Microtubular and Cytoskeletal Mutants

Summary

Microtubules are biochemically one of the simplest, yet functionally most important, cellular organelles in the plant and animal kingdoms. They are integral components of a dynamic, three-dimensional framework referred to as the cytoskeleton. In addition to a fundamental role in intracellular movement, the microtubule, microfilament and intermediate filament arrays of the cytoskeleton are networks upon which asymmetric distribution of subcellular constituents is established and from which these polarized regulatory molecules mediate morphogenesis. The functions of microtubules reflect this common theme of distribution and movement. In plants, four principal microtubular functions are recognized: determination of the division plane; translocation of chromosomes; cell plate/phragmoplast formation; and control of cell morphology. Obviously, from a biotechnological point of view, the ability to control and regulate these processes via modifications of the component parts will have a profound effect on plant growth and development. Microtubules are composed primarily of a single, repeating macromolecular unit — tubulin. Tubulin itself is a heterodimeric protein, composed of two similar subunits: alpha-(α) and beta-(β)tubulin. The α- and β-tubulins are typically encoded by gene families, and these give rise to various tubulin isotypes that are differentially expressed and modified during growth and development. In addition, a ubiquitous and diverse class of proteins that bind to microtubules, known as microtubule associated proteins (MAPs), are believed to be important in nucleation, stabilization and bundling of microtubules.

Knowledge of the cytoskeleton and its fundamental role in cell morphogenesis and intracellular movement is central to an understanding of plant growth and development. Tubulin mutants have proven extremely useful for the analysis of microtubule and cytoskeletal function in algal, fungal and higher plant cells. The best-characterized mutations typically alter microtubule stability in the presence of numerous agents such as temperature, the antimicrotubular fungicides and herbicides, as well as various antitumour drugs (see Chap. 9). Tubulin gene mutations that produce non-conservative amino acid substitutions in tubulin proteins, and in turn may cause changes in the electrophoretic pattern of tubulin subunits, have been identified in Chlamydomonas reinhardtii, Saccharomyces cerevisiae, Aspergillus nidulans, Neurospora crassa and the higher plants Eleusine indica (goosegrass) and Nicotiana plumbaginifolia. Only goosegrass appears to possess naturally occurring site of action mutations. However, at least six other species of higher plants have been reported to have resistant biotypes, and thus may harbor similar mutations. Although these mutations can occur in either tubulin subunit, there may very well be a limited number of mutable sites resulting in phenotypes detectable as alterations in microtubule stability (e.g. sites that are critical for monomer-monomer or dimer-dimer binding). In Chlamydomonas, for example, single amino acid substitutions in the α- or β-tubulin subunits confer resistance to the dinitroaniline herbicides, increase microtubule stability and produce electrophoretic differences in the respective proteins as compared to wild-type tubulin. This chapter will review our current knowledge of mutant tubulins and other mutants suspected of affecting microtubule or cytoskeletal functioning in seed plants, algae and fungi. Also, it will explore and speculate on the biotechnological applications of these mutants.