predominantly Arabidopsis thaliana, because of the ease with which genetic and molecular biological studies can be done using this model system.
The Arabidopsis SAM, a collection of identical-appearing cells at the tip of each shoot, serves as the stem cell population for shoot, leaf, and flower development. The ultimate stem cells number around 20–30 and are at the tip of the meristem, a region called the central zone, which sits adjacent to the peripheral zone, from which leaves and flowers arise, and above the rib meristem, from which the stem and stem vasculature originate.
In the 1990s my laboratory identified genes that control the cell numbers in the central and rib regions. Molecular cloning of the genes showed that the central zone synthesizes a peptide ligand, CLAVATA3, that is secreted from the central zone cells and activates a transmembrane receptor kinase, CLAVATA1, in rib meristem cells, eventually causing reduction in the size of the rib meristem. This signaling network controls the size of the meristem.
One major question raised by the CLAVATA signaling system is how the gene expression domains are defined and maintained—that is, how is it that CLAVATA3 is expressed in (and defines) the cells of the central zone and that CLAVATA1 is expressed in (and defines) the cells of the rib meristem? Recently we discovered that the plant hormone cytokinin plays a role in this, and to understand this control we are finding the expression domains and interactions of the many components of the cytokinin signal transduction network.
Another set of problems posed by the actions of the SAM is its formation of leaf and flower primordia. We have analyzed the pattern in which new leaf and floral primordia arise in the meristematic peripheral zone. The origin of this pattern, the phyllotactic pattern, is a long-standing question in plant biology: the first mathematical approaches to this question were taken almost 150 years ago. In Arabidopsis and in most other plants, the phyllotactic pattern is a spiral with successive primordia appearing approximately 140 degrees radially from each other. By combining live imaging and computational modeling we have provided a detailed mechanistic and causal hypothesis for the phyllotactic pattern, based on feedback in the transport of the plant hormone auxin with its efflux carrier, the PINFORMED1 (PIN1) protein. Each premise of the model is explicit and tested, and together they provide a solution to the old problem of the formation of this characteristic plant pattern. The solution is a new class of developmental model—a regulated transport model—which adds to the two earlier-known classes of models for the generation of pattern by groups of cells (reaction-diffusion models and mutual inhibition models).
One premise of the phyllotaxis model raised a new set of possibilities for plant development. We and others have demonstrated that the PIN1 protein at the plasma membrane of each epidermal cell of the SAM is found adjacent to the neighboring cell with the highest auxin concentration; it is this feedback that gives the meristem its dynamic pattern-forming capabilities. Our experiments indicate that the mechanism used by a cell to sense the auxin concentration of its neighbor is physical: auxin causes meristem cells to expand by relaxing their cell walls, and the stress that such expansion creates on their neighbors creates signals for cellular polarization and for PIN1 relocalization. Further exploration of physical signals between meristem cells has shown that the microtubule cytoskeleton is also organized by physical stress (by a mechanism different from the one that organizes PIN1). As microtubules serve as guides for cellulose synthesis in the cell walls, and cellulose anisotropically reinforces the walls, stress input leads to changes both in auxin flow and cell wall reinforcement (and therefore direction of cell expansion), which in turn change the stress pattern, leading to a feedback between stress, cell expansion, and auxin signaling that explains much of the patterning of the meristem.
One additional property of SAMs is their ability to form de novo, during both embryogenesis and leaf growth and, even more remarkably, during regeneration in tissue culture. How a mass of cultured cells, callus, can give rise to highly organized and functioning shoot (and root) meristems is a mystery. One set of experiments in my laboratory is directed toward a mechanistic understanding of the processes of pattern formation in shoot regeneration. These studies are taking two directions. The first is to understand the nature of callus tissue, which according to the literature of plant tissue culture is an undifferentiated set of cells that forms and grows in response to hormonal cues, with the relevant hormones being auxin and cytokinins. We have recently shown that the literature is incorrect, and that Arabidopsis callus tissue—whether formed from roots, cotyledons, or petals—resembles lateral root primordia in its spatial patterns of gene expression and in the genetic requirements for its formation.
We have also started the analysis of the earliest events in SAM regeneration from growing callus tissue. Although we no longer consider this to be pattern formation from undifferentiated or dedifferentiated cells, it is still a remarkable example of transdifferentiation of root into shoot cells. We are using live imaging and laser scanning confocal microscopy to observe the changes in hormone response and gene activity as new meristems form. We also use computational models to test and suggest hypotheses. We have developed models for phyllotactic pattern and for stress-regulated changes in microtubules arrays, and for gene detailed expression as it is controlled by cell-cell signalling in the shoot meristem.
Grants from the Howard Hughes Medical Institute, Department of Energy, National Institutes of Health, and the Gordon and Betty Moore Foundation provided partial support for these projects.
As of February 15, 2016