I. The Disastereoselective Alkylation of Chiral 2-Oxazolidinone Imide Enolates. II. Efforts Directed Toward the Enantioselective Total Synthesis of Ferensimycin B

The diastereoselective alkylation of chiral 2-oxazolidinone imide enolates is described. The requisite chiral N-acyl 2-oxazolididones are prepared from readily available amino acid and amino alcohol precursors. The lithium and sodium enolates, derived from these chiral imides, react in a highly diastereoselective manner with a variety of electrophiles. Furthermore, the diasteromers are often separable by liquid chromateography affording products with a diastereomeric purity ≥ 99:1. Several methods are described for the non–destructive removal of the chiral auxiliary to afford enantiomerically pure alcohols, aldehydes, carboxylic acids, acid chlorides, esters, hydrazides, and ketones. Through the use of chiral imides 16 and 20 either of the enantiomeric products can be obtained. [Structural formula. See abstract in scanned thesis for details.] An approach to the enantioselective total synthesis of the poly- ether ionophore antibiotic ferensimycin B (2) is described. The synthesis employs the diastereoselective alkylation and aldol condensation of chiral 2-oxazolidinone imide enolates to both construct the carbon backbone and generate the necessary stereo- centers. This research has culminated in the preparation of the advanced intermediate 49. [Structural formula. See abstract in scanned thesis for details.] An enantioselective total synthesis of (R) and (S)-thiorphan [N-(1-oxo-2-mercaptomethyl-3-phenylpropyl)glycine] via a six-step sequence is reported. The key step, establishing the absolute stereochemistry, is the diastereoselective alkylation of the enolate derived from chiral 2-oxazolidinone imide 16 (R = PhCH2) or 20 (R = PhCH2) with benzyl bromomethyl sulfide. The level of alkylation diastereoselection is in excess of 95:5.</p

RDGB , a PtdIns-PtdOH transfer protein, regulates G-protein-coupled PtdIns(4,5)P2 signalling during Drosophila phototransduction

Many membrane receptors activate phospholipase C (PLC) during signalling, triggering changes in the levels of several plasma membrane lipids including phosphatidylinositol (PtdIns), phosphatidic acid (PtdOH) and phosphatidylinositol 4,5-bisphosphate [PtdIns(4,5)P2]. It is widely believed that exchange of lipids between the plasma membrane and endoplasmic reticulum (ER) is required to restore lipid homeostasis during PLC signalling, yet the mechanism remains unresolved. RDGBα (hereafter RDGB) is a multi-domain protein with a PtdIns transfer protein (PITP) domain (RDGB-PITPd). We find that, in vitro, the RDGB-PITPd binds and transfers both PtdOH and PtdIns. In Drosophila photoreceptors, which experience high rates of PLC activity, RDGB function is essential for phototransduction. We show that binding of PtdIns to RDGB-PITPd is essential for normal phototransduction; however, this property is insufficient to explain the in vivo function because another Drosophila PITP (encoded by vib) that also binds PtdIns cannot rescue the phenotypes of RDGB deletion. In RDGB mutants, PtdIns(4,5)P2 resynthesis at the plasma membrane following PLC activation is delayed and PtdOH levels elevate. Thus RDGB couples the turnover of both PtdIns and PtdOH, key lipid intermediates during G-protein-coupled PtdIns(4,5)P2 turnover

Mating type idiomorphs of Pyrenophora teres in Turkey

Pyrenophora teres f. maculata (Ptm) and Pyrenophora teres f. teres (Ptt) causes spot form and net form of net blotch diseases of barley, respectively. Although both forms of P. teres are morphologically similar, their symptoms and genetic background differ. In this study, 175 single spore (109 Ptm and 66 Ptt) isolates obtained from different regions of Turkey were evaluated for their mating type distribution and prevalence. Fungal isolates of both forms were verified using species-speci.c polymerase chain reaction (PCR) primers. For mating type determination studies, duplex PCR was performed using MAT-specific single nucleotide polymorphism primers. Sixty and 49 of 109 Ptm isolates were found as MAT1-1 and MAT1-2 types, respectively and 43 and 23 of 66 Ptt isolates were found as MAT1-1 and MAT1-2 types, respectively. These results show the possibility of sexual reproduction among the Ptm isolates in Turkey and Ptt population of Central Anatolia, Turkey. However, the overall pattern of Ptt isolates did not support the sexual reproduction hypothesis in Turkey. Sexual reproduction in the life cycle of P. teres is important since it could lead to genetic and pathogenic variation. As a result of new sexual combinations more virulent pathotypes of P. teres may occur

