Combined influence of maternal and paternal quality on sex allocation in red-capped robins

Sex allocation theory predicts females will adaptively manipulate sex ratios to maximize their progeny's reproductive value. Recently, the generality of biased sex allocation in birds has been questioned by meta-analytic reviews, which demonstrate that many previously reported significant results may simply reflect sampling error. Here, we utilize a robust sample size and powerful statistical approach to determine whether parental quality is correlated with biased sex allocation in red-capped robins. Indices of maternal quality (including interactive effects of age and condition) were strongly related to sex allocation. These relationships were in the predicted directions, with larger effect sizes than those of previous studies in this field. There were also paternal correlates, involving age and the source of paternity. We propose that biased sex allocation occurs in this species, and is maintained by differing production costs of each sex and genetic benefits to females of producing sons when fertilized by high-quality males.     

Very Few Substitutions in a Germ Line Antibody Are Required To Initiate Significant Domain Exchange

2G12 is a broadly neutralizing anti-HIV-1 monoclonal human IgG1 antibody reactive with a high-mannose glycan cluster on the surface of glycoprotein gp120. A key feature of this very highly mutated antibody is domain exchange of the heavy-chain variable region (V_H) with the V_H of the adjacent Fab of the same immunoglobulin, which assembles a multivalent binding interface composed of two primary binding sites in close proximity. A non-germ line-encoded proline in the elbow between V_H and C_H1 and an extensive network of hydrophobic interactions in the V_H/V_H′ interface have been proposed to be crucial for domain exchange. To investigate the origins of domain exchange, a germ line version of 2G12 that behaves as a conventional antibody was engineered. Substitution of 5 to 7 residues for those of the wild type produced a significant fraction of domain-exchanged molecules, with no evidence of equilibrium between domain-exchanged and conventional forms. Two substitutions not previously implicated, A^H14 and E^H75, are the most crucial for domain exchange, together with I^H19 at the V_H/V_H′ interface and P^H113 in the elbow region. Structural modeling gave clues as to why these residues are essential for domain exchange. The demonstration that domain exchange can be initiated by a small number of substitutions in a germ line antibody suggests that the evolution of a domain-exchanged antibody response in vivo may be more readily achieved than considered to date.Protein oligomers are able to exchange or swap an element of their secondary structure or an entire protein domain. The functional unit in domain-exchanged proteins thereby stays preserved, as only the linking hinge loop changes conformation significantly (4, 17, 27). Analogous to other domain-swapped proteins, antibodies can exchange an entire domain, in this case the heavy-chain variable region (V_H), with an equivalent heavy-chain variable region of an adjacent Fab (V_H′) within the same immunoglobulin (Ig) molecule (11). The advantages of domain-exchanged proteins, including antibodies, are higher local concentrations of active sites, a larger binding surface, and a potential secondary active site at the new subunit interface (27, 45). The one and only antibody shown to be domain exchanged to date is 2G12 (7, 11), but this arrangement is potentially possible for any Ig and could have been overlooked at least in some instances.2G12 is one of only a few high-affinity monoclonal antibodies with broad neutralizing activity against different subtypes of HIV-1 (5, 30, 40, 43). The antibody binds a dense cluster of N-linked high-mannose glycans (Man_8-9GlcNAc₂) on the envelope surface glycoprotein gp120 (10, 35, 36, 41). The domain-exchanged arrangement forms a multivalent binding site composed of two primary binding sites in close proximity and a proposed secondary binding site formed by the novel V_H/V_H′ interface (11). 2G12 provides protection against infection in animal models (19, 31) and has been shown to induce neutralization escape following passive immunization in humans (39).Consensus has grown that a successful HIV-1 vaccine will need to include a component that elicits broadly neutralizing antibodies (8, 18, 21, 26, 32, 42). All attempts to elicit 2G12-like antibodies with the desired specificity and neutralization activity have failed to date (22, 29, 44), conceivably due to difficulties in generating adequate mimicry of the glycan cluster and tolerance mechanisms or, very likely, the inability to induce domain exchange (1). Unraveling the mechanism of domain exchange and how this conformation might have evolved is highly desirable to direct future HIV-1 vaccine design to elicit 2G12-like antibodies.By comparison with other domain-exchanged proteins (27), the following three mechanisms have been proposed to contribute to the unique structure of 2G12 compared to the structure of a conventional antibody: destabilization of the “closed” V_H/V_L interface, conformational change in the elbow between V_H and C_H1, and an energetically favorable “open” V_H/V_H′ interface (11). Key residues involved in promoting domain exchange were predicted based on examination of interacting residues at the two interfaces and by the effects of alanine substitutions on the binding of wild-type 2G12 to gp120. However, the importance of these key residues for domain exchange was not directly demonstrated experimentally (11).Here, we explored the minimal requirements for domain exchange of 2G12, starting with a germ line version of the antibody that adopts a conventional antibody structure. Although wild-type 2G12 is heavily somatically mutated, only five to seven substitutions in the germ line version of the antibody were shown to produce a significant fraction of domain-exchanged molecules. The results suggest the evolution of domain-exchanged antibody responses may be more facile than considered to date.     

