Ten years of life in compost: temporal and spatial variation of North German Caenorhabditis elegans populations

The nematode Caenorhabditis elegans is a central laboratory model system in almost all biological disciplines, yet its natural life history and population biology are largely unexplored. Such information is essential for in‐depth understanding of the nematode's biology because its natural ecology provides the context, in which its traits and the underlying molecular mechanisms evolved. We characterized natural phenotypic and genetic variation among North German C. elegans isolates. We used the unique opportunity to compare samples collected 10 years apart from the same compost heap and additionally included recent samples for this and a second site, collected across a 1.5‐year period. Our analysis revealed significant population genetic differentiation between locations, across the 10‐year time period, but for only one location a trend across the shorter time frame. Significant variation was similarly found for phenotypic traits of likely importance in nature, such as choice behavior and population growth in the presence of pathogens or naturally associated bacteria. Phenotypic variation was significantly influenced by C. elegans genotype, time of isolation, and sampling site. The here studied C. elegans isolates may provide a valuable, genetically variable resource for future dissection of naturally relevant gene functions.     

A Flexible Docking Scheme Efficiently Captures the Energetics of Glycan-Cyanovirin Binding

Cyanovirin-N (CVN), a cyanobacterial lectin, exemplifies a class of antiviral agents that inhibit HIV by binding to the highly glycosylated envelope protein gp120. Here, we investigate the energetics of glycan recognition using a computationally inexpensive flexible docking approach, backbone perturbation docking (BP-Dock). We benchmarked our method using two mutants of CVN: P51G-m4-CVN, which binds dimannose with high affinity through domain B, and CVN^(mutDB), in which binding to domain B has been abolished through mutation of five polar residues to small nonpolar side chains. We investigated the energetic contribution of these polar residues along with the additional position 53 by docking dimannose to single-point CVN mutant models. Analysis of the docking simulations indicated that the E41A/G and T57A mutations led to a significant decrease in binding energy scores due to rearrangements of the hydrogen-bond network that reverberated throughout the binding cavity. N42A decreased the binding score to a level comparable to that of CVN^(mutDB) by affecting the integrity of the local protein structure. In contrast, N53S resulted in a high binding energy score, similar to P51G-m4-CVN. Experimental characterization of the five mutants by NMR spectroscopy confirmed the binding affinity pattern predicted by BP-Dock. Despite their mostly conserved fold and stability, E41A, E41G, and T57A displayed dissociation constants in the millimolar range. N53S showed a binding constant in the low micromolar range, similar to that observed for P51G-m4-CVN. No binding was observed for N42A. Our results show that BP-Dock is a useful tool for rapidly screening the relative binding affinity pattern of in silico-designed mutants compared with wild-type, supporting its use to design novel mutants with enhanced binding properties.     

Arabidopsis RETINOBLASTOMA-RELATED Is Required for Stem Cell Maintenance, Cell Differentiation, and Lateral Organ Production

Several genes involved in the regulation of postembryonic organ initiation and growth have been identified. However, it remains largely unclear how developmental cues connect to the cell cycle. RETINOBLASTOMA RELATED (RBR) is a plant homolog of the tumor suppressor Retinoblastoma (pRb), which is a key regulator of the cell cycle. Using inducible RNA interference (RNAi) against Arabidopsis thaliana RBR (RBRi), we reduced RBR expression levels at different stages of plant development. Conditional reduction or loss of RBR function disrupted cell division patterns, promoted context-dependent cell proliferation, and negatively influenced establishment of cell differentiation. Several lineages of toti- and pluripotent cells, including shoot apical meristem stem cells, meristemoid mother cells, and procambial cells, failed to produce appropriately differentiated cells. Meristem activity was altered, leading to a disruption of the CLAVATA-WUSCHEL feedback loop and inhibition of lateral organ formation. Release of RBR from RNAi downregulation restored meristem activity. Gene profiling analyses soon after RBRi induction revealed that a change in RBR homeostasis is perceived as a stress, even before genes regulated by RBR-E2F become deregulated. The results establish RBR as a key cell cycle regulator required for coordination of cell division, differentiation, and cell homeostasis.     

Negative Potentials Across Biological Membranes Promote Fusion by Class II and Class III Viral Proteins

Voltage was investigated as a factor in the fusion of virions. Virions, pseudotyped with a class II, SFV E1 or VEEV E, or a class III protein, VSV G, were prepared with GFP within the core and a fluorescent lipid. This allowed both hemifusion and fusion to be monitored. Voltage clamping the target cell showed that fusion is promoted by a negative potential and hindered by a positive potential. Hemifusion occurred independent of polarity. Lipid dye movement, in the absence of content mixing, ceased before complete transfer for positive potentials, indicating that reversion of hemifused membranes into two distinct membranes is responsible for voltage dependence and inhibition of fusion. Content mixing quickly followed lipid dye transfer for a negative potential, providing a direct demonstration that hemifusion induced by class II and class III viral proteins is a functional intermediate of fusion. In the hemifused state, virions that fused exhibited slower lipid transfer than did nonfusing virions. All viruses with class II or III fusion proteins may utilize voltage to achieve infection.     

X inactivation and disease

X inactivation is the mechanism by which mammals adjust the X-linked gene dosage between the sexes. The dosage difference between XX females and XY males is functionally equalized by silencing one of the two X chromosomes in female cells. This dosage-compensation mechanism is based on the long functional Xist RNA. Here, we review our understanding of dosage compensation and Xist function in the context of disease.     

Binding site on human immunoglobulin G for the affinity ligand HWRGWV

Affinity ligand HWRGWV has demonstrated the ability to isolate human immunoglobulin G (hIgG) from mammalian cell culture media. The ligand specifically binds hIgG through its Fc portion. This work shows that deglycosylation of hIgG has no influence on its binding to the HWRGWV ligand and the ligand does not compete with Protein A or Protein G in binding hIgG. It is suggested by the mass spectrometry (MS) data and docking simulation that HWRGWV binds to the pFc portion of hIgG and interacts with the amino acids in the loop Ser383–Asn389 (SNGQPEN) located in the C_H3 domain. Subsequent modeling has suggested a possible three‐dimensional minimized solution structure for the interaction of hIgG and the HWRGWV ligand. The results support the fact that a peptide as small as a hexamer can have specific interactions with large proteins such as hIgG. Copyright © 2009 John Wiley & Sons, Ltd.     

The role of dendritic cells in the pathogenesis of systemic lupus erythematosus

The etiology of the autoimmune disease systemic lupus erythematosus is not known, but aberrant apoptosis and/or insufficient clearance of apoptotic material have been assigned a pivotal role. During apoptosis, nucleosomes and several endogenous danger-associated molecular patterns are incorporated in blebs. Recent data indicate that apoptotic blebs induce maturation of myeloid dendritic cells, resulting in IL-17 production by T cells. In this review we summarize current knowledge on the role of dendritic cells in the pathogenesis of systemic lupus erythematosus with special emphasis on the uptake of apoptotic blebs by dendritic cells, and the subsequent induction of Th17 cells.