Comprehensive and quantitative analysis of intracellular structure polarization at the apical-basal axis in elongating Arabidopsis zygotes.

A comprehensive and quantitative evaluation of multiple intracellular structures or proteins is a promising approach to provide a deeper understanding of and new insights into cellular polarity. In this study, we developed an image analysis pipeline to obtain intensity profiles of fluorescent probes along the apical-basal axis in elongating Arabidopsis thaliana zygotes based on two-photon live-cell imaging data. This technique showed the intracellular distribution of actin filaments, mitochondria, microtubules, and vacuolar membranes along the apical-basal axis in elongating zygotes from the onset of cell elongation to just before asymmetric cell division. Hierarchical cluster analysis of the quantitative data on intracellular distribution revealed that the zygote may be compartmentalized into two parts, with a boundary located 43.6% from the cell tip, immediately after fertilization. To explore the biological significance of this compartmentalization, we examined the positions of the asymmetric cell divisions from the dataset used in this distribution analysis. We found that the cell division plane was reproducibly inserted 20.5% from the cell tip. This position corresponded well with the midpoint of the compartmentalized apical region, suggesting a potential relationship between the zygote compartmentalization, which begins with cell elongation, and the position of the asymmetric cell division.

Coordinate Normalization of Live-Cell Imaging Data Reveals Growth Dynamics of the Arabidopsis Zygote.

Polarization of the zygote defines the body axis during plant development. In Arabidopsis (Arabidopsis thaliana), the zygote becomes polarized and elongates in the longitudinal direction, ultimately forming the apical-basal axis of the mature plant. Despite its importance, the mechanism for this elongation remains poorly understood. Based on live-cell imaging of the zygote, we developed new image analysis methods, referred to as coordinate normalization, that appropriately fix and align positions in an image, preventing fluctuation across a temporal sequence of images. Using these methods, we discovered that the zygote elongates only at its apical tip region, similar to tip-growing cells such as pollen tubes and root hairs. We also investigated the spatiotemporal dynamics of the apical tip contour of the zygote and observed that the zygote tip retains its isotropic, hemispherical apical shape during cell elongation. By looking at the elliptical fitting of the contour over time, we further discovered that the apical cell tip becomes thinner at first and then thickens, with a transient increase in growth speed that is followed by the first cell division. We performed the same series of analyses using root hairs and established that both the hemispherical tip shape and the changes in growth rate associated with changes in tip size are specific to the zygote. In summary, the Arabidopsis zygote undergoes directional elongation as a tip-growing cell, but its tip retains an unusual isotropic shape, and the manner of growth changes with the developmental stage.