Four-dimensional quantitative analysis of cell plate development in Arabidopsis using lattice light sheet microscopy identifies robust transition points between growth phases.

Abstract

Cell plate formation during cytokinesis entails multiple stages occurring concurrently and requiring orchestrated vesicle delivery, membrane remodelling, and timely deposition of polysaccharides, such as callose. Understanding such a dynamic process requires dissection in time and space; this has been a major hurdle in studying cytokinesis. Using lattice light sheet microscopy (LLSM), we studied cell plate development in four dimensions, through the behavior of yellow fluorescent protein (YFP)-tagged cytokinesis-specific GTPase RABA2a vesicles. We monitored the entire duration of cell plate development, from its first emergence, with the aid of YFP-RABA2a, in both the presence and absence of cytokinetic callose. By developing a robust cytokinetic vesicle volume analysis pipeline, we identified distinct behavioral patterns, allowing the identification of three easily trackable cell plate developmental phases. Notably, the phase transition between phase I and phase II is striking, indicating a switch from membrane accumulation to the recycling of excess membrane material. We interrogated the role of callose using pharmacological inhibition with LLSM and electron microscopy. Loss of callose inhibited the phase transitions, establishing the critical role and timing of the polysaccharide deposition in cell plate expansion and maturation. This study exemplifies the power of combining LLSM with quantitative analysis to decode and untangle such a complex process.

Plant phenotyping relevance

LLSMによる4次元画像取得と、細胞板の小胞体積を定量化する解析パイプラインの開発が研究の中心であり、植物細胞の形態・発達状態を抽出する方法として substantive です。

Code and data availability

The paper deposits representative 4D lattice light sheet microscopy datasets (YFP–RABA2a cell plate imaging) used for its quantitative analysis on Zenodo, a paper-specific public asset. No author analysis code repository with explicit availability language is stated; the other URLs are method guidelines, not paper data