Invasive forms of apicomplexan parasites contain secretory organelles called rhoptries that are essential for entry into host cells. We present a detailed characterization of an unusual rhoptry protein of the human malaria parasite Plasmodium falciparum, the rhoptryassociated membrane antigen (RAMA) that appears to have roles in both rhoptry biogenesis and host cell invasion. RAMA is synthesized as a 170-kDa protein in early trophozoites, several hours before rhoptry formation and is transiently localized within the endoplasmic reticulum and Golgi within lipid-rich microdomains. Regions of the Golgi membrane containing RAMA bud to form vesicles that later mature into rhoptries in a process that is inhibitable by brefeldin A. Other rhoptry proteins such as RhopH3 and RAP1 are found in close apposition with RAMA suggesting direct protein-protein interactions. We suggest that RAMA is involved in trafficking of these proteins into rhoptries. In rhoptries, RAMA is proteolytically processed to give a 60-kDa form that is anchored in the inner face of the rhoptry membrane by means of the glycosylphosphatidylinositol anchor. The p60 RAMA form is discharged from the rhoptries of free merozoites and binds to the red blood cell membrane by its most C-terminal region. In early ring stages RAMA is found in association with the parasitophorous vacuole.Plasmodium falciparum malaria is one of the most important infectious diseases of humans, accounting for ϳ2 million deaths each year. The stages of the parasite that grow and multiply in red blood cells (RBCs) 1 cause all the pathological effects associated with the disease, and accordingly invasion of red blood cells is one of the most important steps in the parasite life cycle. Three sets of secretory organelles, the rhoptries, micronemes, and dense granules are involved in and are essential for the invasion process. The understanding of the role of these organelles provides important knowledge about the basic biology of malaria and potential therapeutical targets.Rhoptries of Plasmodium parasites are paired club-shaped organelles located at the apical end of merozoites, the form of the parasite that invades RBCs. Following the attachment of merozoites to the RBC surface, rhoptries discharge their contents onto the RBC membrane (1). Rhoptry organelles disappear after internalization of merozoites and thus are formed de novo with each erythrocytic cycle. Rhoptry formation occurs late in the erythrocytic stages of the parasite, and elucidation of rhoptry biogenesis of malaria parasites has been hindered by the lack of early organelle markers. Most of our knowledge is based on microscopic examinations, which suggest that rhoptry biogenesis follows the secretory pathway route, with rhoptry organelles being formed by sequential fusion of post-Golgi vesicles (2, 3), although why particular vesicles are selected is unclear.Rhoptry contents include both protein and lipid components, which assemble to form membrane-like structures. Protein constituents of the rhoptry contents are stil...

Synthesis of the wound-inducible alkaloid, nicotine, in roots of the allotetraploid species Nicotiana tabacum L. is strongly influenced by the presence of two non-allelic genes, A and B. Together, these loci affect baseline transcript levels of genes dedicated to secondary metabolism (e.g. PMT and A622) as well as genes with roles in separate areas of primary metabolism (e.g. ODC, ADC, SAMS — polyamines; QPT — pyridine nucleotide cycle). Experiments comparing high alkaloid variety NC 95 (AABB genotype) and near-isogenic low alkaloid N. tabacum variety LAFC 53 (aabb genotype) indicate that together, mutations in the A and B loci diminish, but do not ablate, the propensity of roots to increase transcript levels of genes involved in alkaloid metabolism after damage to aerial tissues or direct treatment with the wound hormone, methyl jasmonate. Accordingly, roots of aabb genotype can increase their nicotine content somewhat in response to these treatments. Additionally, we show that transcript levels of genes associated with polyamine metabolism (ODC, ADC, SamDC, SAMS and SS) but not alkaloid synthesis (PMT, QPT, A622) are elevated in leaves of N. tabacum in response to wounding. Moreover, respective increases in transcript levels of each gene are similar in wounded leaves of NC 95 and LAFC 53, suggesting that these increases are not controlled by combined action of genes encoded by the A and B loci. Further detailed analysis of wounded leaves of AABB genotype indicates that although transcript levels of these genes of polyamine metabolism and associated enzyme activities for ODC, ADC and SamDC, are markedly increased in leaves in response to wounding, putrescine levels remain unaltered whilst spermidine and spermine levels are reduced to 50–60% of controls levels, when analysed up to 6 h post-wounding. These observations may indicate that any wound-induced increases in polyamine biosynthesis that do occur in leaf cells during this time frame are consumed by metabolic reactions involved in repair and / or strengthening of wounded leaf tissues.

SummaryFructans are the main storage carbohydrates of temperate grasses, sustaining regrowth immediately after defoliation, as well as contributing to the nutritive value of feed. Fructan metabolism is based on the substrate sucrose and involves fructosyltransferases (FTs) for biosynthesis and fructan exohydrolases (FEHs) for degradation. Sucrose is also utilized by invertases (INVs), which hydrolyse it into its constituent monosaccharides for use in metabolism. The isolation, molecular characterization, functional analysis, and phylogenetic relationships of genes encoding FTs, FEHs, and INVs from temperate grasses are reviewed, with an emphasis on perennial ryegrass ( Lolium perenne L.). The roles these enzymes play in fructan accumulation and remobilization, and future biotechnological applications in molecular plant breeding are discussed.

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