Effects of divalent metal ions on the alphaB-crystallin chaperone-like activity: spectroscopic evidence for a complex between copper(II) and protein

AlphaB-crystallin is a small heat shock protein, showing chaperone-like activity, that is expressed in the lens and in several other tissues. The role of some metal ions in the alphaB-crystallin biology starts to be well documented. In some neuro-degenerative pathologies, like Parkinson and Alzheimer's diseases, alphaB-crystallin is expressed at high levels. In the same pathologies an accumulation of divalent metal cations is observed. In order to investigate the interactions between human alphaB-crystallin and divalent metal ions, the effect of copper, zinc and calcium on the chaperone-like activity of the protein has been studied. Copper and zinc at concentrations 0.1 and 1 mM significantly increase the chaperone-like activity, whereas calcium 1 mM completely inhibits activity. Electron paramagnetic resonance (EPR) and circular dichroism (CD) spectra indicate the possible complex formation between Cu(II) and protein at physiological pH. Molecular modeling calculations, carried out for the probable Cu(II) binding site, suggest that a complex with three histidine residues is possible.     

Longli is not a Hybrid of Longan and Lychee as Revealed by Genome Size Analysis and Trichome Morphology

Lychee, longan, longli, and rambutan are closely related, commercially important fruit trees in the Sapindaceae family. Longli fruits are morphologically similar to both lychee and longan, displaying a yellow-brown pericarp like longan, and small, sharp protuberances like lychee. These similarities have led to the hypothesis that longli is the result of intergeneric hybridization between longan and lychee. Scanning electron microscopy and flow cytometry were used to examine trichome morphology and genome size, respectively, to test this hypothesis. Longan, lychee, longli (D. confinis), and ‘Malesianus’ (D. longan sub spp malesianus) had morphologically distinct trichomes. The genome sizes for lychee (554 Mb), longan (444 Mb), ‘Malesianus’ (404 Mb), and rambutan (339 Mb) are distinctive and in a narrow range. ‘Malesianus’ has a genome 9% smaller than that of longan and 27% smaller than that of lychee. It is likely a species that evolved independently in northern Borneo island, and could be classified as a species, Dimocarpus malesianus, not a sub-species of longan as presently stated. Flow cytometry revealed a 50% variation in genome sizes among longli varieties, with genome sizes ranging from 450 to 678 Mb, beyond the range between longan and lychee. The genome size variation and distinct leaf hair morphology suggest that longli is not an intergeneric hybrid, and it is likely a separate genus evolved independently. The tested cultivars with distinctive genome sizes within D. confinis could be classified as separate species.     

Systems analysis of primary Sjögren's syndrome pathogenesis in salivary glands identifies shared pathways in human and a mouse model

Bone tissue has an exceptional quality to regenerate to native tissue in response to injury. However, the fracture repair process requires mechanical stability or a viable biological microenvironment or both to ensure successful healing to native tissue. An improved understanding of the molecular and cellular events that occur during bone repair and remodeling has led to the development of biologic agents that can augment the biological microenvironment and enhance bone repair. Orthobiologics, including stem cells, osteoinductive growth factors, osteoconductive matrices, and anabolic agents, are available clinically for accelerating fracture repair and treatment of compromised bone repair situations like delayed unions and nonunions. Preclinical and clinical studies using biologic agents like recombinant bone morphogenetic proteins have demonstrated an efficacy similar or better than that of autologous bone graft in acute fracture healing. A lack of standardized outcome measures for comparison of biologic agents in clinical fracture repair trials, frequent off-label use, and a limited understanding of the biological activity of these agents at the bone repair site have limited their efficacy in clinical applications.     

Health and chemical burdens of fish species from polluted and hyper-eutrophic freshwater ecosystems in South Africa

Three aquatic ecosystems in South Africa, the Hartbeespoort, Klipvoor and Bospoort Dams, are classified as hyper-eutrophic, because of high nutrient loads and chemical pollution. Water and two fish species, Clarias gariepinus and Cyprinus carpio, were collected from these dams to assess the impact of eutrophication and chemical pollutants on their health status. Water and muscle samples were analysed for organic and inorganic chemicals. Condition factor was determined and a necropsy performed to note any macroscopic abnormalities. A histology-based fish health assessment was done on the liver, kidney, gills and gonads. A number of fish from the three dams exhibited livers with fatty change and focal discoloration, skin lesions and parasites within the visceral cavity. The prevalence and severity of histopathology in the liver resulted in higher liver index values than the index values for kidneys and gills. Aluminium, silicon and chromium were detected in the water and muscle tissue. The DDT metabolite p,p’-DDE was present in both species, as well as in fish from the reference site, Marico-Bosveld Dam. Only C. gariepinus from Hartbeespoort Dam had p,p’-DDD levels higher than 5 µg g^?1 per edible portion. Water from hyper-eutrophic dams adversely affects the health of freshwater fish.     

