Boxwood blight: an ongoing threat to ornamental and native boxwood

Applied Microbiology and Biotechnology - Boxwood blight is an emerging disease of ornamental and native boxwood plants in the family Buxaceae. First documented in the 1990s at a single location in...     

Contributions of the two accessory subunits,RNASEH2B and RNASEH2C,to the activity and properties of the human RNase H2 complex

Eukaryotic RNase H2 is a heterotrimeric enzyme. Here, we show that the biochemical composition and stoichiometry of the human RNase H2 complex is consistent with the properties previously deduced from genetic studies. The catalytic subunit of eukaryotic RNase H2, RNASEH2A, is well conserved and similar to the monomeric prokaryotic RNase HII. In contrast, the RNASEH2B and RNASEH2C subunits from human and Saccharomyces cerevisiae share very little homology, although they both form soluble B/C complexes that may serve as a nucleation site for the addition of RNASEH2A to form an active RNase H2, or for interactions with other proteins to support different functions. The RNASEH2B subunit has a PIP-box and confers PCNA binding to human RNase H2. Unlike Escherichia coli RNase HII, eukaryotic RNase H2 acts processively and hydrolyzes a variety of RNA/DNA hybrids with similar efficiencies, suggesting multiple cellular substrates. Moreover, of five analyzed mutations in human RNASEH2B and RNASEH2C linked to Aicardi-Goutières Syndrome (AGS), only one, R69W in the RNASEH2C protein, exhibits a significant reduction in specific activity, revealing a role for the C subunit in enzymatic activity. Near-normal activity of four AGS-related mutant enzymes was unexpected in light of their predicted impairment causing the AGS phenotype.     

Surfactant protein D decreases pollen-induced IgE-dependent mast cell degranulation

Mast cells play a key role in allergy and asthma. They reside at the host-environment interface and are among the first cells to make contact with inhaled microorganisms and particulate antigens. Pulmonary surfactant proteins A and D (SP-A and SP-D) function in lung host defense by enhancing microbe phagocytosis and mediating other immune cell functions, but little is known about their effects on mast cells. We hypothesized that SP-A and/or SP-D modulate IgE-dependent mast cell functions. Pollen starch granules (PSG) extracted from Dactylis glomerata and coated with trinitrophenol (TNP) were used as a model of an inhaled organic particulate allergen. Our data revealed that SP-D inhibited by 50% the release of beta-hexosaminidase by peritoneal mast cells sensitized with IgE anti-TNP and stimulated with TNP-PSG. In contrast, SP-A had no effect. Furthermore, SP-D aggregated PSG in a dose-dependent manner, and this aggregation was mediated by SP-D's carbohydrate recognition domain. A single arm SP-D mutant (RrSP-Dser15,20) neither aggregated PSG nor inhibited degranulation, suggesting that multimerization of SP-D is required for maximal PSG aggregation and inhibition of PSG-induced mast cell degranulation. This study is the first to demonstrate that SP-D modulates IgE-mediated mast cell functions, which are important in asthma and allergic inflammation.     

Differential modulation of Alzheimer's disease amyloid beta-peptide accumulation by diverse classes of metal ligands

Biometals have an important role in AD (Alzheimer's disease) and metal ligands have been investigated as potential therapeutic agents for treatment of AD. In recent studies the 8HQ (8-hydroxyquinoline) derivative CQ (clioquinol) has shown promising results in animal models and small clinical trials; however, the actual mode of action in vivo is still being investigated. We previously reported that CQ-metal complexes up-regulated MMP (matrix metalloprotease) activity in vitro by activating PI3K (phosphoinositide 3-kinase) and JNK (c-jun N-terminal kinase), and that the increased MMP activity resulted in enhanced degradation of secreted Abeta (amyloid beta) peptide. In the present study, we have further investigated the biochemical mechanisms by which metal ligands affect Abeta metabolism. To achieve this, we measured the effects of diverse metal ligands on cellular metal uptake and secreted Abeta levels in cell culture. We report that different classes of metal ligands including 8HQ and phenanthroline derivatives and the sulfur compound PDTC (pyrrolidine dithiocarbamate) elevated cellular metal levels (copper and zinc), and resulted in substantial loss of secreted Abeta. Generally, the ability to inhibit Abeta levels correlated with a higher lipid solubility of the ligands and their capacity to increase metal uptake. However, we also identified several ligands that potently inhibited Abeta levels while only inducing minimal change to cellular metal levels. Metal ligands that inhibited Abeta levels [e.g. CQ, 8HQ, NC (neocuproine), 1,10-phenanthroline and PDTC] induced metal-dependent activation of PI3K and JNK, resulting in JNK-mediated up-regulation of metalloprotease activity and subsequent loss of secreted Abeta. The findings in the present study show that diverse metal ligands with high lipid solubility can elevate cellular metal levels resulting in metalloprotease-dependent inhibition of Abeta. Given that a structurally diverse array of ligands was assessed, the results are consistent with the effects being due to metal transport rather than the chelating ligand interacting directly with a receptor.     

