MicrobiologyBytes

BSE, commonly known as mad cow disease, is caused by an infectious and abnormal form of the prion protein that is present on cells within the nervous system, but it is unclear as to what causes the abnormality to occur. Prion diseases are unique in that their infectious nature is not dependent on nucleic acid but is instead attributed to a misfolded protein, the prion protein. This misfolded prion protein is capable of inducing the misfolding of the normal form of the prion protein that is present on the surface of neurons and other cells in the body. However, the site in the cell at which this misfolding occurs and whether other proteins are involved remains controversial. Researchers have addressed these questions by investigating how the normal form of the prion protein is targeted to specialised domains on the plasma membrane termed cholesterol-rich lipid rafts. They showed that targeting is due, in part, to a particular heparin sulfate proteoglycan called glypican-1.

A new treatment route for bovine spongiform encephalopathy (BSE) and its human form Creutzfeldt Jakob disease (CJD) could be a step closer. This new works shows that the presence of Glypican-1 causes the numbers of abnormal prion proteins to rise. This suggests that Glypican-1 acts as a scaffold bringing the two forms of the prion protein together and that this contact causes normal prions to mutate into the infectious form. These findings may have implications for the treatment of both BSE and the human form of the disease, CJD. Although the scientists mainly conducted experiments using cells infected with prions, it is also possible that Glypican-1 is involved in other diseases of the nervous system, although experiments have not shown any link with other neurodegenerative diseases such as Alzheimer’s disease.

Glypican-1 Mediates Both Prion Protein Lipid Raft Association and Disease Isoform Formation. 2009 PLoS Pathog 5(11): e1000666. doi:10.1371/journal.ppat.1000666
In prion diseases, the cellular form of the prion protein, PrPC, undergoes a conformational conversion to the infectious isoform, PrPSc. PrPC associates with lipid rafts through its glycosyl-phosphatidylinositol (GPI) anchor and a region in its N terminal domain which also binds to heparan sulfate proteoglycans (HSPGs). We show that heparin displaces PrPC from rafts and promotes its endocytosis, suggesting that heparin competes with an endogenous raft-resident HSPG for binding to PrPC. We then utilised a transmembrane-anchored form of PrP (PrP-TM), which is targeted to rafts solely by its N-terminal domain, to show that both heparin and phosphatidylinositol-specific phospholipase C can inhibit its association with detergent-resistant rafts, implying that a GPI-anchored HSPG targets PrPC to rafts. Depletion of the major neuronal GPI anchored HSPG, glypican-1, significantly reduced the raft association of PrP-TM and displaced PrPC from rafts, promoting its endocytosis. Glypican-1 and PrPC colocalised on the cell surface and both PrPC and PrPSc co-immunoprecipitated with glypican-1. Critically, treatment of scrapie-infected N2a cells with glypican-1 siRNA significantly reduced PrPSc formation. In contrast, depletion of glypican-1 did not alter the inhibitory effect of PrPC on the b-secretase cleavage of the Alzheimer’s amyloid precursor protein. These data indicate that glypican-1 is a novel cellular cofactor for prion conversion and we propose that it acts as a scaffold facilitating the interaction of PrPC and PrPSc in lipid rafts.

Related:

• Prions and Alzheimers disease
• Novel mutation linked to mad cow disease
• What the heck are prions for?
• Drugs for treatment of prion infections

Lipid droplets consist of a core of neutral lipids surrounded by an outer phospholipid monolayer and associated proteins. These organelles are thought to be generated when neutral lipids accumulate in the endoplasmic reticulum (ER) bilayer. Recently, lipid droplets have been shown to play a role in several cellular processes, including lipid storage, lipid trafficking, and protein storage and degradation. The importance of this organelle is underscored by the fact that lipid droplets have been linked to several metabolic diseases, most notably diabetes and obesity. Lipid droplets have also been shown to play a critical role in the replication of several pathogens. One of the most well characterized examples is hepatitis C virus (HCV).