Antibody 2G12 Recognizes Di-Mannose Equivalently in Domain- and Nondomain-Exchanged Forms but Only Binds the HIV-1 Glycan Shield if Domain Exchanged

The broadly neutralizing anti-human immunodeficiency virus type 1 (HIV-1) antibody 2G12 targets the high-mannose cluster on the glycan shield of HIV-1. 2G12 has a unique V_H domain-exchanged structure, with a multivalent binding surface that includes two primary glycan binding sites. The high-mannose cluster is an attractive target for HIV-1 vaccine design, but so far, no carbohydrate immunogen has elicited 2G12-like antibodies. Important questions remain as to how this domain exchange arose in 2G12 and how this unusual event conferred unexpected reactivity against the glycan shield of HIV-1. In order to address these questions, we generated a nondomain-exchanged variant of 2G12 to produce a conventional Y/T-shaped antibody through a single amino acid substitution (2G12 I19R) and showed that, as for the 2G12 wild type (2G12 WT), this antibody is able to recognize the same Manα1,2Man motif on recombinant gp120, Candida albicans, and synthetic glycoconjugates. However, the nondomain-exchanged variant of 2G12 is unable to bind the cluster of mannose moieties on the surface of HIV-1. Crystallographic analysis of 2G12 I19R in complex with Manα1,2Man revealed an adaptable hinge between V_H and C_H1 that enables the V_H and V_L domains to assemble in such a way that the configuration of the primary binding site and its interaction with disaccharide are remarkably similar in the nondomain-exchanged and domain-exchanged forms. Together with data that suggest that very few substitutions are required for domain exchange, the results suggest potential mechanisms for the evolution of domain-exchanged antibodies and immunization strategies for eliciting such antibodies.The broadly neutralizing anti-human immunodeficiency virus type 1 (HIV-1) human monoclonal antibody 2G12 recognizes a highly conserved cluster of oligomannose residues on the glycan shield of the HIV-1 envelope glycoprotein gp120 (9, 10, 36, 39, 44, 45). The antibody binds terminal Manα1,2Man-linked sugars of high-mannose glycans (Man_8-9GlcNAc₂) with nanomolar affinity using a unique domain-exchanged structure in which the variable domains of the heavy chains swap to form a multivalent binding surface that includes two conventional antigen-combining sites and a third potential noncanonical binding site at the novel V_H/V_H′ interface (10). gp120 is one of the most heavily glycosylated proteins identified to date, with approximately 50% of its mass arising from host-derived N-linked glycans (24). These glycans play an important role in shielding the virus from the host immune system (34). Carbohydrates are generally poorly immunogenic, and the dense covering of glycans is often referred to as the “silent face” (52). The oligomannose glycans on gp120 in particular are closely packed, forming a tight cluster, and the unique domain-exchanged structure of 2G12 has been proposed as a means to recognize this cluster (10).The attraction of 2G12 as a template for HIV-1 vaccine design has recently been highlighted in a study that showed the antibody can protect macaques against simian-human immunodeficiency virus (SHIV) challenge at remarkably low serum neutralizing titers (18, 30, 43). When using 2G12 as a template for design of a carbohydrate immunogen, some important considerations must be taken into account. First, 2G12 is unusual in its specificity (targeting host cell-derived glycan motifs presented in a “nonself” arrangement), and although the 2G12 epitope is common to many HIV-1 envelopes, 2G12-like antibodies are rarely elicited (5, 38). Second, due to inherently weak carbohydrate-protein interactions (49, 50), it can be assumed that in order for a carbohydrate-specific antibody to achieve the affinity required to neutralize HIV-1, the avidity of the interaction must be enhanced by both Fab arms of the IgG-contacting glycan motifs simultaneously on the HIV-1 envelope. Third, the unique domain-exchanged structure of 2G12 has not been described for any other antibody (10). These considerations raise a number of questions. Which antigen or sequence of antigens elicited 2G12? Is domain exchange the only solution for recognition of highly clustered oligomannoses? If so, can domain exchange be elicited by immunization with clustered oligomannose motifs (38)?Efforts to design immunogens that elicit responses to the glycan shield of HIV-1 and neutralize the virus have to date been unsuccessful (2, 3, 14, 20, 21, 28, 29, 32, 46-48). Immunogen design strategies that mimic the 2G12 epitope have focused on both chemical and biochemical methods to generate multivalent and clustered displays of both high-mannose sugars (Man_8-9GlcNAc₂) (13, 15, 20, 21, 27-29, 32, 47) and truncated versions of such sugars (Man₉ and Man₄ linked via a 5-carbon linker) (3, 46). These constructs typically bind 2G12 with a lower affinity (on the order of 1 to 3 logs) than recombinant gp120. Although mannose-specific antibodies have been elicited by these immunogens, no HIV-1-neutralizing activities have been described. In a study by Luallen et al., antibodies against recombinant gp120 were generated by immunization with yeast cells that had been mutated to display only Man₈GlcNAc₂ glycans (27, 29). However, no neutralization activity against the corresponding pseudovirus was noted. It was proposed that this was due to either the low abundance of the gp120-specific antibodies in the serum or the antibodies elicited being against carbohydrate epitopes that differed from the 2G12 epitope (27, 29).To gain a better understanding of the importance of domain exchange for glycan recognition and how 2G12 may have been induced, we analyzed the binding characteristics of a nondomain-exchanged (conventional Y/T-shaped) 2G12 variant antibody. This variant was generated by a single point mutation, I19R, that disrupts the V_H/V_H′ interface. We show that the mutant is still able to recognize the Manα1,2Man motif arrayed on yeast, synthetic glycoconjugates, and recombinant gp120 in enzyme-linked immunosorbent assay (ELISA) format but is unable to recognize the discrete, dense mannose clusters found on the surface of the HIV-1 envelope (as measured by neutralization activity and binding to HIV-1-transfected cells). We further show that a major conformational change in the elbow region between V_H and C_H1 in this nondomain-exchanged variant of 2G12 allows the variable domains to assemble in similar orientations with respect to each other, as in the 2G12 wild type (WT), with an identical primary binding site, although with dramatically different orientations with respect to the constant domains. Thus, we conclude that 2G12 recognizes Manα1,2Man motifs in an identical manner in both conventional and domain-exchanged configurations, and the 2G12 specificity for Manα1,2Man likely first arose in a conventional IgG predecessor of 2G12. Subsequent domain exchange was the key event that then enabled high-affinity recognition of the tight oligomannose clusters on HIV-1.     

