Functional conservation of an insect odorant receptor gene across 250 million years of evolution

Pest insects have a profound negative impact on agriculture and human health. Significant global losses of crops, stored agricultural products, timber and livestock can be attributed to damage and destruction by insects . Blood-feeding insects such as mosquitoes, flies and ticks transmit many of humanity's most devastating infectious diseases. Insect-borne diseases account for more than one million annual fatalities, and insect-associated illnesses surpass 300 million annual reported cases . The medical and economic impact of these animals can be ascribed in part to the sensitivity and selectivity of their olfactory systems, essential for location of their preferred plant and animal hosts.     

L-type Ca2+ currents in ventricular myocytes from neonatal and adult rats

Postnatal changes in the slow Ca2+ current (I(Ca)(L)) were investigated in freshly isolated ventricular myocytes from neonatal (1-7 days old) and adult (2-4 months old) rats, using whole-cell voltage clamp and single-channel recordings. The membrane capacitance (mean+/-SEM) averaged 23.2+/-0.5 pF in neonates (n = 163) and 140+/-4.1 pF in adults (n = 143). I(Ca)(L) was measured as the peak inward current at a test potential of +10 mV (or +20 mV) by applying a 300-ms pulse from a holding potential of -40 mV; 1.8 mM Ca2+ was used as charge carrier. The basal ICa(L) density was 6.7+/-0.2 pA/pF in neonatal and 7.8+/-0.2 pA/pF in adult cells (p < 0.05). The time course of inactivation of the fast component (at +10 ms) was significantly longer in the neonatal (10.7+/-1.4 ms) than in the adult (6.6+/-0.4 ms) cells (p < 0.05). Ryanodine (10+/-M) significantly increased this value to 18.0+/-1.9 in neonate (n = 8) and to 17.7+/-2.0 in adult (n = 9). For steady-state inactivation, the half-inactivation potential (Vh) was not changed in either group. For steady-state activation, Vh was 5.1 mV in the neonatal (n = 6) and -7.9 mV in the adult cells (n = 7). Single-channel recordings revealed that long openings (mode-2 behavior) were occasionally observed in the neonatal cells (11 events from 1080 traces/11 cells), but not in the adult cells (400 traces/4 cells). Slope conductance was 24 pS in both the neonatal and adult cells. Results in rat ventricular myocytes suggest the following: (i) the peak Ca2+ current density is already well developed in the neonatal period (being about 85% of the adult value); (ii) the fast component of inactivation is slower in neonates than in adults; and (iii) naturally occurring long openings are occasionally observed in the neonatal stage but not in the adult. Thus, the L-type Ca2+ channels of the neonate were slightly lower in density, were inactivated more slowly, and occasionally exhibited mode-2 behavior as compared with those of the adult.     

Hydroxyethylamine-based inhibitors of BACE1: P1–P3 macrocyclization can improve potency,selectivity, and cell activity

We describe a systematic study of how macrocyclization in the P₁–P₃ region of hydroxyethylamine-based inhibitors of β-site amyloid precursor protein (APP)-cleaving enzyme (BACE1) modulates in vitro activity. This study reveals that in a number of instances macrocyclization of bis-terminal dienes leads to improved potency toward BACE1 and selectivity against cathepsin D (CatD), as well as greater amyloid β-peptide (Aβ)-lowering activity in HEK293T cells stably expressing APP_SW. However, for several closely related analogs the benefits of macrocyclization are attenuated by the effects of other structural features in different regions of the molecules. X-ray crystal structures of three of these novel macrocyclic inhibitors bound to BACE1 revealed their binding conformations and interactions with the enzyme.     