Mechanisms of A beta mediated neurodegeneration in Alzheimer's disease

Development of a comprehensive therapeutic treatment for the neurodegenerative Alzheimer's disease (AD) is limited by our understanding of the underlying biochemical mechanisms that drive neuronal failure. Numerous dysfunctional mechanisms have been described in AD, ranging from protein aggregation and oxidative stress to biometal dyshomeostasis and mitochondrial failure. In this review we discuss the critical role of amyloid-beta (A beta) in some of these potential mechanisms of neurodegeneration. The 39-43 amino acid A beta peptide has attracted intense research focus since it was identified as a major constituent of the amyloid deposits that characterise the AD brain, and it is now widely recognised as central to the development of AD. Familial forms of AD involve mutations that lead directly to altered A beta production from the amyloid-beta A4 precursor protein, and the degree of AD severity correlates with specific pools of A beta within the brain. A beta contributes directly to oxidative stress, mitochondrial dysfunction, impaired synaptic transmission, the disruption of membrane integrity, and impaired axonal transport. Further study of the mechanisms of A beta mediated neurodegeneration will considerably improve our understanding of AD, and may provide fundamental insights needed for the development of more effective therapeutic strategies.     

Membrane-targeted strategies for modulating APP and Aβ-mediated toxicity

Alzheimer's disease (AD) is a progressive neurodegenerative disease characterized by numerous pathological features including the accumulation of neurotoxic amyloid-β (Aβ) peptide. There is currently no effective therapy for AD, but the development of therapeutic strategies that target the cell membrane is gaining increased interest. The amyloid precursor protein (APP) from which Aβ is formed is a membrane-bound protein, and Aβ production and toxicity are both membrane mediated events. This review describes the critical role of cell membranes in AD with particular emphasis on how the composition and structure of the membrane and its specialized regions may influence toxic or benign Aβ/APP pathways in AD. The putative role of copper (Cu) in AD is also discussed, and we highlight how targeting the cell membrane with Cu complexes has therapeutic potential in AD.     

Contributions of phenylalanine 335 to ligand recognition by human surfactant protein D: ring interactions with SP-D ligands

Surfactant protein D (SP-D) is an innate immune effector that contributes to antimicrobial host defense and immune regulation. Interactions of SP-D with microorganisms and organic antigens involve binding of glycoconjugates to the C-type lectin carbohydrate recognition domain (CRD). A trimeric fusion protein encoding the human neck+CRD bound to the aromatic glycoside p-nitrophenyl-alpha-D-maltoside with nearly a log-fold higher affinity than maltose, the prototypical competitor. Maltotriose, which has the same linkage pattern as the maltoside, bound with intermediate affinity. Site-directed substitution of leucine for phenylalanine 335 (Phe-335) decreased affinities for the maltoside and maltotriose without significantly altering the affinity for maltose or glucose, and substitution of tyrosine or tryptophan for leucine restored preferential binding to maltotriose and the maltoside. A mutant with alanine at this position failed to bind to mannan or maltose-substituted solid supports. Crystallographic analysis of the human neck+CRD complexed with maltotriose or p-nitrophenyl-maltoside showed stacking of the terminal glucose or nitrophenyl ring with the aromatic ring of Phe-335. Our studies indicate that Phe-335, which is evolutionarily conserved in all known SP-Ds, plays important, if not critical, roles in SP-D function.     

Informing and influencing the interface between biodiversity science and biodiversity policy in South Africa

South Africa, as a megadiverse country (±21 700 vascular plants, 4800 vertebrates and 68 900 invertebrates described), is presently engaged with an extended, modified Global Strategy for Plant Conservation (GSPC). The country is fortunate in having a strong tradition of systematics research and, inter alia, houses several million preserved plant specimens (±1 million databased and georeferenced), allowing taxonomists and conservationists to track both the occurrence and distribution of indigenous and naturalized plant species. These rich local resources have been extensively drawn upon to deliver, with varying degrees of success, the 16 outcome-oriented GSPC 2010 Targets. The National Environmental Management: Biodiversity Act (NEMBA, 2004), the National Biodiversity Strategy and Action Plan (NBSAP) and the National Biodiversity Framework (NBF) have provided a robust legislative, enabling and policy framework for making operational and advancing GSPC-related efforts. However, within an emerging economy, the conservation of biodiversity has competed for government resources with housing, sanitation, primary education, basic health care and crime prevention, delivery of which translates to the currency of politicians: votes. A key challenge identified by local (and global) biodiversity scientists for the current GSPC phase is broad-scale advocacy, communicating the changing state of nature, and the inter-relatedness of biodiversity and human well-being. The nature of meeting this challenge is explored.