Vstx1, a modifier of Kv channel gating, localizes to the interfacial region of lipid bilayers

VSTx1 is a tarantula venom toxin which binds to the archaebacterial voltage-gated potassium channel KvAP. VSTx1 is thought to access the voltage sensor domain of the channel via the lipid bilayer phase. In order to understand its mode of action and implications for the mechanism of channel activation, it is important to characterize the interactions of VSTx1 with lipid bilayers. Molecular dynamics (MD) simulations (for a total simulation time in excess of 0.2 micros) have been used to explore VSTx1 localization and interactions with zwitterionic (POPC) and with anionic (POPE/POPG) lipid bilayers. In particular, three series of MD simulations have been used to explore the net drift of VSTx1 relative to the center of a bilayer, starting from different locations of the toxin. The preferred location of the toxin is at the membrane/water interface. Although there are differences between POPC and POPE/POPG bilayers, in both cases the toxin forms favorable interactions at the interface, maximizing H-bonding to lipid headgroups and to water molecules while retaining interactions with the hydrophobic core of the bilayer. A 30 ns unrestrained simulation reveals dynamic partitioning of VSTx1 into the interface of a POPC bilayer. The preferential location of VSTx1 at the interface is discussed in the context of Kv channel gating models and provides support for a mode of action in which the toxin interacts with the Kv voltage sensor "paddle" formed by the S3 and S4 helices.     

A trans-specific polymorphism in ZC3HAV1 is maintained by long-standing balancing selection and may confer susceptibility to multiple sclerosis

The human ZC3HAV1 gene encodes an antiviral protein. The longest splicing isoform of ZC3HAV1 contains a C-terminal PARP-like domain, which has evolved under positive selection in primates. We analyzed the evolutionary history of this same domain in humans and in Pan troglodytes. We identified two variants that segregate in both humans and chimpanzees; one of them (rs3735007) does not occur at a hypermutable site and accounts for a nonsynonymous substitution (Thr851Ile). The probability that the two trans-specific polymorphisms have occurred independently in the two lineages was estimated to be low (P = 0.0054), suggesting that at least one of them has arisen before speciation and has been maintained by selection. Population genetic analyses in humans indicated that the region surrounding the shared variants displays strong evidences of long-standing balancing selection. Selection signatures were also observed in a chimpanzee population sample. Inspection of 1000 Genomes data confirmed these findings but indicated that search for selection signatures using low-coverage whole-genome data may need masking of repetitive sequences. A case-control study of more than 1,000 individuals from mainland Italy indicated that the Thr851Ile SNP is significantly associated with susceptibility to multiple sclerosis (MS) (odds ratio [OR] = 1.47, 95% confidence intervals [CI]: 1.08-1.99, P = 0.011). This finding was confirmed in a larger sample of 4,416 Sardinians cases/controls (OR = 1.18, 95% CI: 1.037-1.344, P = 0.011), but not in a population from Belgium. We provide one of the first instances of human/chimpanzee trans-specific coding variant located outside the major histocompatibility complex region. The selective pressure is likely to be virus driven; in modern populations, this variant associates with susceptibility to MS, possibly via the interaction with environmental factors.     

