No abstract

Spring wheat (Triticum aestivum L. cv. TRISO) was grown for three consecutive seasons in a free‐air carbon dioxide (CO2) enrichment (FACE) field experiment in order to examine the effects on crop yield and grain quality. CO2 enrichment promoted aboveground biomass (+11.8%) and grain yield (+10.4%). However, adverse effects were predominantly observed on wholegrain quality characteristics. Although the thousand‐grain weight remained unchanged, size distribution was significantly shifted towards smaller grains, which may directly relate to lower market value. Total grain protein concentration decreased significantly by 7.4% under elevated CO2, and protein and amino acid composition were altered. Corresponding to the decline in grain protein concentration, CO2 enrichment resulted in an overall decrease in amino acid concentrations, with greater reductions in non‐essential than essential amino acids. Minerals such as potassium, molybdenum and lead increased, while manganese, iron, cadmium and silicon decreased, suggesting that adjustments of agricultural practices may be required to retain current grain quality standards. The concentration of fructose and fructan, as well as amounts per area of total and individual non‐structural carbohydrates, except for starch, significantly increased in the grain. The same holds true for the amount of lipids. With regard to mixing and rheological properties of the flour, a significant increase in gluten resistance under elevated CO2 was observed. CO2 enrichment obviously affected grain quality characteristics that are important for consumer nutrition and health, and for industrial processing and marketing, which have to date received little attention.

In preparation for being shed into the environment as infectious cysts, trophozoites of Giardia spp. synthesize and deposit large amounts of extracellular matrix into a resistant extracellular cyst wall. Functional aspects of this developmentally regulated process were investigated by expressing a series of chimeric cyst wall protein 1 (CWP1)-green fluorescent protein (GFP) reporter proteins. It was demonstrated that a short 110 bp 5Ј flanking region of the CWP1 gene harbors all necessary cis-DNA elements for strictly encystation-specific expression of a reporter during in vitro encystation, whereas sequences in the 3Ј flanking region are involved in modulation of steady-state levels of its mRNA during encystation. Encysting Giardia expressing CWP1-GFP chimeras showed formation and maturation of labeled dense granule-like vesicles and subsequent incorporation of GFP-tagged protein into the cyst wall, dependent on which domains of CWP1 were included. The N-terminal domain of CWP1 was required for targeting GFP to regulated compartments of the secretory apparatus, whereas a central domain containing leucine-rich repeats mediated association of the chimera with the extracellular cyst wall. We show that analysis of protein transport using GFP-tagged molecules is feasible in an anaerobic organism and provides a useful tool for investigating the organization of primitive eukaryotic vesicular transport. INTRODUCTIONGiardia duodenalis (syn. G. intestinalis, G. lamblia) is a flagellated protozoan parasite of medical significance because it is a major cause of waterborne enteric disease worldwide (Adam, 1991). The biology of this parasite is of particular interest because it represents one of the earliest branching lineages of eukaryotic descent, based on phylogenetic analysis of ribosomal and protein coding sequences (Sogin et al., 1989;Leipe et al., 1993;Gupta et al., 1994;Drouin et al., 1995;Roger et al., 1999). Giardia also lacks organelles typical for higher eukaryotes such as mitochondria and peroxisomes. Asexually dividing, motile trophozoites colonize the upper intestine of humans and other mammals, where they attach to the intestinal epithelium (Adam, 1991). Encysting Giardia trophozoites undergo a series of developmental changes in the course of which they synthesize, export, and assemble an extensive fibrillar extracellular matrix composed of proteins but also a large proportion of carbohydrate, mainly in the form of N-acetylgalactosamine (Manning et al., 1992). Formation of environmentally resistant cysts is a key step in Giardia development as in other eukaryotic pathogens (Eichinger, 1997;Taratuto and Venturiello, 1997;Dubey et al., 1998) that has not been well characterized on a molecular level. Therefore, a better understanding of the mechanisms that govern stage-specific regulation of genes and the synthesis, transport, and assembly of the cyst wall is needed.Several genes that are up-regulated during Giardia encystation have been identified recently by biochemical approaches, molecular genetics, or metab...

No abstract

Transmission of the protozoan parasite Giardia intestinalis to vertebrate hosts presupposes the encapsulation of trophozoites into an environmentally resistant and infectious cyst form. We have previously shown that cyst wall proteins were faithfully sorted to large encystation-specific vesicles (ESVs), despite the absence of a recognizable Golgi apparatus. Here, we demonstrate that sorting to a second constitutively active pathway transporting variant-specific surface proteins (VSPs) to the surface depended on the cytoplasmic VSP tail. Moreover, pulsed endoplasmic reticulum (ER) export of chimeric reporters containing functional signals for both pathways showed that protein sorting was done at or very soon after export from the ER. Correspondingly, we found that a limited number of novel transitional ER-like structures together with small transport intermediates were generated during encystation. Colocalization of transitional ER regions and early ESVs with coat protein (COP) II and of maturing ESVs with COPI and clathrin strongly suggested that ESVs form by fusion of ER-derived vesicles and subsequently undergo maturation by retrograde transport. Together, the data supported the hypothesis that in Giardia, a primordial secretory apparatus is in operation by which proteins are sorted in the early secretory pathway, and the developmentally induced ESVs carry out at least some Golgi functions.