Cryo‐EM structures of human ABCA7 provide insights into its phospholipid translocation mechanisms

Phospholipid extrusion by ABC subfamily A (ABCA) exporters is central to cellular physiology, although the specifics of the underlying substrate interactions and transport mechanisms remain poorly resolved at the molecular level. Here we report cryo‐EM structures of lipid‐embedded human ABCA7 in an open state and in a nucleotide‐bound, closed state at resolutions between 3.6 and 4.0 Å. The former reveals an ordered patch of bilayer lipids traversing the transmembrane domain (TMD), while the latter reveals a lipid‐free, closed TMD with a small extracellular opening. These structures offer a structural framework for both substrate entry and exit from the ABCA7 TMD and highlight conserved rigid‐body motions that underlie the associated conformational transitions. Combined with functional analysis and molecular dynamics (MD) simulations, our data also shed light on lipid partitioning into the ABCA7 TMD and localized membrane perturbations that underlie ABCA7 function and have broader implications for other ABCA family transporters.     

<Emphasis Type="Italic">C. elegans</Emphasis> behavior of preference choice on bacterial food

Caenorhabditis elegans is a free living soil nematode and thus in its natural habitat, C. elegans encounters many different species of soil bacteria. Although some soil bacteria may be excellent sources of nutrition for the worm, others may be pathogenic. Thus, we undertook a study to understand how C. elegans can identify their preferred food using a simple behavioral assay. We found that there are various species of soil bacteria that C. elegans prefers in comparison to the standard laboratory E. coli strain OP50. In particular, two bacterial strains, Bacillus mycoides and Bacillus soli, were preferred strains. Interestingly, the sole feeding of these bacteria to wild type animals results in extended lifespan through the activation of the autophagic process. Further studies will be required to understand the precise mechanism controlling the behavior of identification and selection of food in C. elegans.     

The effect of provocative tests on electrodiagnosis criteria in clinical carpal tunnel syndrome

Introduction and objectiveNerve conduction study is the most sensitive test for diagnosis of carpal tunnel syndrome (CTS). This test is normal in some patients with mild CTS. Median nerve conduction study evaluation after a provocative test (e.g. wrist flexion) may be helpful for diagnosis of mild CTS. This study aimed to determine the effect of wrist flexion on median nerve conduction in patients suspected to CTS and in healthy subjects.Materials and methodsIn this case-control study, 20 patients (20 hands) with clinical signs of CTS and normal routine electrodiagnosis test results and 20 healthy subjects were investigated. Measured parameters included: median nerve distal sensory latency (DSL), nerve conduction velocity (NCV) across wrist, compound nerve action potential (CNAP), distal motor latency (DML) and compound muscle action potential amplitude (CAMPAMP). The above noted parameters were measured before and after 5 min of full wrist flexion. Data were analyzed using paired T-test.ResultsDistal sensory latency increment and NCV decrimental after 5 min of wrist flexion in the patients group were statistically significant (p < 0.01). The same parameters did not show significant incremental or detrimental changes in the control group.ConclusionMedian nerve DSL and NCV measurement after 5 min of wrist flexion may be helpful in determining more sensitive parameters in the electrodiagnosis of CTS.     

Protective Effects of Garlic and Silymarin on NDEA-Induced Rats Hepatotoxicity

Background ­— The present study was conducted to investigate the chemopreventive effects of garlic extract and silymarin on N-nitrosodiethylamine (NDEA) and carbon tetrachloride (CCl₄)-induced hepatotoxicity in male albino rats. Methods and Results — Animals were pretreated with garlic, silymarin or both for one week prior to the injection of NDEA. Then animals received a single injection of NDEA followed by weekly subcutaneous injections of CCl₄ for 6 weeks. Oral administration was then continued along with the injection of CCl₄ for the duration of the experiment. Serum aspartate transaminase (AST), alanine transaminase (ALT), alkaline phosphatase (ALP), hepatic lipid peroxidation (LPO), superoxide dismutase (SOD), reduced glutathione (GSH), glutathione-S-transferase (GST) and glutathione reductase (GSR) were measured. Injection of NDEA induced a significant elevation in serum AST, ALT and ALP. In the liver, NDEA increased oxidative stress through the increase in LPO and decrease in SOD, and GSH-dependent enzymes. Although administration of garlic or silymarin significantly reduced the liver toxicity, combined administration was more effective in preventing the development of hepatotoxicity. Conclusion — These novel findings suggest that silymarin and garlic have a synergistic effect, and could be used as hepatoprotective agents against hepatotoxicity.     