Phylogenetic relationships among superfamilies of Hymenoptera

The first comprehensive analysis of higher‐level phylogeny of the order Hymenoptera is presented. The analysis includes representatives of all extant superfamilies, scored for 392 morphological characters, and sequence data for four loci (18S, 28S, COI and EF‐1α). Including three outgroup taxa, 111 terminals were analyzed. Relationships within symphytans (sawflies) and Apocrita are mostly resolved. Well supported relationships include: Xyeloidea is monophyletic, Cephoidea is the sister group of Siricoidea + [Xiphydrioidea + (Orussoidea + Apocrita)]; Anaxyelidae is included in the Siricoidea, and together they are the sister group of Xiphydrioidea + (Orussoidea + Apocrita); Orussoidea is the sister group of Apocrita, Apocrita is monophyletic; Evanioidea is monophyletic; Aculeata is the sister group of Evanioidea; Proctotrupomorpha is monophyletic; Ichneumonoidea is the sister group of Proctotrupomorpha; Platygastroidea is sister group to Cynipoidea, and together they are sister group to the remaining Proctotrupomorpha; Proctotrupoidea s. str. is monophyletic; Mymarommatoidea is the sister group of Chalcidoidea; Mymarommatoidea + Chalcidoidea + Diaprioidea is monophyletic. Weakly supported relationships include: Stephanoidea is the sister group of the remaining Apocrita; Diaprioidea is monophyletic; Ceraphronoidea is the sister group of Megalyroidea, which together form the sister group of [Trigonaloidea (Aculeata + Evanioidea)]. Aside from paraphyly of Vespoidea within Aculeata, all currently recognized superfamilies are supported as monophyletic. The diapriid subfamily Ismarinae is raised to family status, Ismaridae stat. nov. © The Will Henning Society 2011.     

External mouthpart morphology in the Tenuipalpidae (Tetranychoidea): Raoiella a case study

The use of low-temperature scanning electron microscopy (LTSEM) to study external mouthpart morphology in the Tenuipalpidae, in particular the genus Raoiella, has brought some aspects of the mechanics of feeding in this group into question. In addition, an LTSEM study on the specialized feeding behaviour of Raoiella indica Hirst (Tetranychoidea: Tenuipalpidae) revealed host plant use in this species could be affected by stomatal complex morphology.     

Regional carbon footprint analysis of dairy feeds for milk production in the USA

Purpose

A greenhouse gas emissions analysis (carbon footprint) was conducted for cultivation, harvesting, and production of common dairy feeds used for the production of dairy milk in the USA. The goal was to determine the carbon footprint (grams CO₂ equivalents (gCO₂e)/kg of dry feed) in the USA on a regional basis, identify key inputs, and make recommendations for emissions reduction.

Methods

Commonly used dairy feeds in the USA, such as soybeans, alfalfa, corn, and others, were identified based on a recent literature review and information from dairy farm surveys. The following input data for the cultivation and harvesting of dairy feeds were collected for five US regions: crop production data, energy input, soil amendments, and crop protection chemicals. Life cycle inventory input data were mainly collected from the US Department of Agriculture National Agricultural Statistical Service on a state-by-state basis as well as from state extension services forage crop budget estimates. In addition to consulting other life cycle assessment studies and published articles and reports, this cradle-to-farm gate carbon footprint analysis was conducted using the Ecoinvent? unit processes in SimaPro version 7.1? (PRé Consultants 2009).

Results

The final carbon footprint results (gCO₂e/kg of dry dairy feed) varied regionally depending on a number of factors such as lime and fertilizer application rates. The average national US carbon footprint results of the main feeds were: corn grain (390), corn silage (200), dried distillers grains with solubles (910 dry mill, 670 wet mill), oats (850), soybeans (390), soybean meal (410), winter wheat (430), alfalfa hay (170), and forage mix (160).

Conclusions and recommendations

The southeast dairy region generally showed a relatively high level of carbon footprint for most feeds, and this is attributable to the higher application rates of both synthetic fertilizers and lime. The highest contributor to carbon footprint for most regions (apart from soybeans and soybean meal) was due to the application of inorganic nitrogen fertilizer. Efficient transfer of knowledge to farmers with regards to fertilizer best management practices such as precision application of farm nutrients may contribute significantly to reducing regional crop carbon footprints.