Anthrax Toxin Receptor 2 Determinants that Dictate the pH Threshold of Toxin Pore Formation

The anthrax toxin receptors, ANTXR1 and ANTXR2, act as molecular clamps to prevent the protective antigen (PA) toxin subunit from forming pores until exposure to low pH. PA forms pores at pH ∼6.0 or below when it is bound to ANTXR1, but only at pH ∼5.0 or below when it is bound to ANTXR2. Here, structure-based mutagenesis was used to identify non-conserved ANTXR2 residues responsible for this striking 1.0 pH unit difference in pH threshold. Residues conserved between ANTXR2 and ANTXR1 that influence the ANTXR2-associated pH threshold of pore formation were also identified. All of these residues contact either PA domain 2 or the neighboring edge of PA domain 4. These results provide genetic evidence for receptor release of these regions of PA as being necessary for the protein rearrangements that accompany anthrax toxin pore formation

Oxidative discolouration in whole-head and cut lettuce: biochemical and environmental influences on a complex phenotype and potential breeding strategies to improve shelf-life

Lettuce discolouration is a key post-harvest trait. The major enzyme controlling oxidative discolouration has long been considered to be polyphenol oxidase (PPO) however, levels of PPO and subsequent development of discolouration symptoms have not always correlated. The predominance of a latent state of the enzyme in plant tissues combined with substrate activation and contemporaneous suicide inactivation mechanisms are considered as potential explanations for this phenomenon. Leaf tissue physical properties have been associated with subsequent discolouration and these may be influenced by variation in nutrient availability, especially excess nitrogen and head maturity at harvest. Mild calcium and irrigation stress has also been associated with a reduction in subsequent discolouration, although excess irrigation has been linked to increased discolouration potentially through leaf physical properties. These environmental factors, including high temperature and UV light intensities, often have impacts on levels of phenolic compounds linking the environmental responses to the biochemistry of the PPO pathway. Breeding strategies targeting the PALand PPOpathway biochemistry and environmental response genes are discussed as a more cost-effective method of mitigating oxidative discolouration then either modified atmosphere packaging or post-harvest treatments, although current understanding of the biochemistry means that such programs are likely to be limited in nature and it is likely that they will need to be deployed alongside other methods for the foreseeable future

Anthrax Toxin Receptor 2–Dependent Lethal Toxin Killing In Vivo

Anthrax toxin receptors 1 and 2 (ANTXR1 and ANTXR2) have a related integrin-like inserted (I) domain which interacts with a metal cation that is coordinated by residue D683 of the protective antigen (PA) subunit of anthrax toxin. The receptor-bound metal ion and PA residue D683 are critical for ANTXR1-PA binding. Since PA can bind to ANTXR2 with reduced affinity in the absence of metal ions, we reasoned that D683 mutant forms of PA might specifically interact with ANTXR2. We show here that this is the case. The differential ability of ANTXR1 and ANTXR2 to bind D683 mutant PA proteins was mapped to nonconserved receptor residues at the binding interface with PA domain 2. Moreover, a D683K mutant form of PA that bound specifically to human and rat ANTXR2 mediated killing of rats by anthrax lethal toxin, providing strong evidence for the physiological importance of ANTXR2 in anthrax disease pathogenesis

Tumor Suppressor Cylindromatosis (CYLD) Controls HIV transcription in an NF-κB-dependent manner

Characterizing the cellular factors that play a role in the HIV replication cycle is fundamental to fully understanding mechanisms of viral replication and pathogenesis. Whole-genome small interfering RNA (siRNA) screens have identified positive and negative regulators of HIV replication, providing starting points for investigating new cellular factors. We report here that silencing of the deubiquitinase cylindromatosis protein (CYLD), increases HIV infection by enhancing HIV long terminal repeat (LTR)-driven transcription via the NF-κB pathway. CYLD is highly expressed in CD4+ T lymphocytes, monocyte-derived macrophages, and dendritic cells. We found that CYLD silencing increases HIV replication in T cell lines. We confirmed the positive role of CYLD silencing in HIV infection in primary human CD4+ T cells, in which CYLD protein was partially processed upon activation. Lastly, Jurkat T cells latently infected with HIV (JLat cells) were more responsive to phorbol 12-myristate 13-acetate (PMA) reactivation in the absence of CYLD, indicating that CYLD activity could play a role in HIV reactivation from latency. In summary, we show that CYLD acts as a potent negative regulator of HIV mRNA expression by specifically inhibiting NF-κB driven transcription. These findings suggest a function for this protein in modulating productive viral replication as well as in viral reactivation

BIRC2/cIAP1 is a Negative Regulator of HIV-1 Transcription and Can Be Targeted by Smac Mimetics to Promote Reversal of Viral Latency

