Sum frequency generation (SFG) microscopy analysis of cellulose microfibrils in Physcomitrium patens gametophore leaf

Physcomitrium patens is a good model system to study the structure and function relationships of cellulose synthase (CESA) proteins since the life cycle of moss is short and genetic modification is relatively easy. Here, we report a microscopic sum frequency generation (SFG) vibrational spectroscopy analysis of P. patens leaves in a never-dried and fully hydrated state. Using imaging-SFG spectroscopy, cellulose microfibril (CMF) organization in the midrib, laminar, and marginal regions of the gametophore leaf were analyzed separately. The azimuth angle dependence of SFG signals showed that CMFs in the thick stereid cell walls in the midrib are highly aligned along the microfibril angle of similar to 30 degrees with respect to the midrib axis. CMFs in the marginal cell walls also are preferentially tilted toward the longitudinal axis of leaf with the characteristic features of secondary cell walls. On the other hand, CMFs in the lamina cell walls have no preferential orientation and their SFG intensity is low, which is consistent with the primary cell wall property. The thick walls and highly aligned CMFs in the midrib and marginal cells of P. patens leaves are consistent with their support function.

Automated summary

This study used microscopic sum frequency generation (SFG) vibrational spectroscopy to analyze the cellulose microfibril (CMF) organization in different regions of P. patens leaves. The results showed that CMFs in the midrib and marginal cell walls exhibited highly aligned and tilted characteristics, while CMFs in the lamina cell walls had no preferential orientation. These findings reveal the structural differences of cell walls in P. patens leaves and their functional implications.