Molecular basis for substrate recognition and septum cleavage by AtlA,the major N-acetylglucosaminidase of Enterococcus faecalis

The cleavage of septal peptidoglycan at the end of cell division facilitates the separation of the two daughter cells. The hydrolases involved in this process (called autolysins) are potentially lethal enzymes that can cause cell death; their activity, therefore, must be tightly controlled during cell growth. In Enterococcus faecalis, the N-acetylglucosaminidase AtlA plays a predominant role in cell separation. atlA mutants form long cell chains and are significantly less virulent in the zebrafish model of infection. The attenuated virulence of atlA mutants is underpinned by a limited dissemination of bacterial chains in the host organism and a more efficient uptake by phagocytes that clear the infection. AtlA has structural homologs in other important pathogens, such as Listeria monocytogenes and Salmonella typhimurium, and therefore represents an attractive model to design new inhibitors of bacterial pathogenesis. Here, we provide a 1.45 Å crystal structure of the E. faecalis AtlA catalytic domain that reveals a closed conformation of a conserved β-hairpin and a complex network of hydrogen bonds that bring two catalytic residues to the ideal distance for an inverting mechanism. Based on the model of the AtlA–substrate complex, we identify key residues critical for substrate recognition and septum cleavage during bacterial growth. We propose that this work will provide useful information for the rational design of specific inhibitors targeting this enterococcal virulence factor and its orthologs in other pathogens.     

A three-step method for isolating a few to several hundred milligrams of protein

An operationally simple general protein isolation method was devised from three previously available separation tools, and was tested by application to two demanding fractionation problems and for yield. One test system was the isolation by gel electrofocusing of two model proteins with pI values of 4.6 and 4.8, bovine serum albumin and ovalbumin, with a load of 220 mg each. The other test was the isolation of 10 mg of human growth hormone isohormone B from a mixture of closely migrating other isohormones. The three-step procedure comprises of: (1) separation into zones of homogeneous protein by gel electrofocusing; (2) excision of the zones of homogeneous protein from the gel followed by concentration of the protein to a small volume of solution by means of Steady-State Stacking; (3) purification from polyacrylamide-like contaminants and non-volatile buffers by gel filtration followed by lyophilization. The average overall recovery was 70--80%.     

Les corps myéloïdes de l'épithelium pigmentaire rétinien

Résumé La morphologie (forme, taille, structure) des corps myéloïdes de l'épithelium pigmentaire rétinien est décrite, ainsi que leur répartition chez les différentes classes de vertébrés. On définit les critères d'identification de ces corps: Ils ne sont jamais limités par une membrane, sont en continuité avec le reticulum endoplasmique lisse, sont formés de saccules aplatis, liés deux à deux par des complexes de jonction. Ils établissent des relations de continuité avec la membrane nucléaire. Les confusions entre corps myéloïdes et diverses autres structures (phagosomes en particulier) sont discutées. Il n'existe pas de corps myéloïdes dans la rétine des Mammifères.

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Myeloid bodies of the retinal pigment epitheliumI. Distribution, morphology and connections with cytoplasmic organels

Summary The morphology (shape, dimensions, structure) of the myeloid bodies of the retinal pigment epithelium and their distribution through several classes of vertebrates is described. The criteria of identity of these bodies are defined: They are never membrane-bounded, they are in direct continuity with the smooth endoplasmic reticulum, they are made of flattened discs linked two and two by junction complexes. They are in direct relation with the nuclear membrane. The confusion between myeloïd bodies and other structures, like phagosomes, is discussed. The retina of Mammals does not contain myeloïd bodies.

Chargé de recherche à l'INSERM. Travail réalisé grace à une subvention de l'INSERM.     

Electron microscopic localization of a fucose-binding protein in acrosome reacted boar spermatozoa by the fucosyl-peroxidase-gold method

In this study we have examined the behaviour and the localization of the fucose-binding protein (FBP) in boar spermatozoa during ionophore induced acrosome reaction (AR) by means of normal TEM and specimen preparation in toto. During early stages of AR the FBP is first localized at the border between equatorial segment and anterior acrosome. With the propagation of the AR the FBP is dramatically expressed and visible over the entire surface of the acrosome and equatorial segment. TEM pictures of this stages show that the FBP is associated with the OAM. At later stages of AR, when acrosomal ghost formation occur, the FBP is associated with the acrosomal ghost, and equatorial segment and to a very low degree also with the IAM. It is concluded from this data that the FBP is responsible for the specific binding of the ghost-sperm unit to the zona pellucida.     

Wrinkling instabilities for biologically relevant fiber-reinforced composite materials with a case study of Neo-Hookean/Ogden–Gasser–Holzapfel bilayer

Biomechanics and Modeling in Mechanobiology - Wrinkling is a ubiquitous surface phenomenon in many biological tissues and is believed to play an important role in arterial health. As arteries are...