Raman imaging of Micrasterias: new insights into shape formation.

The algae Micrasterias with its star-shaped cell pattern is a perfect unicellular model system to study morphogenesis. How the indentations are formed in the primary cell wall at exactly defined areas puzzled scientists for decades, and they searched for chemical differences in the primary wall of the extending tips compared to the resting indents. We now tackled the question by Raman imaging and scanned in situ Micrasterias cells at different stages of development. Thousands of Raman spectra were acquired from the mother cell and the developing semicell to calculate chemical images based on an algorithm finding the most different Raman spectra. Each of those spectra had characteristic Raman bands, which were assigned to molecular vibrations of BaSO4, proteins, lipids, starch, and plant cell wall carbohydrates. Visualizing the cell wall carbohydrates revealed a cell wall thickening at the indentations of the primary cell wall of the growing semicell and uniplanar orientation of the cellulose microfibrils to the cell surface in the secondary cell wall. Crystalline cellulose dominated in the secondary cell wall spectra, while in the primary cell wall spectra, also xyloglucan and pectin were reflected. Spectral differences between the indent and tip region of the primary cell wall were scarce, but a spectral mixing approach pointed to more cellulose fibrils deposited in the indent region. Therefore, we suggest that cell wall thickening together with a denser network of cellulose microfibrils stiffens the cell wall at the indent and induces different cell wall extensibility to shape the lobes.