TOBIAS BASKIN'S LAB

1. Role of actin in mediating plant responses to auxin.
To better understand how auxin regulates root growth, we have quantified cell division, elemental elongation, and examined actin filaments in the primary root of Arabidopsis thaliana. In treatments for 48 h that inhibit root elongation rate by 50%, auxins and auxin-transport inhibitors can be put into two classes based on their effects on cell division, elongation, and actin organization. The auxins, IAA and NAA, and the auxin-transport inhibitor, TIBA, inhibit root growth primarily through reducing the length of the growth zone rather than the maximal rate of elemental elongation and they do not reduce cell production rate. These three compounds increase the fluorescence of filamentous actin, as imaged either with chemical fixation and immuno-cytochemistry or with reporter in living cells. In contrast, another auxin, 2,4-D, and and another inhibitor of transport, NPA, inhibit root growth primarily by reducing cell production rate. These compounds cause depolymerization of actin, stop cytoplasmic streaming, yet do not lead to the mislocalization of the putative efflux carrier, PIN2. The effects of 2,4-D and NPA on root growth as well as on actin were mimicked by a bona fide actin inhibitor, latrunculin B. This inhibitor stimulates actin depolymerization. There is a different actin inhibitor, called jasplakinolide, which stabilizes actin and causes extra polymerization. Too much actin is as deleterious as too little. The rotation project will be to quantify the effects of jasplakinolide, in comparison to previous results latrunculin and the other compounds. The project will involve high resolution analysis of root growth, confocal imaging of actin and PIN2, in both living and fixed cells, and real time measurements of endocytosis and cytoplasmic streaming.

2. How long is a cellulose microfibril?
This project aims to answer a fundamental question in plant biology; namely, how long is a microfibril. What is a microfibril? The plant cell wall is a composite material, resembling fiberglass. Both materials have stiff fibers embedded in an amorphous matrix. In the case of the cell wall, the stiff fibers are cellulose microfibrils. These are polymers of glucose linked end to end and many chains associate side-by-side to form a microfibril. The amorphous matrix is made up of the other polysaccharides and proteins in the cell wall. The reason we care about how long the microfibrils are is because the length of the stiff rod is a crucial parameter in determining the mechanical properties of the material. Certain models for the behavior of growing cell walls make predictions about microfibril length but so far these models have not been testable.

Recently, a protocol appeared for measuring microfibril length in another context. The protocol involves making a cell wall pellet, removing all material other than cellulose and then viewing the resulting microfibrils in the transmission electron microscope. The rotation project would involve adapting the protocol from tissue culture cells to a plant organ, such as a stem or root, and also to examine treatments that change the mechanical properties of the cell wall, changes that have been hypothesized to be controlled by the microfibril length. Another goal will be to scale the protocol down so that it works on sufficiently small amounts of material so that we can examine microfibril length in arabidopsis roots, where the phenotypes of certain mutants could be explained by altered microfibril length.

This project features biochemistry and electron microscopy.

3. What is the spatial profile of relative expansion rate of selected root morphology mutants?
A major research goal in my laboratory is to understand how a plant builds an organ with a specific shape. To that end, I have isolated a series of root morphology mutants, in which the shape of the root is aberrant. Some of these mutations turn out to be in cytoskeletal proteins, others in proteins active in cell wall synthesis; however, it is not always clear how the genetic defect generates the altered morphology of the root. The shape of the root is determined by expansion in length and width and these parameters vary at an essentially cellular scale. In my lab, we have developed a platform for high-resolution measurements of growth based on digital image processing. The project would be to use this platform to characterize the growth profile of the mutants at high resolution. Such characterization is required for us to understand the mutant phenotype clearly.

This project features computer-based image processing and requires a reasonable comfort level for mathematics.

I am also open to hosting rotations on other projects on-going in the lab. Please come see me if you would like further information (106 Morrill south; 5-1533; baskin@bio.umass.edu). Further information about my lab can be found at http://www.bio.umass.edu/biology/baskin/