Viperin is an interferon (IFN)-induced antiviral protein that is induced upon HCV infection and inhibits HCV replication. Like the HCV NS proteins, viperin has been shown to localize to the cytosolic face of the ER through an N-terminal amphipathic
alpha-helix. This N-terminal amphipathic
alpha-helix is essential for viperin to inhibit HCV and influenza, as mutants lacking this domain have greatly reduced antiviral activity. Although the precise mechanism by which viperin inhibits HCV is still unknown, viperin was previously shown to inhibit influenza virus budding by disrupting plasma membrane lipid raft microdomains, which are sites of influenza virion assembly and budding. Independent of viral infection, the N-terminal amphipathic
alpha-helix of viperin inhibits protein secretion and appears to induce ER membrane curvature.

Numerous questions remain about how lipid droplets are generated and used by viruses. This paper shows that the IFN-induced antiviral protein viperin, which localizes to the cytosolic face of the ER and inhibits HCV, localizes to lipid droplets. This paper shows that the N-terminal amphipathic alpha-helix of viperin that is responsible for ER localization is also necessary and sufficient to localize both viperin and the fluorescent protein dsRed to lipid droplets. Point mutations in the alpha-helix that prevent ER association also disrupt lipid droplet association, and sequential deletion mutants indicate that the same number of helical turns are necessary for ER and lipid droplet association. The N-terminal amphipathic alpha-helix of the hepatitis C viral protein NS5A can localize dsRed and viperin to lipid droplets. These findings indicate that the amphipathic alpha-helices of viperin and NS5A are lipid droplet-targeting domains and suggest that viperin inhibits HCV by localizing to lipid droplets using a domain and mechanism similar to that used by HCV itself.

The antiviral protein, viperin, localizes to lipid droplets via its N-terminal amphipathic αalpha-helix. PNAS USA November 17 2009. doi: 10.1073/pnas.0911679106

Related:

• Viperin (cig5), an IFN-inducible antiviral protein directly induced by human cytomegalovirus. 2001 PNAS USA 98(26): 15125-15130
• Viroporins
• Hepatitis C Virus: a mountain to climb
• The Interferon Antiviral Response

Arsenic (As) is a natural and ubiquitous element that presents in many environmental compartments and is released through various natural processes or by human inputs. It is recognized as carcinogenic and chronic exposure to arsenic results in a wide range of adverse health effects. Depending on the physical–chemical conditions of the environment, some arsenic compounds can be easily solubilized in water and taken up by microorganisms, resulting in high levels of bioavailability. The most notable case was observed in India and Bangladesh where over 50 million people were exposed to highly contaminated water or food.

Owing to its extreme toxicity, arsenic is ranked number one on the US Environmental Protection Agency’s (EPA) priority list of drinking water contaminants and effective from 2006 the maximum contaminant level for arsenic in drinking water was reduced by the EPA from 50 ppb to 10 ppb. According to the Natural Resources Defense Council, over 56 million Americans in the 25 reporting states consume water containing arsenic at levels presenting a potential fatal cancer risk. Several treatment technologies have been applied in laboratory-scale and/or field-scale testing for the removal of arsenic from waters, such as coagulation, filtration, ion exchange, adsorption, and reverse osmosis. However, these technologies are either too expensive or ineffective for low arsenic concentration treatment. To comply with the current regulatory limit of 10 ppb would require extensive technological developments that are highly selective and economically competitive.

In nature, microbes respond to arsenic in a variety of different ways. Depending on the species of different microorganisms, the responses could be chelation, compartmentalization, exclusion, and immobilization. Understanding the molecular and genetic level of arsenic metabolism will be, therefore, an important knowledge base for developing efficient and selective arsenic bioremediation approaches, which has so far been considered as a cost-effective and environmental friendly way for heavy-metal removal. This review highlights the natural arsenic metabolism in different microbes and their impact on environmental arsenic contamination. In addition, the potential utility of these natural metabolisms for arsenic remediation is discussed, as is our current understanding of the biochemistry and molecular biology involved in these natural arsenic metabolisms, and some successful examples of engineered microbes by harnessing these natural mechanisms for effective remediation.

Arsenic metabolism by microbes in nature and the impact on arsenic remediation. Curr Opin Biotechnol. Oct 30 2009

Related:

• Biotransformation of arsenic by algae
• Molecular approaches to bioremediation
• Can fungi clean up radiation releases?

