The mechanism of cellulose synthesis has been studied by characterizing the motility of cellulose synthase complexes tagged with a fluorescent protein; however, this approach has been used exclusively on the hypocotyl of Arabidopsis thaliana. Here we characterize cellulose synthase motility in the model grass, Brachypodium distachyon. We generated lines in which mEGFP is fused N-terminal to BdCESA3 or BdCESA6 and which grew indistinguishably from the wild type (Bd21-3) and had dense fluorescent puncta at or near the plasma membrane. Measured with a particle tracking algorithm, the average speed of GFP-BdCESA3 particles in the mesocotyl was 164 ± 78 nm min−1 (error gives standard deviation [SD], n = 1451 particles). Mean speed in the root appeared similar. For comparison, average speed in the A. thaliana hypocotyl expressing GFP-AtCESA6 was 184 ± 86 nm min−1 (n = 2755). For B. distachyon, we quantified root diameter and elongation rate in response to inhibitors of cellulose (dichlorobenylnitrile; DCB), microtubules (oryzalin), or actin (latrunculin B). Neither oryzalin nor latrunculin affected the speed of CESA complexes; whereas, DCB reduced average speed by about 50% in B. distachyon and by about 35% in A. thaliana. Evidently, between these species, CESA motility is well conserved.
(La0.9Sr0.1)0.95Cr0.85Mg0.10Ni0.05O3 (LSCMN) ceramics sintered at temperatures 1100–1700°C in air were characterized using powder x-ray diffraction, field emission scanning electron microscopy coupled with energy dispersive x-ray spectroscopy, transmission electron microscopy, differential scanning calorimetry, and inductively-coupled plasma spectroscopic analysis. Pervoskite ceramics with the highest density (porosity between 2–5%) were sintered at 1650°C for 24 hours. Secondary phases at a level of 3–5% porousity have been detected in the LSCMN initial powder received and sintered samples. LSCMN initial powder and ceramics exist in orthorhombic phase at room temperature and exhibits a first order phase transition into rhombohedral phase in the temperature range 70–95°C. Temperature of phase transition depends on grain size. Hardness and fracture toughness of LSCMN were studied by nanoindentation and microindentation methods. At low indentation depths hardness values depend significantly on the number of grains effected by the indent and crack formation. Indentation size effect was quantified in terms of Nix-Feng and power-low models. At high loads the apparent hardness is almost two times less than hardness of LSCMN monocrystalls.
The polysaccharide cellulose is the main component of plant cell walls, so it is the most abundant polymer on the earth. While it is widely used in industry due to its remarkable properties, such as renewability and biodegradability, its biosynthesis is still not well understood. The large transmembrane protein complex responsible for synthesizing cellulose contains several cellulose synthase A (CESA) subunits that polymerize UDP glucose into the constituent glucan chains of cellulose. Here, we used variable angle epi-fluorescence microscopy in combination with single-particle tracking to characterize the motion of GFP labeled CESA complexes in the root and mesocotyl of Brachypodium distachyon seedlings that are a 3 to 4 days old. We show that CESA complexes move through the plasma membrane at approximately 165 nm/minute. Their motion is known to be guided by cortical microtubules, but no molecular motors are involved. Rather, the motion is thought to be driven by the polymerization and crystallization of the cellulose. A mean-squared displacement analysis shows that CESA complexes move diffusively on short time scales and undergo a transition to super-diffusive motion on a time scale of about 10 s. We also report on the effect of actin and microtubule inhibitors on CESA motion.
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