Revealing the Architecture of the Cell Wall in Living Plant Cells by Bioimaging and Enzymatic Degradation

Plant cell walls consist mostly of crystalline cellulose fibrils embedded in a matrix of complex polysaccharides, but information on their morphological features has generally been limited to that obtained from nonliving plant specimens. Here, we characterized the primary cell wall of a living plant cell (from the tobacco BY-2 suspension culture) at nanometer resolution using high-speed atomic force microscopy and at micrometer resolution using confocal laser scanning microscopy. Our results showed aligned and disordered cellulose fibrils coexisting in the outermost layer of the cell wall. We investigated the orientation of the aligned cellulose fibrils in the outer lamellae of the cell wall of living plant cells after removing cellulose, hemicellulose, and pectin by enzymatic degradation to make the cellulose fibrils more visible and, accordingly, to reveal the structure of the nanoachitecture formed by these fibrils within the cell wall. We observed that the cellulose fibrils in the outermost layer were usually oriented close to the direction of cell growth, whereas the orientation of the cellulose fibrils in the successive lamellae further inward changed randomly. Such organization should be crucial to render the plant cell wall both rigid and flexible. This finding provides insight not only into the structure of the functional plant cell wall but also into its growth mechanism.