The cell wall of Arabidopsis thaliana influences actin network dynamics

Abstract

Number of tables: 3Word count: 5624 words Number of supplementary items: 7 Running titleCell wall affects actin cytoskeleton dynamics in A. thaliana HighlightActin cytoskeleton dynamics, as quantified with two new image analysis tools, were reduced in plasmolysed cells and cells treated with isoxaben to disrupt cell wall structure. AbstractIn plant cells, molecular connections link the cell wall-plasma membrane-actin cytoskeleton to form a continuum. It is hypothesised that the cell wall provides stable anchor points around which the actin cytoskeleton remodels. Here we use live cell imaging of fluorescently labelled marker proteins to quantify the organisation and dynamics of the actin cytoskeleton and to determine the impact of disrupting connections within the continuum. Labelling of the actin cytoskeleton with FABD2-GFP resulted in a network composed of fine filaments and thicker bundles that appeared as a highly-dynamic remodelling meshwork. This differed substantially from the GFP-Lifeact-labelled network that appeared much more sparse with thick bundles that underwent 'simple movement', in which the bundles slightly change position, but in such a manner that the structure of the network was not substantially altered during the time of observation. Label-dependent differences in actin network morphology and remodelling necessitated development of two new image analysis techniques. The first of these, Pairwise image subtraction , was applied to measurement of the more rapidlyremodelling actin network labelled with GFP-FABD2 while the second, Cumulative fluorescence intensity, was used to measure bulk remodelling of the actin cytoskeleton when labelled with GFP-Lifeact. In each case, these analysis techniques show that the actin cytoskeleton has a decreased rate of bulk remodelling when the cell wall-plasma membraneactin continuum is disrupted either by plasmolysis or with isoxaben, a drug that specifically inhibits cellulose deposition. Changes in the rate of actin remodelling also affect its functionality as observed by alteration in Golgi body motility.

Cell wall contributes to the stability of plasma membrane nanodomain organization of Arabidopsis thaliana FLOTILLIN2 and HYPERSENSITIVE INDUCED REACTION1 proteins

“…Increase in immobility may be connected with enhanced internalization in case of At HIR1–YFP and with accumulation of At FLOT2–GFP into sparser but more pronounced domains in these conditions. Plasmolysis induced by similar concentrations of mannitol or sorbitol caused dramatic changes in several cellular processes including reduced movement of endoplasmic reticulum structures (Cheng et al , ) or decreased actin dynamics (Tolmie et al , ) and polymerization (Yu et al , ). It is therefore possible that reported increase in mobility may not be a direct effect of PM detachment but rather a result of complex changes induced by plasmolysis in a cell which probably involve also cytoskeleton dynamics alteration that were in many cases also documented to affect PM protein mobility (see the Discussion above).…”

The cell wall regulates dynamics and size of plasma-membrane nanodomains in Arabidopsis

“…The plant cell wall is also required for normal localisation of PIN2 in the membrane and hence regulation of cell polarity (19). These examples highlight the possibility that the components of the cytoskeleton / PM / cell wall continuum can regulate each other, with cell-wall regulation of plasma-membrane, and cytoskeleton organization already observed (5, 20).…”

The Cytoskeleton and Its Role in Determining Cellulose Microfibril Angle in Secondary Cell Walls of Woody Tree Species

“…This continuum is presumably responsible for transmitting cell wall perturbations to the cytoskeleton resulting in reorganisation. In this respect, Tolmie et al [92] reported an increase in actin network stability in plasmolysed cells or cells treated with isoxaben, a cellulose synthesis inhibitor, and normal dynamics were recovered in re-hydrated cells, demonstrating changes in actin cytoskeleton dynamics were highly correlated with cell wall disruption. In combination, these findings suggest a role of the plant cytoskeleton in regulating cell wall biosynthesis by determining cellulose deposition sites and cellulose microfibril orientation through the cytoskeleton-plasma membrane-cell wall continuum.…”