Mechanism of XIAP-mediated inhibition of caspase-9

The inhibitor of apoptosis (IAP) proteins potently inhibit the catalytic activity of caspases. While profound insight into the inhibition of the effector caspases has been gained in recent years, the mechanism of how the initiator caspase-9 is regulated by IAPs remains enigmatic. This paper reports the crystal structure of caspase-9 in an inhibitory complex with the third baculoviral IAP repeat (BIR3) of XIAP at 2.4 A resolution. The structure reveals that the BIR3 domain forms a heterodimer with a caspase-9 monomer. Strikingly, the surface of caspase-9 that interacts with BIR3 also mediates its homodimerization. We demonstrate that monomeric caspase-9 is catalytically inactive due to the absence of a supporting sequence element that could be provided by homodimerization. Thus, XIAP sequesters caspase-9 in a monomeric state, which serves to prevent catalytic activity. These studies, in conjunction with other observations, define a unified mechanism for the activation of all caspases.     

Polyesters Based on Linoleic Acid for Biolubricant Basestocks: Low-Temperature,Tribological and Rheological Properties

Presently, plant oils which contain high percentage of linoleic acid 1 are perceived to be a viable alternative to mineral oil for biolubricant applications due to their biodegradability and technical properties. In order to get biodegradable lubricant, triester derivatives compounds (1–5) were synthesized and characterized. The processes involved were monoepoxidation of linoleic acid 2, oxirane ring opening 3, esterification 4 and acylation 5. The structures of the products were confirmed by FTIR, ¹H and ¹³C-NMR and LC-MS. The results that showed lowest temperature properties were obtained for triester 5, with a pour point value (PP) of -73°C, highest onset temperature (260°C) and lowest volatility at 0.30%. Viscosity index (VI) increased for the ester’s synthetic compounds (2, 3, 4, 5), while the PP decreased. This behavior is the result of the increase of the chain length of the branching agents. Triester based linoleic acid has improved properties such as low-temperature and tribological properties. These results will make it feasible for plant oil to be used for biolubricants, fuels in chain saws, transmission oil and brake fluid.     

The C-terminal tail of presenilin regulates Omi/HtrA2 protease activity

Presenilin mutations are responsible for most cases of autosomal dominant inherited forms of early onset Alzheimer disease. Presenilins play an important role in amyloid beta-precursor processing, NOTCH receptor signaling, and apoptosis. However, the molecular mechanisms by which presenilins regulate apoptosis are not fully understood. Here, we report that presenilin-1 (PS1) regulates the proteolytic activity of the serine protease Omi/HtrA2 through direct interaction with its regulatory PDZ domain. We show that a peptide corresponding to the cytoplasmic C-terminal tail of PS1 dramatically increases the proteolytic activity of Omi/HtrA2 toward the inhibitor of apoptosis proteins and beta-casein and induces cell death in an Omi/HtrA2-dependent manner. Consistent with these results, ectopic expression of full-length PS1, but not PS1 lacking the C-terminal PDZ binding motif, potentiated Omi/HtrA2-induced cell death. Our results suggest that the C terminus of PS1 is an activation peptide ligand for the PDZ domain of Omi/HtrA2 and may regulate the protease activity of Omi/HtrA2 after its release from the mitochondria during apoptosis. This mechanism of Omi/HtrA2 activation is similar to the mechanism of activation of the related bacterial DegS protease by the outer-membrane porins.     

The relationship between nitric oxide and leptin in the lung of rat with streptozotocin-induced diabetes

