KORRIGAN1 (KOR1) is a membrane-bound cellulase implicated in cellulose biosynthesis. PttCel9A1 from hybrid aspen (Populus tremula L. x tremuloides Michx.) has high sequence similarity to KOR1 and we demonstrate here that it complements kor1-1 mutants, indicating that it is a KOR1 ortholog. We investigated the function of PttCel9A1/KOR1 in Arabidopsis secondary growth using transgenic lines expressing 35S::PttCel9A1 and the KOR1 mutant line irx2-2. The presence of elevated levels of PttCel9A1/KOR1 in secondary walls of 35S::PttCel9A1 lines was confirmed by in muro visualization of cellulase activity. Compared with the wild type, 35S::PttCel9A1 lines had higher trifluoroacetic acid (TFA)-hydrolyzable glucan contents, similar Updegraff cellulose contents and lower cellulose crystallinity indices, as determined by (13)C solid-state nuclear magnetic resonance (NMR) spectroscopy. irx2-2 mutants had wild-type TFA-hydrolyzable glucan contents, but reduced Updegraff cellulose contents and higher than wild-type cellulose crystallinity indices. The data support the hypothesis that PttCel9A1/KOR1 activity is present in cell walls, where it facilitates cellulose biosynthesis in a way that increases the amount of non-crystalline cellulose.

Sphinganine or dihydrosphingosine (d18:0, DHS), one of the most abundant free sphingoid Long Chain Base (LCB) in plants, has been recently shown to induce both cytosolic and nuclear calcium transient increases and a correlated Programmed Cell Death (PCD) in tobacco BY-2 cells. In this study, in order to get deeper insight into the LCB signaling pathway leading to cell death, the putative role of Reactive Oxygen Species (ROS) has been investigated. We show that DHS triggers a rapid dose-dependent production of H₂O₂ that is blocked by diphenyleniodonium (DPI), indicating the involvement of NADPH oxidase(s) in the process. In addition, while DPI does not block DHS-induced calcium increases, the ROS production is inhibited by the broad spectrum calcium channel blocker lanthanum (La³+). Therefore, ROS production occurs downstream of DHS-induced Ca²+ transients. Interestingly, DHS activates expression of defense-related genes that is inhibited by both La³+ and DPI. Since DPI does not prevent DHS-induced cell death, these results strongly indicate that DHS-induced H₂O₂ production is not implicated in PCD mechanisms but rather would be associated to basal cell defense mechanisms.

Increases in the concentration of free calcium in the cytosol are one of the general events that relay an external stimulus to the internal cellular machinery and allow eukaryotic organisms, including plants, to mount a specific biological response. Different lines of evidence have shown that other intracellular organelles contribute to the regulation of free calcium homeostasis in the cytosol. The vacuoles, the endoplasmic reticulum and the cell wall constitute storage compartments for mobilizable calcium. In contrast, the role of organelles surrounded by a double membrane (e.g. mitochondria, chloroplasts and nuclei) is more complex. Here, we review experimental data showing that these organelles harbor calcium-dependent biological processes. Mitochondria, chloroplasts as well as nuclei are equipped to generate calcium signal on their own. Changes in free calcium in a given organelle may also favor the relocalization of proteins and regulatory components and therefore have a profound influence on the integrated functioning of the cell. Studying, in time and space, the dynamics of different components of calcium signaling pathway will certainly give clues to understand the extraordinary flexibility of plants to respond to stimuli and mount adaptive responses. The availability of technical and biological resources should allow breaking new grounds by unveiling the contribution of signaling networks in integrative plant biology.