Cytoplasmic microtubules of higher plant cells visualised with anti-tubulin antibodies

Abstract

IT has been known for some time from electron microscopy that eukaryotic cells contain a fibrillar cytoskeleton. Recently, immunofluorescence has greatly advanced our understanding of how the cytoskeleton functions in controlling the shape and behaviour of cells¹ by allowing immunological identification and by displaying the three-dimensional distribution of the constituent proteins. As plant cell walls exclude antibodies, such work has been carried out almost entirely on animal cells. It is not possible to extrapolate from animals to plants because the problems are not the same: plant microtubules occur in cortical hoops at right angles to the axis of cell elongation unlike, say, neuronal outgrowth; they overlap to form bundles; they do not appear to grow from focal points around the nucleus and they are conspicuously cross-bridged to each other and to the plasma membrane². In a rare study on higher plant cells, Franke et al.³ made use of the immunological cross-reactivity between mammalian anti-tubulin and plant microtubules to study the giant mitotic apparatus in the endosperm of the March cup flower. However, such temporary nutritive tissue may have no significant wall deposition and although ideal for studying the nucleus is unsuitable for studying the cytoplasmic microtubules involved with cellulose in controlling plant cell shape. Franke et al.³ saw no cytoplasmic microtubules and for the study reported here we therefore used single, elongated cells from carrot suspension cultures which we know from electron microscopy (EM) studies (unpublished) to contain cytoplasmic microtubules. A controlled cellulase treatment and detergent extraction to allow the entry of antibodies to these cells was used, and we believe this to be the first report of the immunofluorescent staining of cytoplasmic microtubules in higher plant cells.