Dynamic apico-basal enrichment of the F-actin during cytokinesis in Arabidopsis cells embedded in their tissues

Abstract

Cell division is a tightly regulated mechanism, notably in tissues where malfunctions can lead to tumour formation or developmental defects. This is particularly true in land plants, where cells cannot relocate and therefore cytokinesis determines tissue topology. In plants, cell division is executed in radically different manners than in animals, with the appearance of new structures and the disappearance of ancestral mechanisms. Whilst F-actin and microtubules closely co-exist, recent studies mainly focused on the involvement of microtubules in this key process. Here, we used a root tracking system to image the spatio-temporal dynamics of both F-actin reporters and cell division markers in dividing cells embedded in their tissues. In addition to the F-actin accumulation at the phragmoplast, we observed and quantified a dynamic apico-basal enrichment of F-actin from the prophase/metaphase transition until the end of the cytokinesis.

Keywords: Arabidopsis; cell division; cytoskeleton; live cell imaging; root-tracking.

Graphical abstract

Figure 1.

Representatives images of a z-projection of a time-lapse analysis in Arabidopsis root meristem, expressing Ub10:Lifeact-YFPv. White arrows, F-actin enrichment at the apico-basal part of the cells; empty arrows, F-actin in the phragmoplast; n, nucleus. Scale bar = 5 μm.

Figure 2.

Representatives images of a z-projection of a time-lapse analysis in Arabidopsis root meristem, expressing mGFP-ABD2. White arrows, F-actin enrichment at the apico-basal part of the cells; empty arrows, F-actin in the phragmoplast; n, nucleus. Scale bar = 5 μm.

Figure 3.

(a) Representative images of a three-dimensional projection of a dividing cell in Arabidopsis root meristem expressing Ub10:LifeAct-2xmTU2. White arrows, F-actin enrichment at the apico-basal part of the cells; empty arrows, F-actin in the phragmoplast; n, nucleus. The yellow arrows marked the four rotated images obtained by the rotation in ImageJ of the initial z-projected image presented in the left panel. Scale bar = 10 μm. (b) Representative images of the distribution of LifeAct-2xmTU2 in dividing cell and non-dividing cells used for the quantification. Scale bar = 15 μm. (c) Quantification of the localization index of LifeAct-2xmTU2 between the apico-basal part of the cell and the lateral part of the cell of dividing versus non-dividing cells. (d) Quantification of the localization index of LifeAct-2xmTU2 between the apico-basal or the lateral part of the cell and the cytoplasm of dividing versus non-dividing cells. (e) Quantification of the localization index of LifeAct-YFPv between the apico-basal or the lateral part of the cells at key steps of the cell division, visualised thanks to the microtubule marker RFP-MBD. In the plots, the middle horizontal bars represent the median, whilst the bottom and top of each box represent the 25th and 75th percentiles, respectively. At most, the whiskers extend to 1.5 times the IQR and exclude data beyond that range. For the range of values under 1.5 times the IQR, the whiskers represent the range of maximum and minimum values.

Figure 4.

(a) Representatives images of a time-lapse analysis in Arabidopsis root meristem, expressing LifeAct-tdTom (green) together with KNOLLE-YFP (magenta). (b) Semi-quantitative measurement of the signal intensity for both LifeAct-tdTom (green) and KNOLLE-YFP (magenta) along the yellow line. White arrows, F-actin enrichment at the apico-basal part of the cells; orange arrows, accumulation of F-actin in the phragmoplast; n, nucleus. Scale bar = 5 μm.

Figure 5.

Representatives images of a time-lapse analysis in Arabidopsis root meristem, expressing LifeAct-tdTom (green) together with GFP-PHGAP1 (magenta). Yellow and white arrows, F-actin enrichment at the apico-basal part of the cells; empty arrows, the position of the cortical division zone; n, nucleus. Scale bar = 5 μm.

Figure 6.

(a) Representatives images of a time-lapse analysis in Arabidopsis root meristem, expressing LifeAct-YFPv (green) together with RFP-MBD (magenta). White arrows, F-actin enrichment; empty arrows, the phragmoplast; blue arrowhead, the position of the F-actin in the phragmoplast; n, nucleus. Scale bar = 10 μm. (b) Semi-quantitative analysis of the fluorescence intensity corresponding to LifeAct-YFPv (green) and RFP-MBD (magenta) in a rectangular ROI (h22xw106 pixels) represented in yellow on the merged image in (a). The grey area corresponds to the external part of the dividing cell and the blue arrowhead corresponds to the position of the F-actin in the phragmoplast.