Seasonal variations in the diversity and abundance of diazotrophic communities across soils

The nitrogen (N)-fixing community is a key functional community in soil, as it replenishes the pool of biologically available N that is lost to the atmosphere via anaerobic ammonium oxidation and denitrification. We characterized the structure and dynamic changes in diazotrophic communities, based on the nifH gene, across eight different representative Dutch soils during one complete growing season, to evaluate the amplitude of the natural variation in abundance and diversity, and identify possible relationships with abiotic factors. Overall, our results indicate that soil type is the main factor influencing the N-fixing communities, which were more abundant and diverse in the clay soils (n=4) than in the sandy soils (n=4). On average, the amplitude of variation in community size as well as the range-weighted richness were also found to be higher in the clay soils. These results indicate that N-fixing communities associated with sandy and clay soil show a distinct amplitude of variation under field conditions, and suggest that the diazotrophic communities associated with clay soil might be more sensitive to fluctuations associated with the season and agricultural practices. Moreover, soil characteristics such as ammonium content, pH and texture most strongly correlated with the variations observed in the diversity, size and structure of N-fixing communities, whose relative importance was determined across a temporal and spatial scale.     

The mitochondrial contact site complex, a determinant of mitochondrial architecture

Mitochondria are organelles with a complex architecture. They are bounded by an envelope consisting of the outer membrane and the inner boundary membrane (IBM). Narrow crista junctions (CJs) link the IBM to the cristae. OMs and IBMs are firmly connected by contact sites (CS). The molecular nature of the CS remained unknown. Using quantitative high-resolution mass spectrometry we identified a novel complex, the mitochondrial contact site (MICOS) complex, formed by a set of mitochondrial membrane proteins that is essential for the formation of CS. MICOS is preferentially located at the CJs. Upon loss of one of the MICOS subunits, CJs disappear completely or are impaired, showing that CJs require the presence of CS to form a superstructure that links the IBM to the cristae. Loss of MICOS subunits results in loss of respiratory competence and altered inheritance of mitochondrial DNA.     

Galactose-modified iNKT cell agonists stabilized by an induced fit of CD1d prevent tumour metastasis

Invariant natural killer T (iNKT) cells are known to have marked immunomodulatory capacity due to their ability to produce copious amounts of effector cytokines. Here, we report the structure and function of a novel class of aromatic α-galactosylceramide structurally related glycolipids with marked Th1 bias in both mice and men, leading to superior tumour protection in vivo. The strength of the Th1 response correlates well with enhanced lipid binding to CD1d as a result of an induced fit mechanism that binds the aromatic substitution as a third anchor, in addition to the two lipid chains. This induced fit is in contrast to another Th1-biasing glycolipid, α-C-GalCer, whose CD1d binding follows a conventional key-lock principle. These findings highlight the previously unexploited flexibility of CD1d in accommodating galactose-modified glycolipids and broaden the range of glycolipids that can stimulate iNKT cells. We speculate that glycolipids can be designed that induce a similar fit, thereby leading to superior and more sustained iNKT cell responses in vivo.     

The styryl dye FM1-43 suppresses odorant responses in a subset of olfactory neurons by blocking cyclic nucleotide-gated (CNG) channels

Many olfactory receptor neurons use a cAMP-dependent transduction mechanism to transduce odorants into depolarizations. This signaling cascade is characterized by a sequence of two currents: a cation current through cyclic nucleotide-gated channels followed by a chloride current through calcium-activated chloride channels. To date, it is not possible to interfere with these generator channels under physiological conditions with potent and specific blockers. In this study we identified the styryl dye FM1-43 as a potent blocker of native olfactory cyclic nucleotide-gated channels. Furthermore, we characterized this substance to stain olfactory receptor neurons that are endowed with cAMP-dependent transduction. This allows optical differentiation and pharmacological interference with olfactory receptor neurons at the level of the signal transduction.     

Cutting edge: structural basis for the recognition of β-linked glycolipid antigens by invariant NKT cells

Invariant NKT (iNKT) cells expressing a semi-invariant Vα14 TCR recognize self and foreign lipid Ags when presented by the nonclassical MHCI homolog CD1d. Whereas the majority of known iNKT cell Ags are characterized by the presence of a single α-linked sugar, mammalian self Ags are β-linked glycosphingolipids, posing the interesting question of how the semi-invariant TCR can bind to such structurally distinct ligands. In this study, we show that the mouse iNKT TCR recognizes the complex β-linked Ag isoglobotrihexosylceramide (iGb3; Galα1-3-Galβ1-4-Glcβ1-1Cer) by forcing the proximal β-linked sugar of the trisaccharide head group to adopt the typical binding orientation of α-linked glycolipids. The squashed iGb3 orientation is stabilized by several interactions between the trisaccharide and CD1d residues. Finally, the formation of novel contacts between the proximal and second sugar of iGb3 and CDR2α residues of the TCR suggests an expanded recognition logic that can possibly distinguish foreign Ags from self Ags.     

Impact of valency of a glycoprotein B-specific monoclonal antibody on neutralization of herpes simplex virus

Herpes simplex virus (HSV) glycoprotein B (gB) is an integral part of the multicomponent fusion system required for virus entry and cell-cell fusion. Here we investigated the mechanism of viral neutralization by the monoclonal antibody (MAb) 2c, which specifically recognizes the gB of HSV type 1 (HSV-1) and HSV-2. Binding of MAb 2c to a type-common discontinuous epitope of gB resulted in highly efficient neutralization of HSV at the postbinding/prefusion stage and completely abrogated the viral cell-to-cell spread in vitro. Mapping of the antigenic site recognized by MAb 2c to the recently solved crystal structure of the HSV-1 gB ectodomain revealed that its discontinuous epitope is only partially accessible within the observed multidomain trimer conformation of gB, likely representing its postfusion conformation. To investigate how MAb 2c may interact with gB during membrane fusion, we characterized the properties of monovalent (Fab and scFv) and bivalent [IgG and F(ab')(2)] derivatives of MAb 2c. Our data show that the neutralization capacity of MAb 2c is dependent on cross-linkage of gB trimers. As a result, only bivalent derivatives of MAb 2c exhibited high neutralizing activity in vitro. Notably, bivalent MAb 2c not only was capable of preventing mucocutaneous disease in severely immunodeficient NOD/SCID mice upon vaginal HSV-1 challenge but also protected animals even with neuronal HSV infection. We also report for the first time that an anti-gB specific monoclonal antibody prevents HSV-1-induced encephalitis entirely independently from complement activation, antibody-dependent cellular cytotoxicity, and cellular immunity. This indicates the potential for further development of MAb 2c as an anti-HSV drug.