Risk Factors for Death from Influenza A(H1N1)pdm09, State of S?o Paulo,Brazil, 2009

This case-control study aimed to assess the risk factors for death from influenza A(H1N1)pdm09 in patients with laboratory confirmation, who had severe acute respiratory illness-SARI and were hospitalized between June 28^th and August 29^th 2009, in the metropolitan regions of São Paulo and Campinas, Brazil. Medical charts of all the 193 patients who died (cases) and the 386 randomly selected patients who recovered (controls) were investigated in 177 hospitals. Household interviews were conducted with those who had survived and the closest relative of those who had died. 73.6% of cases and 38.1% of controls were at risk of developing influenza-related complications. The 18-to-59-year age group (OR = 2.31, 95%CI: 1.31–4.10 (reference up to 18 years of age)), presence of risk conditions for severity of influenza (OR = 1.99, 95%CI: 1.11–3.57, if one or OR = 6.05, 95%CI: 2.76–13.28, if more than one), obesity (OR = 2.73, 95%CI: 1.28–5.83), immunosuppression (OR = 3.43, 95%CI: 1.28–9.19), and search for previous care associated with the hospitalization (OR = 3.35, 95%CI: 1.75–6.40) were risk factors for death. Antiviral treatment performed within 72 hours of the onset of symptoms (OR = 0.17, 95%CI: 0.08–0.37, if within 48hours, and OR = 0.30, 95%CI: 0.11–0.81, if between 48 and 72 hours) was protective against death. The identification of high-risk patients and early treatment are important factors for reducing morbi-mortality from influenza.     

Dishevelled,a Wnt signalling component,is involved in mitotic progression in cooperation with Plk1

Wnt signalling is known to promote G1/S progression through the stimulation of gene expression, but whether this signalling regulates mitotic progression is not clear. Here, the function of dishevelled 2 (Dvl2), which transmits the Wnt signal, in mitosis was examined. Dvl2 localized to the spindles and spindle poles during mitosis. When cells were treated with nocodazole, Dvl2 was observed at the kinetochores (KTs). Dvl2 bound to and was phosphorylated at Thr206 by a mitotic kinase, Polo‐like kinase 1 (Plk1), and this phosphorylation was required for spindle orientation and stable microtubule (MT)‐KT attachment. Dvl2 was also found to be involved in the activation of a spindle assembly checkpoint (SAC) kinase, Mps1, and the recruitment of other SAC components, Bub1 and BubR1, to the KTs. However, the phosphorylation of Dvl2 by Plk1 was dispensable for SAC. Furthermore, Wnt receptors were involved in spindle orientation, but not in MT‐KT attachment or SAC. These results suggested that Dvl2 is involved in mitotic progression by regulating the dynamics of MT plus‐ends and the SAC in Plk1‐dependent and ‐independent manners.     

Binding of YC-1/BAY 41-2272 to soluble guanylate cyclase: A new perspective to the mechanism of activation

Soluble guanylate cyclase (sGC), a heterodimeric heme protein, catalyses the conversion of GTP in to cyclic GMP, which acts as a second messenger in cellular signaling. Nitric oxide activates this enzyme several hundred folds over its basal level. Carbon monoxide, along with some activator molecules like YC-1 and BAY, also synergistically activate sGC. Mechanism of this synergistic activation is a matter of debate. Here we review the existing literature to identify the possible binding site for YC-1 and BAY on bovine lung sGC and its mechanism of activation. These two exogenous compounds bind sGC on α subunit inside a pocket and thus exert allosteric effect via subunit interface, which is relayed to the catalytic site. We used docking studies to further validate this hypothesis. We propose that the binding of YC-1/BAY inside the sensory domain of the α subunit modulates the interactions on the subunit interface resulting in rearrangements in the catalytic site into active conformation and this partly induces the cleavage of Fe-His bond.     

Identification of Cys385 in the isolated kinase insertion domain of heme-regulated eIF2 alpha kinase (HRI) as the heme axial ligand by site-directed mutagenesis and spectral characterization

Heme-regulated eIF2alpha kinase (HRI) is an important enzyme that modulates protein synthesis during cellular emergency/stress conditions, such as heme deficiency in red cells. It is essential to identify the heme axial ligand(s) and/or binding sites to establish the heme regulation mechanism of HRI. Previous reports suggest that a His residue in the N-terminal region and a Cys residue in the C-terminal region trans to the His are axial ligands of the heme. Moreover, mutational analyses indicate that a residue located in the kinase insertion (KI) domain between Kinase I and Kinase II domains in the C-terminal region is an axial ligand. In the present study, we isolate the KI domain of mouse HRI and employ site-directed mutagenesis to identify the heme axial ligand. The optical absorption spectrum of the Fe(III) hemin-bound wild-type KI displays a broad Soret band at around 373nm, while that of the Fe(II) heme-bound protein contains a band at 422nm. Spectral titration studies conducted for both the Fe(III) hemin and Fe(II) heme complexes with KI support a 1:1 stoichiometry of heme iron to protein. Resonance Raman spectra of Fe(III) hemin-bound KI suggest that thiol is the axial ligand in a 5-coordinate high-spin heme complex as a major form. Electron spin resonance (ESR) spectra of Fe(III) hemin-bound KI indicate that the axial ligands are OH(-) and Cys. Since Cys385 is the only cysteine in KI, the residue was mutated to Ser, and its spectral characteristics were analyzed. The Soret band position, heme spectral titration behavior and ESR parameters of the Cys385Ser mutant were markedly different from those of wild-type KI. Based on these spectroscopic findings, we conclude that Cys385 is an axial ligand of isolated KI.     