Inactivation of template-directed misfolding of infectious prion protein by ozone

Misfolded prions (PrP(Sc)) are well known for their resistance to conventional decontamination processes. The potential risk of contamination of the water environment, as a result of disposal of specified risk materials (SRM), has raised public concerns. Ozone is commonly utilized in the water industry for inactivation of microbial contaminants and was tested in this study for its ability to inactivate prions (263K hamster scrapie = PrP(Sc)). Treatment variables included initial ozone dose (7.6 to 25.7 mg/liter), contact time (5 s and 5 min), temperature (4°C and 20°C), and pH (pH 4.4, 6.0, and 8.0). Exposure of dilute suspensions of the infected 263K hamster brain homogenates (IBH) (0.01%) to ozone resulted in the in vitro destruction of the templating properties of PrP(Sc), as measured by the protein misfolding cyclic amplification (PMCA) assay. The highest levels of prion inactivation (≥4 log(10)) were observed with ozone doses of 13.0 mg/liter, at pH 4.4 and 20°C, resulting in a CT (the product of residual ozone concentration and contact time) value as low as 0.59 mg · liter(-1) min. A comparison of ozone CT requirements among various pathogens suggests that prions are more susceptible to ozone degradation than some model bacteria and protozoa and that ozone treatment may be an effective solution for inactivating prions in water and wastewater.     

EphB receptors trigger Akt activation and suppress Fas receptor-induced apoptosis in malignant T lymphocytes

Treatment of hematopoietic malignancies often requires allogeneic bone marrow transplantation, and the subsequent graft-versus-leukemia response is crucial for the elimination of malignant cells. Cytotoxic T lymphocytes and NK cells responsible for the immunoelimination express Fas ligand and strongly rely on the induction of Fas receptor-mediated apoptosis for their action. Although cancer cells are removed successfully by graft-versus-leukemia reactions in myeloid malignancies, their efficiency is low in T cell leukemias. This may be partially because of the ability of malignant T cells to escape apoptosis. Our work shows that Eph family receptor EphB3 is consistently expressed by malignant T lymphocytes, most frequently in combination with EphB6, and that stimulation with their common ligands, ephrin-B1 and ephrin-B2, strongly suppresses Fas-induced apoptosis in these cells. This effect is associated with Akt activation and with the inhibition of the Fas receptor-initiated caspase proteolytic cascade. Akt proved to be crucial for the prosurvival response, because inhibition of Akt, but not of other molecules central to T cell biology, including Src kinases, MEK1 and MEK2, blocked the antiapoptotic effect. Overall, this demonstrates a new role for EphB receptors in the protection of malignant T cells from Fas-induced apoptosis through Akt engagement and prevention of caspase activation. Because Fas-triggered apoptosis is actively involved in the graft-versus-leukemia response and cytotoxic T cells express ephrin-Bs, our observations suggest that EphB receptors are likely to support immunoevasivenes of T cell malignancies and may represent promising targets for therapies, aiming to enhance immunoelimination of cancerous T cells.     

Can plants serve as a vector for prions causing chronic wasting disease?

Prions, the causative agent of chronic wasting disease (CWD) enter the environment through shedding of bodily fluids and carcass decay, posing a disease risk as a result of their environmental persistence. Plants have the ability to take up large organic particles, including whole proteins, and microbes. This study used wheat (Triticum aestivum L.) to investigate the uptake of infectious CWD prions into roots and their transport into aerial tissues. The roots of intact wheat plants were exposed to infectious prions (PrP^TSE) for 24 h in three replicate studies with PrP^TSE in protein extracts being detected by western blot, IDEXX and Bio-Rad diagnostic tests. Recombinant prion protein (PrP^C) bound to roots, but was not detected in the stem or leaves. Protease-digested CWD prions (PrP^TSE) in elk brain homogenate interacted with root tissue, but were not detected in the stem. This suggests wheat was unable to transport sufficient PrP^TSE from the roots to the stem to be detectable by the methods employed. Undigested PrP^TSE did not associate with roots. The present study suggests that if prions are transported from the roots to the stems it is at levels that are below those that are detectable by western blot, IDEXX or Bio-Rad diagnostic kits.     

Isolation and1H-NMR spectroscopic identification of the glucose-containing lipid-linked precursor oligosaccharide ofN-linked carbohydrate chains

The lipid-linked precursor ofN-type glycoprotein oligosaccharides was isolated from porcine thyroid microsomes after in cubation with UDP[³H] Glucose. The carbohydrate was released from dolichol pyrophosphate by mild acid hydrolysis, purified by gel filtration and characterized by 500-MHz¹H-NMR spectroscopy in combination with enzymatic degradation. The parent oligosaccharide was found to be Glc₃Man₉Glc-NAc₂. The three glucose residues are present in the linear sequence Glcα1-2Glα1-3 Glc, the latter being α(1-3)-linked to one of the mannose residues. In order to establish the branch location of the triglucosyl unit, the parent compound was digested with jack-bean α-mannosidase. The oligosaccharide product was purified by gel filtration, and identified by¹H-NMR as Glc₃Man₅GlcNAc₂ lacking the mannose residues A, D₂, B and D₃. Therefore, the structure of the precursor oligosaccharide is as follows: $$\begin{gathered} c b a D_1 C 4 \hfill \\ Glc\alpha 1 - 2Glc\alpha 1 - 3Glc\alpha 1 - 3Man\alpha 1 - 2Man\alpha 1 - 2Man\alpha 1 \hfill \\ 3 \swarrow 3 2 1 \hfill \\ Man\alpha 1 - 2Man\alpha 1 Man\beta 1 - 4GlcNAc\beta 1 - 4GlcNAc \hfill \\ D_{2 } A 3 6 \hfill \\ Man\alpha 1 \hfill \\ 6 \hfill \\ Man\alpha 1 - 2Man\alpha 1 \nwarrow 4 \hfill \\ D_3 B \hfill \\ \end{gathered} $$