The detergent-resistant cytoskeleton of higher plant protoplasts contains nucleus-associated fibrillar bundles in addition to microtubules

Abstract

Cytoskeletons prepared from carrot protoplasts often retain the nucleus, which is associated with bundles of 7 nm fibrils. These were also seen in cytoskeletons from oil palm, maize and Italian rye grass cells. By examining such cytoskeletons by immunofluorescence, negative staining and stereo scanning electron microscopy it appears that the bundles could be part of a more delicate system of fibrils distal to the nucleus. Optical diffraction of the bundles, staining with heavy meromyosin, drug treatments and phalloidin-staining all confirm that the bundles are not composed of actin. Bundles could not be isolated from such cytoskeletons in quantities sufficient for analysis, but by disrupting protoplasts prepared from stationary-phase cells it was possible to purify fibrillar bundles that appeared in the lysate. These were similar in size and properties to the nucleus-associated bundles. The former were shown by sodium dodecyl sulphate/polyacrylamide gel electrophoresis to be composed of several proteins and were sensitive to reducing agents. Since they are not composed of actin, the possibility was explored that these fibrils are related to the P-proteins of phloem sieve tube and accessory cells.