Nice video from the University of Wolverhampton:

The HIV-1 pandemic represents one of the great plagues in human history and a major challenge for medicine, public health and biological research. Infection with HIV-1 causes AIDS, a syndrome that was first described in 1981. Continuous research has allowed the development of effective treatments that have transformed HIV-1 infection from a fatal illness into a chronic disease. Currently, 25 different active compounds belonging to six different drug families have been developed. However, regardless of the use of highly active antiretroviral therapy (HAART), a cure is not yet achievable, and HIV-1 persistence in reservoirs represents the major obstacle for its eradication.

All lentiviruses can infect macrophage lineage cells, in which they generate a persistent infection. HIV-1 is a lentivirus that has developed a broader tropism, leading to preferential infection of CD4+ T cells, which are severely destroyed during the illness. This target provides two potential environments for the virus, allowing latency in resting CD4+ T cells and massive viral replication in activated cells. The molecular mechanisms that lead to HIV-1 reactivation have been characterized in detail, but the study of virus latency remains limited. Recently, the identification of factors that restrict retroviral infections, the characterization of chromatin structure in the setting of viral integration, the discovery of new systems that regulate gene expression (such as small interfering RNAs (siRNAs)) and the development of new techniques for analysing HIV-1 latency have provided a new perspective on this concealed state. Latency should not be considered a merely passive event but, rather, an active process that is maintained by cellular elements. This review analyses the molecular mechanisms that are necessary for the establishment of HIV-1 latency and their relationships with different cellular and anatomical reservoirs, and discusses the current treatment strategies for targeting viral persistence in reservoirs, their main limitations and future perspectives.

Understanding HIV-1 latency provides clues for the eradication of long-term reservoirs. Nature Reviews Microbiology 7, 798 (2009)

Related:

• Why is HIV a pathogen?
• How does HIV cause AIDS?
• Immune exhaustion in HIV infection
• HIV “can never be cured”
• HIV-2 – a kinder AIDS virus?

Fungus growing on spilt birdseed in my garden. I don’t know what species this is – maybe someone could hazard a guess?

Stinking lagoons of pig poo created by thousands of animals in giant pig farms can pollute rivers, poison groundwater and pump out clouds of methane and carbon dioxide. Using microorganisms to break down slurry makes sense for two reasons. The first is environmental protection, but the methane produced by anaerobic digestion can also be used to generate electricity. The problem was that no one has been certain which way of doing it makes the most electricity for the least greenhouse gas production.

Denmark is famous for lots of things, but one of them is pigs. A Danish team has analysed the various ways in which firms in that country treat pig manure and use it to generate electricity in systems such as anaerobic digesters or incinerators. In anaerobic digestion, bacteria break down waste material by warming it in an oxygen-free vessel, releasing methane which is used in gas turbines. Incinerators burn material to boil water and drive a steam turbine. The team found that for high-efficiency energy production, anaerobic digestion is the best answer. But if minimising greenhouse gas emissions takes priority, the best option was to separate the solid from the liquid waste, dry the solids and incinerate them.

Energy Production, Nutrient Recovery and Greenhouse gas Emission Potentials From Integrated pig Manure Management Systems. Waste Manag Res. Sep 1. 2009 doi: 10.1177/0734242×09338728
Improper management of pig manure has resulted in environmental problems such as surface water eutrophication, ground water pollution, and greenhouse gas emissions. This study develops and compares 14 alternative manure management scenarios aiming at energy and nutrient extraction. The scenarios based on combinations of thermal pretreatment, anaerobic digestion, anaerobic co-digestion, liquid/solid separation, drying, incineration, and thermal gasification were compared with respect to their energy, nutrient and greenhouse gas balances. Both sole pig manure and pig manure mixed with other types of waste materials were considered. Data for the analyses were obtained from existing waste treatment facilities, experimental plants, laboratory measurements and literature. The assessment reveals that incineration combined with liquid/solid separation and drying of the solids is a promising management option yielding a high potential energy utilization rate and greenhouse gas savings. If maximum electricity production is desired, anaerobic digestion is advantageous as the biogas can be converted to electricity at high efficiency in a gas engine while allowing production of heat for operation of the digestion process. In conclusion, this study shows that the choice of technology has a strong influence on energy, nutrient and greenhouse gas balances. Thus, to get the most reliable results, it is important to consider the most representative (and up-to-date) technology combined with data representing the area or region in question.