Structural basis for innate immune sensing by M-ficolin and its control by a pH-dependent conformational switch

Ficolins are soluble oligomeric proteins with lectin-like activity, assembled from collagen fibers prolonged by fibrinogen-like recognition domains. They act as innate immune sensors by recognizing conserved molecular markers exposed on microbial surfaces and thereby triggering effector mechanisms such as enhanced phagocytosis and inflammation. In humans, L- and H-ficolins have been characterized in plasma, whereas a third species, M-ficolin, is secreted by monocytes and macrophages. To decipher the molecular mechanisms underlying their recognition properties, we previously solved the structures of the recognition domains of L- and H-ficolins, in complex with various model ligands (Garlatti, V., Belloy, N., Martin, L., Lacroix, M., Matsushita, M., Endo, Y., Fujita, T., Fontecilla-Camps, J. C., Arlaud, G. J., Thielens, N. M., and Gaboriaud, C. (2007) EMBO J. 24, 623-633). We now report the ligand-bound crystal structures of the recognition domain of M-ficolin, determined at high resolution (1.75-1.8 A), which provides the first structural insights into its binding properties. Interaction with acetylated carbohydrates differs from the one previously described for L-ficolin. This study also reveals the structural determinants for binding to sialylated compounds, a property restricted to human M-ficolin and its mouse counterpart, ficolin B. Finally, comparison between the ligand-bound structures obtained at neutral pH and nonbinding conformations observed at pH 5.6 reveals how the ligand binding site is dislocated at acidic pH. This means that the binding function of M-ficolin is subject to a pH-sensitive conformational switch. Considering that the homologous ficolin B is found in the lysosomes of activated macrophages (Runza, V. L., Hehlgans, T., Echtenacher, B., Zahringer, U., Schwaeble, W. J., and Mannel, D. N. (2006) J. Endotoxin Res. 12, 120-126), we propose that this switch could play a physiological role in such acidic compartments.     

Soybean-derived phosphatidylinositol inhibits in vivo low concentrations of amyloid beta protein-induced degeneration of hippocampal neurons in V337M human tau-expressing mice

In our previous reports using primary cultured rat hippocampal neurons, pathophysiological concentrations (< or =10 nM) of amyloid beta proteins (Abetas) showed neurotoxicity via a phosphatidylinositol metabolism disorder, and soybean-derived phosphatidylinositol protected the neurons against the Abeta's neurotoxicity. In the present study, such a neurotoxic effect of Abeta and a neuroprotective effect of phosphatidylinositol were examined in vivo using transgenic mice expressing V337 M human tau. Intrahippocampal CA1 injection of 1.5 mul of 100 nM or 1 microM Abeta25-35 increased the number of degenerating neurons with an apoptotic feature in bilateral hippocampal CA1, CA2, CA3 and dentate gyrus regions in 1 month, demonstrating an in vivo neurotoxic effect of Abeta at lower concentrations after diffusion. Intrahippocampal co-injection or intracerebroventricular administration of 1.5 microl of 500 nM phosphatidylinositol prevented the Abeta25-35-induced neuronal degeneration in all the hippocampal regions, while co-injection of another acidic phospholipid, phosphatidylserine (1.5 microl, 500 nM) with Abeta25-35 showed no protective effects. Thus, exogenously applied phosphatidylinositol appeared to minimize the toxic effects of Abeta in vivo. These results suggest that soybean-derived phosphatidylinositol may be effective in the treatment of Alzheimer's disease.     

4-Pyridone derivatives as new inhibitors of bacterial enoyl-ACP reductase FabI

Bacterial FAS provides essential fatty acids for use in the assembly of key cellular components. Among them, FabI is an enoyl-ACP reductase which catalyzes the final and rate-limiting step of bacterial FAS. It is a potential target for selective antibacterial action, because it shows low overall sequence homology with mammalian enzymes. Until today, various compounds have been reported as inhibitors of bacterial FabI-inhibitory compounds. To discover novel small-molecular FabI inhibitors, we initially screened our compound library for inhibitory activity toward FabI of Escherichia coli. And discovered 4-pyridone derivatives as a lead compound. Structure optimization studies yielded 4-pyridone derivatives 7n having strong FabI-inhibitory and antibacterial activities against Staphylococcus aureus. There have been no reports concerning 4-pyridone derivatives as FabI inhibitor.