Research Theme 1:
How do plant cells polarize before dividing asymmetrically?
|
Higher plants and animals are made of different organs and tissues that all have specialized jobs. Within tissues, cells must be arranged properly to correctly form these tissues such that they function correctly. The arrangement of cells relative to one another and the specific function a cell acquires are tightly linked through asymmetric cell division. Prior to asymmetric division, cells must polarize. We study the process of how cells receive signals that induce them to polarize, and the subsequent cellular processes required to carry out polarization.
|
Figure 1: Maize cells expressing YFP-tubulin (green) and BRK1-CFP (magenta). The three left-most cells have polarized and are in preprophase. The three right most cells are in the process of, or have just completed, an asymmetric division. The cells are either in telophase (top), anaphase (middle) or recently completed cytokinesis (bottom).
|
Plant stomata and the power of corn
Figure 2. Mature maize epidermis stained with propidium iodide to highlight cell walls and nuclei.
|
The Facette lab studies asymmetric divisions in plant cells. Unlike animal cells, which can move, plant cells are fixed in place. Therefore cell division and differential cell expansion are the only ways cells can organize themselves within a tissue.
We study the formation of stomata in the maize (corn) leaf epidermis. In maize and other grasses, the stomata are composed of 4 cells: 2 guard cells flanked by 2 subsidiary cells (Figure 2). The divisions that form this 4-cell complex are highly stereotypical. Moreover, the leaves of maize and other grasses develop in a linear gradient, where new cells are "born" at the leaf base, and the most mature cells are at the leaf tip. This allows us to examine the development of these cells in a single piece of epidermis . |
BRK and PAN protiens: Molecular players in the asymmetric divisions that form maize stomata
|
We focus on the molecular signalling proteins that are required for the mother cells that form subsidiaries cells to polarize before they divide asymmetrically. These proteins include: BRK1 and BRK3, members of the evolutionarily conserved SCAR/WAVE complex that is a major actin nucleator; and PAN1 & PAN2, receptor-like proteins that act down-stream of BRK proteins. ROPs are small GTPases that act downstream of PAN protiens that are also required for polarization. In brk, pan or rop mutants, subsidiary mother cells fail to polarize resulting in abnormal stomata (Figure 3).
To study the cell polarization process in maize, we use live cell markers and microscopy, mutational analyses, and biochemical assays. We are currently identifying new players in the pathway through a combination of forward genetics (through an enhancer screen) and reverse genetics (using CRISPR-Cas9 or transposons to knock out proteins that interact with pan1, pan2 or brk1). Several different projects are available in the Facette lab to study plant cell polarization. |
Figure 3. Normal stomata in wildtype (top) and abnormal stomata in pan2 mutants (bottom).
|
Research Theme 2:
How do subsidiary cells function in the grass stomatal complex?
Grass stomata open and close quickly, presumably due in part to the action of the subsidiary cells that flank the guard cells, although the molecular mechanisms of how subsidiary cells might do this are unknown
Are stomatal movements affected in pan and brk mutants?
We are trying to understand if plants with aberrant subsidiary cells open and close more slowly in response to environmental cues. We are examining this in the juvenile leaves (i.e., the first ~5 leaves of the maize plant) where many subsidiary cells are aberrantly shaped in these mutants, as well as in adult leaves (i.e., approximately leaves 7 and older), which have fewer aberrant cells. Studies from other plants suggest molecules affecting actin nucleation that operate with BRK proteins, as well as a receptor protein similar to PAN2 are involved in stomatal function.
Do stomatal variants in maize perform differently?
|
Figure 4. Stomata from 5 different maize inbreds, with different subsidiary cell and guard cell shapes and sizes.
|
Since little is known about the mechanism of subsidiary cell interaction, and the subsidiary cell-guard cell interaction, we are examining the behavior of stomata in diverse maize inbred lines. We are determining if there are morphological features (such as size or shape) or genetic features (gene expression or gene variants) that contribute to efficient stomatal opening and closing. Ultimately, we will determine if variation in stomatal function translates to differences in plant water use efficiency.