Combination antiretroviral therapy (ART) is able to suppress HIV-1 replication to undetectable levels. However, the persistence of latent viral reservoirs allows for a rebound of viral load upon cessation of therapy. Thus, therapeutic strategies to eradicate the viral latent reservoir are critically needed. Employing a targeted RNAi screen, we identified the ubiquitin ligase BIRC2 (cIAP1), a repressor of the noncanonical NF-κB pathway, as a potent negative regulator of LTR-dependent HIV-1 transcription. Depletion of BIRC2 through treatment with small molecule antagonists known as Smac mimetics enhanced HIV-1 transcription, leading to a reversal of latency in a JLat latency model system. Critically, treatment of resting CD4+ T cells isolated from ART-suppressed patients with the histone deacetylase inhibitor (HDACi) panobinostat together with Smac mimetics resulted in synergistic activation of the latent reservoir. These data implicate Smac mimetics as useful agents for shock-and-kill strategies to eliminate the latent HIV reservoir

Oxidative Stability of Polyunsaturated Edible Oils Mixed With Microcrystalline Cellulose

The oxidative stability of mixtures of edible oils containing polyunsaturated fatty acids (PUFA) and microcrystalline cellulose (MCC) was investigated. The mixtures studied consisted of oils of either camelina (CAM), cod liver (CLO), or salmon (SO) mixed with either colloidal or powdered MCC. A 50:50 (w/w) ratio of oil:MCC resulted in an applicable mixture containing high levels of PUFA edible oil and dietary fiber. The oxidative stability of the formulated mixtures and the pure oils was investigated over a period of 28 days. The peroxide value (PV) was assessed as a parameter for primary oxidation products and dynamic headspace gas chromatography mass spectrometry (GC/MS) was used to analyze secondary volatile organic compounds (VOC). CAM and the respective mixtures were oxidatively stable at both 4 and 22 °C during the storage period. The marine oils and the respective mixtures were stable at 4 °C. At 22 °C, an increase in hydroperoxides was found, but no increase in VOC was detected during the time-frame investigated. At 42 °C, prominent increases in PV and VOC were found for all oils and mixtures. Hexanal, a common marker for the degradation of n-6 fatty acids, propanal and 2,4-heptadienal (E,E), common indicators for the degradation of n-3 fatty acids, were among the volatiles detected in the headspace of oils and mixtures. This study showed that a mixture containing a 50:50 ratio of oil:MCC can be obtained by a low-tech procedure that does not induce oxidation when stored at low temperatures during a period of 1 month

Anthrax Toxin Receptor 2 Determinants that Dictate the pH Threshold of Toxin Pore Formation

The anthrax toxin receptors, ANTXR1 and ANTXR2, act as molecular clamps to prevent the protective antigen (PA) toxin subunit from forming pores until exposure to low pH. PA forms pores at pH ∼6.0 or below when it is bound to ANTXR1, but only at pH ∼5.0 or below when it is bound to ANTXR2. Here, structure-based mutagenesis was used to identify non-conserved ANTXR2 residues responsible for this striking 1.0 pH unit difference in pH threshold. Residues conserved between ANTXR2 and ANTXR1 that influence the ANTXR2-associated pH threshold of pore formation were also identified. All of these residues contact either PA domain 2 or the neighboring edge of PA domain 4. These results provide genetic evidence for receptor release of these regions of PA as being necessary for the protein rearrangements that accompany anthrax toxin pore formation

The role of soluble fiber intake in patients under highly effective lipid-lowering therapy

<p>Abstract</p> <p>Background</p> <p>It has been demonstrated that statins can increase intestinal sterol absorption. Augments in phytosterolemia seems related to cardiovascular disease.</p> <p>Objective</p> <p>We examined the role of soluble fiber intake in endogenous cholesterol synthesis and in sterol absorption among subjects under highly effective lipid-lowering therapy.</p> <p>Design</p> <p>In an open label, randomized, parallel-design study with blinded endpoints, subjects with primary hypercholesterolemia (n = 116) were assigned to receive during 12 weeks, a daily dose of 25 g of fiber (corresponding to 6 g of soluble fibers) plus rosuvastatin 40 mg (n = 28), rosuvastatin 40 mg alone (n = 30), sinvastatin 40 mg plus ezetimibe 10 mg plus 25 g of fiber (n = 28), or sinvastatin 40 mg plus ezetimibe 10 mg (n = 30) alone.</p> <p>Results</p> <p>The four assigned therapies produced similar changes in total cholesterol, LDL-cholesterol, and triglycerides (p < 0.001 vs. baseline) and did not change HDL-cholesterol. Fiber intake decreased plasma campesterol (p < 0.001 vs. baseline), particularly among those patients receiving ezetimibe (p < 0.05 vs. other groups), and β-sitosterol (p = 0.03 vs. baseline), with a trend for lower levels in the group receiving fiber plus ezetimibe (p = 0.07). Treatment with rosuvastatin alone or combined with soluble fiber was associated with decreased levels of desmosterol (p = 0.003 vs. other groups). Compared to non-fiber supplemented individuals, those treated with fibers had weight loss (p = 0.04), reduced body mass index (p = 0.002) and blood glucose (p = 0.047).</p> <p>Conclusion</p> <p>Among subjects treated with highly effective lipid-lowering therapy, the intake of 25 g of fibers added favorable effects, mainly by reducing phytosterolemia. Additional benefits include improvement in blood glucose and anthropometric parameters.</p