Trypanosoma cruzi: incorporation of [3H]-palmitic acid and [3H]-galactose into components shed by trypomastigotes.

Trypomastigotes were metabolically labeled with [3H]-palmitic acid or [3H]-galactose and labeled components were detected in the culture medium. Thin layer chromatography of the shed material showed several lipids in the [3H]-palmitic acid labeled sample while the sugar was mainly incorporated into macromolecules. The material incorporated with the lipidic precursor was fractionated by DEAE-Sephadex (acetate form) and the amount of radioactivity was ten times higher in the acidic lipids than in the neutral lipids. When acidic lipids were further separated by Unisil, 73% of the radioactivity was recovered in the less polar fraction. Different patterns were obtained on comparison of the shed components with the lipids remaining in the parasite.     

BioGraph: unsupervised biomedical knowledge discovery via automated hypothesis generation

We present BioGraph, a data integration and data mining platform for the exploration and discovery of biomedical information. The platform offers prioritizations of putative disease genes, supported by functional hypotheses. We show that BioGraph can retrospectively confirm recently discovered disease genes and identify potential susceptibility genes, outperforming existing technologies, without requiring prior domain knowledge. Additionally, BioGraph allows for generic biomedical applications beyond gene discovery. BioGraph is accessible at .     

The role of Arabidopsis SCAR genes in ARP2-ARP3-dependent cell morphogenesis

The actin-nucleating ARP2-ARP3 complex controls cell shape in plants in many different cell types. Its activity is controlled by a multimeric complex containing BRK1 (also known as HSPC300), NAP1, SRA1, ABI and SCAR/WAVE. In this study, we focus on the function of the five putative SCAR homologues in Arabidopsis and we provide biochemical evidence that AtSCAR2 can activate the ARP2-ARP3 complex in vitro. Among the single mutants, mutations in only AtSCAR2 result in a subtle or weak phenotype similar to ARP2, ARP3 and other ;distorted' mutants. Double-mutant analysis revealed a redundancy with AtSCAR4. Systematic application of the yeast two-hybrid system and Bimolecular Fluorescence Complementation (BiFC) revealed a complex protein-interaction network between the ARP2-ARP3 complex and its genetically defined regulators. In addition to protein interactions known in other systems, we identified several new interactions, suggesting that SPIKE1 may be an integral component of the SCAR/WAVE complex and that SCAR proteins in plants might act as direct effectors of ROP GTPases.     

Arabidopsis N-MYC DOWNREGULATED-LIKE1, a Positive Regulator of Auxin Transport in a G Protein–Mediated Pathway

Root architecture results from coordinated cell division and expansion in spatially distinct cells of the root and is established and maintained by gradients of auxin and nutrients such as sugars. Auxin is transported acropetally through the root within the central stele and then, upon reaching the root apex, auxin is transported basipetally through the outer cortical and epidermal cells. The two Gβγ dimers of the Arabidopsis thaliana heterotrimeric G protein complex are differentially localized to the central and cortical tissues of the Arabidopsis roots. A null mutation in either the single β (AGB1) or the two γ (AGG1 and AGG2) subunits confers phenotypes that disrupt the proper architecture of Arabidopsis roots and are consistent with altered auxin transport. Here, we describe an evolutionarily conserved interaction between AGB1/AGG dimers and a protein designated N-MYC DOWNREGULATED-LIKE1 (NDL1). The Arabidopsis genome encodes two homologs of NDL1 (NDL2 and NDL3), which also interact with AGB1/AGG1 and AGB1/AGG2 dimers. We show that NDL proteins act in a signaling pathway that modulates root auxin transport and auxin gradients in part by affecting the levels of at least two auxin transport facilitators. Reduction of NDL family gene expression and overexpression of NDL1 alter root architecture, auxin transport, and auxin maxima. AGB1, auxin, and sugars are required for NDL1 protein stability in regions of the root where auxin gradients are established; thus, the signaling mechanism contains feedback loops.     

Genetic selection and liquid medium conditions improve the yield of androgenetic plants from diploid potatoes

Summary Solatium tuberosum L. diploid strains with superior androgenetic capacity have been selected for from androgenetic progenies of unselected diploid material. The paper also demonstrates that the use of a liquid medium for culturing potato anthers, instead of the conventional solid agar plates, improves the yield of androgenetic embryoids. The new method, associated with two successive cycles of selection for superior androgenetic response, allows the induction and regeneration of microspore derived plants on a large scale. The best genotype (clone 21 in this paper) regenerates androgenetic plants with a frequency around 30 per each anther plated. Over 80% of the regenerated plants are diploid. It is suggested that the androgenetic embryoids mainly originate from unreduced microspores by a mechanism which maintains a heterozygous or a partly heterozygous genetic situation.     

