Changes of the actin filament system in the green algaMicrasterias denticulata induced by different cytoskeleton inhibitors

Abstract

Summary. Two different techniques have been adapted for Micras-terias denticulata to depict the actin cytoskeleton of both untreated and inhibitor-treated developing cells: the "quickstaining method", where the cells are fixed in a mixture of glutaraldehyde and formaldehyde followed by staining with phalloidin without embedding, and the "methacrylate method", where the cells are also fixed by aldehydes and where the embedding medium is removed prior to incubation with an actin antibody. Both methods produce sufficient preservation and visualization of actin microfilaments (MFs) and confirm earlier observations on the presence of a cortical actin MF network in both the growing and the nongrowing semicell as well as of a basketlike MF arrangement around the migrating nucleus. The results show that a network of actin MFs is essential for the proper development of the young lobes of M. denticulata. Early developmental stages expanding uniformly at the beginning of growth lack any netlike actin MF arrangement. The actin cytoskeleton in developing cells treated with the actin-targeting agents cytocbalasin D and latrunculin B is markedly influenced. Cytochalasin D, which produces the most pronounced effects, causes a breakdown of the network of actin MFs, resulting in bright actin clusters as well as in short and abnormally thick actin fragments particularly in cortical cell regions. In latrunculin B-treated cells remnants of the former actin MF network are still visible, yet most of the actin cytoskeleton appears collapsed and is reduced to short filament pieces. The disturbance of the actin MF system visualized in the present study correlates with the severe morphological and ultrastructural changes occurring in desmid cells as a consequence of both drugs. The dinitroanilin herbicide oryzalin, known to deploymerize cytoplasmic microtubules, causes also an impairment of the actin cytoskeleton in M. denticulata though not sufficient to influence normal cell growth and differentiation.

Involvement of myosin in intracellular motility and cytomorphogenesis in Micrasterias

“…Then the cells were exposed to 0.33 µM Alexa phalloidin (Molecular Probes Eugene, OR, USA) containing 0.5% Triton X-100 in darkness (1 h 15 min), washed in destilled water and embedded in 5.5 mM p-phenylenediamine as antifade (method modified from Pflü gl-Haill et al, 2000). Cells were examined on a Zeiss Confocal Laser Scanning Microscope (LSM 510).…”

Chondramides, novel cyclodepsipeptides from myxobacteria, influence cell development and induce actin filament polymerization in the green algaMicrasterias

“…This green alga is subject to cell biological research for many years and serves as a model organism for investigating morphogenetic processes due to its highly symmetric pattern [e.g., Kiermayer, 1981;Meindl, 1993]. An important advantage of using Micrasterias cells is, that methods for actin visualization in fixed [Ueda and Noguchi, 1988;Pflügl-Haill et al, 2000] and in living cells [Meindl et al, 1994] are well established. It was found by the use of cytochalasins [Tippit and Pickett-Heaps, 1974;Lehtonen, 1983;Pflügl-Haill et al, 2000], jasplakinolide Holzinger, 2000], latrunculin B [Pflügl-Haill et al, 2000], or by microinjection of actin-binding proteins ] that F-actin plays a key role for the morphogenetic process in Micrasterias [for summary see Lütz-Meindl and Menzel, 2000].…”

“…4b). Immunofluorescence and Alexa™ 488 phalloidin staining allows depiction of a net-like F-actin arrangement in cortical regions of the growing and non-growing semicell of Micrasterias denticulata [Meindl et al, 1994;Pflügl-Haill et al, 2000] (Figs. 5a, 6a).…”

Nitric oxide suppresses growth and development in the unicellular green alga Micrasterias denticulata

“…The detailed knowledge about morphogenesis and development, secretion, cytoskeleton function, and cell wall formation of Micrasterias makes this organism attractive for physiological studies (Meindl, 1993;Pflügl-Haill et al, 2000;Oertel et al, 2004) and fulfills the prerequisite for our study to demonstrate the effect of nitric oxide on the secretory pathway, and, consequently, on cell growth and development.…”