Morphological and molecular characterization of Curvularia and related species associated with leaf spot disease of rice in Peninsular Malaysia

Curvularia species are important phytopathogens reported worldwide. They are closely related; consist of major destructive pathogens mainly for grasses and cereal plants including rice (Oryza sativa). A leaf spot symptom of rice is one of the common symptoms found in the rice field and caused reduction of rice yield. However, there are no reports on Curvularia species associated with rice leaves showing spot symptoms. The objectives are to isolate and characterize Curvularia and related species from leaf spot of rice by using morphological and molecular characterization and to determine the phylogenetic relationship between the isolated fungi. Fungal isolation was done from diseased rice leaves showing leaf spot symptoms collected throughout Peninsular Malaysia. Thirty-three isolates were recovered and identified based on their morphological characteristics such as conidia morphology, colony appearance, pigmentation and growth rate for species delimitation. Internal transcribed spacer (ITS) region was amplified to confirm the species identification. The 33 isolates were identified as Bipolaris sorokiniana (10 isolates), Curvularia hawaiiensis (8 isolates), C. geniculata (6 isolates), C. eragrostidis (6 isolates), C. aeria (2 isolates) and C. lunata (1 isolate). A phylogenetic tree was constructed based on ITS sequences using neighbour-joining method. The tree grouped members of Curvularia and Bipolaris into different clades. The phylogenetic tree indicated that the presence of two groups of fungi species; highly virulent and mild pathogens. In conclusion, Curvularia species and Bipolaris sorokiniana were present in rice field in Malaysia and associated with leaf spot of rice

Pyrenophora teres: Profile of an increasingly damaging barley pathogen

Pyrenophora teres, causal agent of net blotch of barley, exists in two forms, designated P. teres f. teres and P. teres f. maculata, which induce net form net blotch (NFNB) and spot form net blotch (SFNB), respectively. Significantly more work has been performed on the net form than on the spot form although recent activity in spot form research has increased because of epidemics of SFNB in barley‐producing regions. Genetic studies have demonstrated that NFNB resistance in barley is present in both dominant and recessive forms, and that resistance/susceptibility to both forms can be conferred by major genes, although minor quantitative trait loci have also been identified. Early work on the virulence of the pathogen showed toxin effector production to be important in disease induction by both forms of pathogen. Since then, several laboratories have investigated effectors of virulence and avirulence, and both forms are complex in their interaction with the host. Here, we assemble recent information from the literature that describes both forms of this important pathogen and includes reports describing the host–pathogen interaction with barley. We also include preliminary findings from a genome sequence survey

A first genome assembly of the barley fungal pathogen Pyrenophora teres f. teres

Background: Pyrenophora teres f. teres is a necrotrophic fungal pathogen and the cause of one of barley’s most important diseases, net form of net blotch. Here we report the first genome assembly for this species based solely on short Solexa sequencing reads of isolate 0-1. The assembly was validated by comparison to BAC sequences, ESTs, orthologous genes and by PCR, and complemented by cytogenetic karyotyping and the first genome-wide genetic map for P. teres f. teres. Results: The total assembly was 41.95 Mbp and contains 11,799 gene models of 50 amino acids or more. Comparison against two sequenced BACs showed that complex regions with a high GC content assembled effectively. Electrophoretic karyotyping showed distinct chromosomal polymorphisms between isolates 0-1 and 15A, and cytological karyotyping confirmed the presence of at least nine chromosomes. The genetic map spans 2477.7 cM and is composed of 243 markers in 25 linkage groups, and incorporates SSR markers developed from the assembly. Among predicted genes, non-ribosomal peptide synthetases and efflux pumps in particular appear to have undergone a P. teres f. teres-specific expansion of non-orthologous gene families. Conclusions: This study demonstrates that paired-end Solexa sequencing can successfully capture coding regions of a filamentous fungal genome. The assembly contains a plethora of predicted genes that have been implicated in a necrotrophic lifestyle and pathogenicity and presents a significant resource for examining the bases for P. teres f. teres pathogenicity