Transposition of the maize transposable element Ac in Solanum tuberosum

Summary The maize transposable element Ac has been introduced into potato via the T-DNA (transferred DNA) of Agrobacterium tumefaciens. Ac was inserted within the untranslated leader region of a neomycin phosphotransferase II (NPT-II) gene such that excision restored NPT-II activity. Two approaches to monitor Ac excision were used. (i) Using an Agrobacterium strain harbouring plasmid pGV3850::pKU3, leaf discs were selected on kanamycin (Km) after exposure to Agrobacterium. (ii) Using a strain containing plasmid pGV3850HPT::pKU3, the leaf discs were selected on hygromycin (Hm) and the resulting shoots were checked for NPT-II expression. Thirteen kanamycin resistant shoots transformed with pGV3850::pKU3 were isolated, suggesting that Ac had excised from the NPT-II gene. Out of 43 hygromycin resistant shoots transformed with pGV3850HPT::pKU3, 22 expressed the NPT-II gene, indicating that Ac had undergone excision in approximately 50% of the hygromycin resistant shoots. Southern analysis revealed that all kanamycin resistant plants contained the DNA restriction fragments expected when Ac excises from the NPT-II gene. The presence of Ac at new locations within the genomic DNA of several transformants was also detected.     

Anther culture of Solanum genotypes

Anther culture response was examined in five Solanum genotypes. Large genotypic differences exist in response to culture and liquid culture media were found to be superior to agar solidified media systems. Pre-treatment effects on embryoid induction were also investigated and two of the genotypes displayed differential response to temperature stress pretreatment. In general the beneficial effect of charcoal in the media was confirmed. Successful embryoid production and plantlet regeneration was obtained from the wild potato species, Solanum papita. The influence of sucrose concentration on embryoid production was also investigated. Large genotype by sucrose interactions were detected and this was mainly due to the differential response of the tuberosum clones to increasing sucrose concentration when compared with S. papita. Embryoids were produced from this species in media containing 15% sucrose although the optimum concentration of sucrose for embryoid production was 9%. The possible role of anther culture techniques in gene introgression and potato improvement is discussed.     

Cytochemical demonstration of hydrogen peroxide in polymorphonuclear leukocyte phagosomes

Phagocytosis by polymorphonuclear leukocytes (PMN) is accompanied by specific morphological and metabolic events which may result in the killing of internalized micro-organism. Hydrogen peroxide is produced in increased amounts during phagocytosis (17) and in combination with myeloperoxidase and halide ions constitute a potent, microbicidal mechanism (8,9,11). There can be direct iodination of micro-organisms (10), or alternatively, other intermediate reaction products, i.e. chloramines and aldehydes (21), can exert a microbicidal effect. The H2O2-peroxidase-halide system is presumed to operate within the phagocytic vacuole (12,18). Myeloperoxidase, present in the primary granules of PMN, enters the phagocytic vacuole during degranulation (1,4,7), and halide ions are probably derived from the extracellular medium or are present in the PMN (see 11, 18). For the operation of this system in intact cells, the presence of H2O2 in the phagocytic vacuole is necessary, and indeed this has been suggested by the work of several investigators (12, 18, 21). In the present investigation, the diaminobenzidine reaction of Graham and Karnovsky (5), modified to utilize endogenous myeloperoxidase and hydrogen peroxide, has been applied to actively phagocytizing PMN to demonstrate cytochemically the presence of H2O2 in the phagocytic vacuole.     

Breeding for nematode and virus resistance in potato via anther culture

Summary In Solanum tuberosum the production by parthenogenesis of 2x plants with 24 chromosomes, and the regeneration of microspores of such dihaploids to yield monohaploid (1x) plants is reproducibly possible, at least for some specific genotypes. Experiments are described using tissue culture techniques in an applied breeding program with the main aim of increasing the level of resistance to the potato cyst nematode Globodera pallida (Stone) and to the potato viruses X, Y and leaf roll. These resistances follow quantitative as well as qualitative modes of inheritance. Using anther culture it is demonstrated that doubled monohaploid clones can be produced which possess the resistance in the homozygous condition. In both ways of inheritance the ratio of resistant clones is rather high. The genotype of the anther donor plant has, however, a strong influence on the total number of androgenetic plants which can be regenerated. Therefore, experiments were initiated with the aim of integrating this capacity for regeneration (tissue culture ability) into valuable genotypes. The results show that the potentiality for regeneration is under genetic control and can be utilized by combination breeding. Its inheritance and physiological basis, as well as the behaviour of complete homozygous clones, is discussed.Dedicated to Prof. Dr. Joseph Straub on the occasion of his 70. birthday