Phloem loading in rice leaves depends strongly on the apoplastic pathway

Phloem loading is the first step in sucrose transport from source leaves to sink organs. The phloem loading strategy in rice remains unclear. To determine the potential phloem loading mechanism in rice, yeast invertase (INV) was overexpressed specifically in the cell wall by 35S promoter to block sugar transmembrane loading in rice. The transgenic lines exhibited obvious phloem loading suppression characteristics accompanied by the accumulation of sucrose and starch, restricted vegetative growth and decreased grain yields. The decreased sucrose exudation rate with p-chloromercuribenzenesulfonic acid (PCMBS) treatment also indicated that rice actively transported sucrose into phloem. Moreover, the expression level of OsSUT1 was much higher than that of other plasma membrane localized OsSUTs in the source leaf. Cross sections of the GUS transgenic plant showed that the signals of OsSUT1 and OsSUT5 occurred in the phloem companion cells. The ossut1 and ossut4 mutants presented a decrease of grain yield, implying important roles of OsSUTs in phloem loading. Based on these results, we conclude that rice uses the apoplastic loading as a major phloem loading strategy.

A Novel Method for the Quantification of White Wine Mannoproteins by a Competitive Indirect Enzyme-Linked Lectin Sorbent Assay (CI-ELLSA)

Mannoproteins (MPs) are cell wall proteoglycans released in wine by yeast during fermentation and ageing on lees, a procedure used for the production of several wines to enrich them in these components with consequences from both a technological and sensory point of view. Given the significance that wine MPs have for wine quality, winemakers would welcome a simple and accurate method for their quantification, as this would allow them to have a better control of this aspect at different winemaking stages. This study develops and validates a novel, simple and accurate method for MPs quantification in white wines based on a competitive indirect enzyme-linked lectin sorbent assay (CI-ELLSA), using the highly mannosylated yeast invertase as the standard. The method utilizes the lectin concanavalin A (ConA) as the immobilized ligand for MPs, and peroxidase, an enzyme rich in mannose, as the competitor for ConA. After addition of the peroxidase substrate, the intensity of the signal produced by the activity of this enzyme (absorbance at 450 nm) is inversely proportional to the amount of mannosylated proteins in the sample. Results have been validated on several wine styles including still, sparkling and sweet wines.

BLACK AND WHITE LAHAR AS INORGANIC SUPPORT FOR THE IMMOBILIZATION OF YEAST INVERTASE

Volcanic ejecta or lahar can serve as an inorganic support for the immobilization of invertase. Pampanga and Bicol lahar samples were pretreated by ignition at 550oC for 5 hrs followed by concentrated hydrochloric acid treatment, activatedby reaction with aminopropyltriethoxysilane (APTS) and then covalently bound to invertase using glutaraldehyde as linker. Chemical tests confirmed the attachment of APTS to lahar and glutaraldehyde to silanized lahar. The quantity of immobilized invertase on Pampanga white, Pampanga black and Bicol black lahar were 98.73%, 96.73% and 84.27%,respectively. Conditions for maximum activity of invertase immobilized on Pampanga white lahar were pH 3.5, 45oC and 0.3 M sucrose concentration. The Kmand Vmaxfor free invertase and immobilized invertase on Pampanga white lahar were 2.37 M and 48.75 mmol/min, and 3.88 M and 38.87 mmol/min, respectively. Invertase bound to Pampanga white lahar was most stable towards repeated and continuous use and towards storage with intermittent use as indicated by its relatively greater activity.