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John Fowler
Associate Professor, Botany and Plant Pathology
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RESEARCH:
A crucial element of plant morphogenesis is the control of the three-dimensional growth characteristics of cells: that is, the direction(s) in which existing cells expand and/or the relative position of new cells, as dictated by the placement of division planes. Thus, plant cells must respond to external signals to expand or divide in an oriented fashion. Cell polarity is implicated in diverse plant developmental processes, from pollen tube growth to the asymmetric cell divisions that generate stomatal complexes and two-celled embryos. The establishment of polarity, via specialized domains in the cytoplasm, plasma membrane, or cell wall of a cell, is an important means of providing three-dimensional information to direct subsequent morphogenesis.
Recent data in higher plants and in zygotes of fucoid algae (Fucus and Pelvetia) support the hypothesis that the actin cytoskeleton and secretion both play important roles in establishing plant cell polarity. Fucoid zygotes have a unique advantage for investigations into cell polarity: they establish oriented molecular and developmental asymmetries in culture media in response to external gradients (e.g., unilateral light) over the course of their first 24-hour cell cycle. Zygotes that are initially apolar orient the localized tip growth of the cell, as well as the first plane of division, with respect to the external vector, producing an embryo with two distinct cell types. An intact actin cytoskeleton and directed secretion are necessary for this process. However, the molecular mechanisms that direct secretion and cytoskeletal organization in plant cells are poorly understood.
The related, highly-conserved Rho and Rab families of small GTPases play important roles in the regulation of cytoskeletal organization and secretion, respectively, in fungal and animal cells. For example, the Cdc42p protein of S. cerevisiae (in the Rho family) is necessary for actin localization and polar growth at the yeast bud; the mammalian Rab8 protein is involved in the polarized transport of vesicles to the plasma membrane in epithelial cells. These proteins act as molecular switches, cycling between an active, GTP-bound form and an inactive, GDP-bound form and are promising targets for investigation of the role of the cytoskeleton and secretion in plant cell polarity.
We are using microinjection and antibody techniques to investigate the function and localization of Rho and Rab GTPases during embryogenesis in Fucus. Using these techniques, in conjunction with fluorescent probes that mark specific asymmetric events in Fucus, we hope to assemble a detailed description of the mechanisms involved in the polar development of this simple organism.
In higher plants, we are using a combined genetic and cell biological approach to study the role of cell polarity in development. We have isolated several Rho family genes from maize, an organism highly amenable to genetic studies. These cloned genes have allowed us to screen for and recover Mutator transposon insertions that inactivate the Rho genes. Currently, we are characterizing phenotypes associated with these mutations, and are targeting other Rho family genes for mutation. Investigation of mutant phenotypes, and localization of the gene products during the establishment of polarized processes (e.g. pollen tube growth, asymmetric division) will help us deduce Rho family functions in higher plant development.