Cellulose Synthase Stoichiometry in Aspen Differs from Arabidopsis and Norway Spruce.

Cellulose is synthesized at the plasma membrane by cellulose synthase complexes (CSCs) containing cellulose synthases (CESAs). Genetic analysis and CESA isoform quantification indicate that cellulose in the secondary cell walls of Arabidopsis (Arabidopsis thaliana) is synthesized by isoforms CESA4, CESA7, and CESA8 in equimolar amounts. Here, we used quantitative proteomics to investigate whether the CSC model based on Arabidopsis secondary cell wall CESA stoichiometry can be applied to the angiosperm tree aspen (Populus tremula) and the gymnosperm tree Norway spruce (Picea abies). In the developing xylem of aspen, the secondary cell wall CESA stoichiometry was 3:2:1 for PtCESA8a/b:PtCESA4:PtCESA7a/b, while in Norway spruce, the stoichiometry was 1:1:1, as observed previously in Arabidopsis. Furthermore, in aspen tension wood, the secondary cell wall CESA stoichiometry changed to 8:3:1 for PtCESA8a/b:PtCESA4:PtCESA7a/b. PtCESA8b represented 73% of the total secondary cell wall CESA pool, and quantitative polymerase chain reaction analysis of CESA transcripts in cryosectioned tension wood revealed increased PtCESA8b expression during the formation of the cellulose-enriched gelatinous layer, while the transcripts of PtCESA4, PtCESA7a/b, and PtCESA8a decreased. A wide-angle x-ray scattering analysis showed that the shift in CESA stoichiometry in tension wood coincided with an increase in crystalline cellulose microfibril diameter, suggesting that the CSC CESA composition influences microfibril properties. The aspen CESA stoichiometry results raise the possibility of alternative CSC models and suggest that homomeric PtCESA8b complexes are responsible for cellulose biosynthesis in the gelatinous layer in tension wood.

Immunomodulation of function of small heat shock proteins prevents their assembly into heat stress granules and results in cell death at sublethal temperatures.

The conformational dynamism and aggregate state of small heat shock proteins (sHSPs) may be crucial for their functions in thermoprotection of plant cells from the detrimental effects of heat stress. Ectopic expression of single chain fragment variable (scFv) antibodies against cytosolic sHSPs was used as new tool to generate sHSP loss-of-function mutants by antibody-mediated prevention of the sHSP assembly in vivo. Anti-sHSP scFv antibodies transiently expressed in heat-stressed tobacco protoplasts were not only able to recognize the endogenous sHSPs but also prevented their assembly into heat stress granula (HSGs). Constitutive expression of the same scFv antibodies in transgenic plants did not alter their phenotype at normal growth temperatures, but their leaves turned yellow and died after prolonged stress at sublethal temperatures. Structural analysis revealed a regular cytosolic distribution of stress-induced sHSPs in mesophyll cells of stress-treated transgenic plants, whereas extensive formation of HSGs was observed in control cells. After prolonged stress at sublethal temperatures, mesophyll cells of transgenic plants suffered destruction of all cellular membranes and finally underwent cell death. In contrast, mesophyll cells of the stressed controls showed HSG disintegration accompanied by appearance of polysomes, dictyosomes and rough endoplasmic reticulum indicating normalization of cell functions. Apparently, the ability of sHSPs to assemble into HSGs as well as the HSG disintegration is a prerequisite for survival of plant cells under continuous stress conditions at sublethal temperatures.