Related:

• New Microbial Fuels
• Microdiesel: microbiology can reduce your carbon footprint
• Termites: wood-munching fuel factories

Hepatitis B virus (HBV) infection is a major health problem, with
400 million people chronically infected worldwide who are at high risk of developing liver cirrhosis and hepatocellular carcinoma (HCC). The epidemiological evidence linking HBV infection to HCC is very strong, and despite the mechanisms underlying HBV-associated carcinogenesis remain to be fully defined, a growing number of studies support a direct role of HBV in the process. The HBV-encoded regulatory protein hepatitis B virus X protein (HBx) is thought to contribute to HBV oncogenicity. HBx transforms SV40-immortalized mouse hepatocytes, induces cell cycle progression within the regenerating liver, causes liver cancer in some transgenic mice, and acts as a cofactor to accelerate cancer development in other mouse models. HBx is a 154-amino acid protein with an N-terminal negative regulatory domain and C-terminal transactivation or coactivation domain that has been detected both in the cytoplasm and in the nuclei of infected hepatocytes. Studies in transfected cells have shown that HBx expression affects cellular functions such as cytoplasmic calcium regulation, cell signaling, transcription, cell proliferation, DNA repair, and apoptosis.

The lack of homology of the X ORF to host protein and its high conservation to other mammalian hepadnaviruses genomes strongly suggest that HBx play a role in virus life cycle. Although initial studies suggested that HBx was not required for virus replication in cell culture, experiments with the highly related woodchuck hepatitis virus (WHV) system indicate that the WHV X protein (WHx) is required for virus replication in vivo. Studies performed using a plasmid-based replication assays that use greater-than-unit-length HBV genomes transfected in HCC cell lines or injected via the mouse tail vein have repeatedly confirmed that HBx potentiate HBV replication. The HBx protein behaves as a promiscuous transactivator of viral and cellular promoters. Although the subcellular localization of HBx seems to be mainly cytoplasmic, a small variable fraction of the protein can be found in the nucleus, and the ability of HBx to activate transcription of host genes is thought to occur indirectly by interaction with nuclear transcription factors or by activation of different signal transduction pathways in the cytoplasm.

Nuclear HBx binds the HBV minichromosome and modifies the epigenetic regulation of cccDNA function. PNAS USA November 11 2009. doi: 10.1073/pnas.0908365106
HBV cccDNA, the template for transcription of all viral mRNAs, accumulates in the nucleus of infected cells as a stable episome organized into minichromosomes by histones and non-histone viral and cellular proteins. Using a cccDNA-specific chromatin immunoprecipitation (ChIP)-based quantitative assay, we have previously shown that transcription of the HBV minichromosome is regulated by epigenetic changes of cccDNA-bound histones and that modulation of the acetylation status of cccDNA-bound H3/H4 histones impacts on HBV replication. We now show that the cellular histone acetyltransferases CBP, p300, and PCAF/GCN5, and the histone deacetylases HDAC1 and hSirt1 are all recruited in vivo onto the cccDNA. We also found that the HBx regulatory protein produced in HBV replicating cells is recruited onto the cccDNA minichromosome, and the kinetics of HBx recruitment on the cccDNA parallels the HBV replication. As expected, an HBV mutant that does not express HBx is impaired in its replication, and exogenously expressed HBx transcomplements the replication defects. p300 recruitment is severely impaired, and cccDNA-bound histones are rapidly hypoacetylated in cells replicating the HBx mutant, whereas the recruitment of the histone deacetylases hSirt1 and HDAC1 is increased and occurs at earlier times. Finally, HBx mutant cccDNA transcribes significantly less pgRNA. Altogether our results further support the existence of a complex network of epigenetic events that influence cccDNA function and HBV replication and identify an epigenetic mechanism (i.e., to prevent cccDNA deacetylation) by which HBx controls HBV replication.

Related:

• Hepatitis B and C viruses
• Hepatitis B virus integration and hepatocarcinogenesis