Lung structural changes and immunoreactivity of endothelial (eNOS)- and inducible nitric oxide synthase (iNOS) were investigated by light microscopy in lungs of treated and untreated diabetic rats. Diabetes was induced by a single intraperitoneal (i.p.) injection of 65 mg kg(-1) streptozotocin (STZ) in Wistar albino male rats. Diabetic rats received daily i.p. doses of dexamethasone (2 mg kg(-1)), leptin (0.5 microg kg(-1)) and intramuscular insulin (20 U kg(-1)) or a combination of these drugs for 1 week starting 4 weeks after the STZ injections. After treatment, the blood levels of glucose, leptin, insulin and nitrate/nitrite (NO(3) (-)/NO(2) (-)) were measured. Dilatation of alveoli and alveolar ducts, partial alveolar wall thickening and increased eNOS- and iNOS characterized the diabetic rat lungs. High blood glucose and nitrate/nitrite levels as well as low insulin and leptin levels were also present. Treatment with insulin, dexamethasone and a combination of these drugs resulted in improvement of the structural and immunohistochemical abnormalities. The most effective treatment was insulin therapy. Leptin administration resulted in increased relative amounts of extracellular material, which led to noticeable respiratory efficiency in the diabetic rat lungs. All treatments except leptin lowered blood glucose levels. The combination of insulin and dexamethasone increased blood leptin and insulin, while the remaining diabetic rats had blood with low leptin and insulin concentrations. These results suggest that therapy with insulin plus dexamethasone but not therapy with leptin is beneficial for diabetics.     

The Microaerophilic Microbiota of De-Novo Paediatric Inflammatory Bowel Disease: The BISCUIT Study

Introduction

Children presenting for the first time with inflammatory bowel disease (IBD) offer a unique opportunity to study aetiological agents before the confounders of treatment. Microaerophilic bacteria can exploit the ecological niche of the intestinal epithelium; Helicobacter and Campylobacter are previously implicated in IBD pathogenesis. We set out to study these and other microaerophilic bacteria in de-novo paediatric IBD.

Patients and Methods

100 children undergoing colonoscopy were recruited including 44 treatment naïve de-novo IBD patients and 42 with normal colons. Colonic biopsies were subjected to microaerophilic culture with Gram-negative isolates then identified by sequencing. Biopsies were also PCR screened for the specific microaerophilic bacterial groups: Helicobacteraceae, Campylobacteraceae and Sutterella wadsworthensis.

Results

129 Gram-negative microaerophilic bacterial isolates were identified from 10 genera. The most frequently cultured was S. wadsworthensis (32 distinct isolates). Unusual Campylobacter were isolated from 8 subjects (including 3 C. concisus, 1 C. curvus, 1 C. lari, 1 C. rectus, 3 C. showae). No Helicobacter were cultured. When comparing IBD vs. normal colon control by PCR the prevalence figures were not significantly different (Helicobacter 11% vs. 12%, p = 1.00; Campylobacter 75% vs. 76%, p = 1.00; S. wadsworthensis 82% vs. 71%, p = 0.312).

Conclusions

This study offers a comprehensive overview of the microaerophilic microbiota of the paediatric colon including at IBD onset. Campylobacter appear to be surprisingly common, are not more strongly associated with IBD and can be isolated from around 8% of paediatric colonic biopsies. S. wadsworthensis appears to be a common commensal. Helicobacter species are relatively rare in the paediatric colon.

Trial Registration

This study is publically registered on the United Kingdom Clinical Research Network Portfolio (9633).     

Exploring gas permeability of cellular membranes and membrane channels with molecular dynamics

Aquaporins are a family of membrane proteins specialized in rapid water conduction across biological membranes. Whether these channels also conduct gas molecules and the physiological significance of this potential function have not been well understood. Here we report 140 ns of molecular dynamics simulations of membrane-embedded AQP1 and of a pure POPE bilayer addressing these questions. The permeability of AQP1 to two types of gas molecules, O2 and CO2, was investigated using two complementary methods, namely, explicit gas diffusion simulation and implicit ligand sampling. The simulations show that the central (tetrameric) pore of AQP1 can be readily used by either gas molecule to permeate the channel. The two approaches produced similar free energy profiles associated with gas permeation through the central pore: a -0.4 to -1.7 kcal/mol energy well in the middle, and a 3.6-4.6 kcal/mol energy barrier in the periplasmic vestibule. The barrier appears to be mainly due to a dense cluster of water molecules anchored in the periplasmic mouth of the central pore by four aspartate residues. Water pores show a very low permeability to O2, but may contribute to the overall permeation of CO2 due to its more hydrophilic nature. Although the central pore of AQP1 is found to be gas permeable, the pure POPE bilayer provides a much larger cross-sectional area, thus exhibiting a much lower free energy barrier for CO2 and O2 permeation. As such, gas conduction through AQP1 may only be physiologically relevant either in membranes of low gas permeability, or in cells where a major fraction of the cellular membrane is occupied by AQPs.