Stabilizing the Native Trimer of HIV-1 Env by Destabilizing the Heterodimeric Interface of the gp41 Postfusion Six-Helix Bundle

The HIV-1 envelope glycoprotein (Env) is a trimer of gp120-gp41 heterodimers and is essential for viral entry. The gp41 subunit in native, prefusion trimeric Env exists in a metastable conformation and attains a stable six-helix bundle (6-HB) conformation comprised of a trimer of N-heptad repeat (NHR) and C-heptad repeat (CHR) heterodimers, that drives the fusion of viral and cellular membranes. We attempted to stabilize native Env trimers by incorporation of mutations at the NHR-CHR interface that disrupt the postfusion 6-HB of gp41. The mutations V570D and I573D stabilize native Env of the HIV-1 JRFL strain and occlude nonneutralizing epitopes to a greater extent than the previously identified I559P mutation that is at the interface of the NHR trimers in the 6-HB. The mutations prevent soluble-CD4 (sCD4)-induced gp120 shedding and 6-HB formation. In the context of cell surface-expressed JRFL Env, introduction of a previously reported additional disulfide between residues A501 and T605 perturbs the native conformation, though this effect is partially alleviated by furin coexpression. The data suggest that positions 570 and 573 are surface proximal in native Env and that the NHR homotrimeric coiled coil in native Env terminates before or close to residue 573. Aspartic acid substitutions at these positions stabilize native trimers through destabilization of the postfusion 6-HB conformation. These mutations can be used to stabilize Env in a DNA vaccine format. IMPORTANCE The major protein on the surface of HIV-1 is the envelope (Env) glycoprotein. Env is a trimer of gp120-gp41 heterodimers. gp120 is involved in receptor/coreceptor binding and gp41 in the fusion of viral and cellular membranes. Like many other viral fusion proteins, the gp41 subunit in native trimeric Env exists in a metastable conformation. gp41 readily forms a stable six-helix bundle (6-HB) conformation comprised of a trimer of N-heptad repeat (NHR) and C-heptad repeat (CHR) heterodimers that drives fusion of viral and cellular membranes. While it is expected that native Env is a good immunogen, its metastability results in exposure of immunodominant nonneutralizing epitopes. In the present study, we stabilize native Env trimers by incorporation of a number of different mutations at the NHR-CHR interface that disrupt the postfusion 6-HB of gp41. The stabilized constructs described here can be incorporated into DNA vaccine candidates.

Nutritional benefits from domatia inhabitants in an ant-plant interaction: interlopers do pay the rent

1. How a symbiosis originates and is maintained are important evolutionary questions. Symbioses in myrmecophytes (plants providing nesting for ants) are believed to be maintained by protection and nutrients provided by specialist plant-ants in exchange for nesting spaces (called domatia) and nourishment offered by ant-plants. However, besides the benefits accrued from housing protective ants, the mechanisms contributing to the fitness advantages of bearing domatia have rarely been examined, especially because the domatia trait is usually constitutively expressed, and many myrmecophytes have obligate mutualisms with single ant species resulting in invariant conditions. 2. In the unspecialized ant-plant Humboldtia brunonis (Fabaceae) that offers extrafloral nectar to ants, only some plants produce domatia in the form of hollow internodes. These domatia have a self-opening slit making them more prone to interlopers and are occupied mostly by non-protective ants and other invertebrates, especially arboreal earthworms. The protection mutualism with ants is restricted in geographical extent, occurring only at a few sites in the southernmost part of this plant's range in the Western Ghats of India. 3. We examined nutrient flux from domatia residents to the plant using stable isotopes. We found that between 9% (earthworms) and 17% (protective or non-protective ants) of nitrogen of plant tissues nearest the domatium came from domatia inhabitants. Therefore, interlopers such as earthworms and non-protective ants contributed positively to the nitrogen budget of localized plant modules of this understorey tree. N-15-enriched feeding experiments with protective ants demonstrated that nutrients flowed from domatia inhabitants to nearby plant modules. Fruit set did not differ between paired hand-pollinated inflorescences on domatia and non-domatia bearing branches. This was possibly due to the nutrient flux from domatia to adjacent branches without domatia within localized modules. 4. This study has demonstrated the nutritive role of non-protective ants and non-ant invertebrates, hitherto referred to as interlopers, in an unspecialized myrmecophyte. Our study suggests that even before the establishment of a specialized ant-plant protection mutualism, nutritional benefits conferred by domatia inhabitants can explain the fitness benefits of bearing domatia, and thus the maintenance of a trait that facilitates the establishment of a specialized ant-plant symbiosis.

Fruit and Seed Volatiles: Multiple Stage Settings, Actors and Props in an Evolutionary Play

Plants emit volatile organic compounds (VOCs) from most parts of their anatomy. Conventionally, the volatiles of leaves, flowers, fruits and seeds have been investigated separately. This review presents an integrated perspective of volatiles produced by fruits and seeds in the context of selection on the whole plant. It suggests that fruit and seed volatiles may only be understood in the light of the chemistry of the whole plant. Fleshy fruit may be viewed as an ecological arena within which several evolutionary games are being played involving fruit VOCs. Fruit odour and colour may be correlated and interact via multimodal signalling in influencing visits by frugivores. The hypothesis of volatile crypsis in the evolution of hard seeds as protection against volatile diffusion and perception by seed predators is reviewed. Current views on the role of volatiles in ant dispersal of seeds or myrmecochory are summarised, especially the suggestion that ants are being manipulated by plants in the form of a sensory trap while providing this service. Plant VOC production is presented as an emergent phenotype that could result from multiple selection pressures acting on various plant parts; the ``plant'' phenotype and VOC profile may receive significant contributions from symbionts within the plant. Viewing the plant as a holobiont would benefit an understanding of the emergent plant phenotype.

And yet it shrinks: A novel method for correcting bias in forest tree growth estimates caused by water-induced fluctuations

Accuracy in tree woody growth estimates is important to global carbon budget estimation and climate-change science. Tree growth in permanent sampling plots (PSPs) is commonly estimated by measuring stem diameter changes, but this method is susceptible to bias resulting from water-induced reversible stem shrinkage. In the absence of bias correction, temporal variability in growth is likely to be overestimated and incorrectly attributed to fluctuations in resource availability, especially in forests with high seasonal and inter-annual variability in water. We propose and test a novel approach for estimating and correcting this bias at the community level. In a 50-ha PSP from a seasonally dry tropical forest in southern India, where tape measurements have been taken every four years from 1988 to 2012, for nine trees we estimated bias due to reversible stem shrinkage as the difference between woody growth measured using tree rings and that estimated from tape. We tested if the bias estimated from these trees could be used as a proxy to correct bias in tape-based growth estimates at the PSP scale. We observed significant shrinkage-related bias in the growth estimates of the nine trees in some censuses. This bias was strongly linearly related to tape-based growth estimates at the level of the PSP, and could be used as a proxy. After bias was corrected, the temporal variance in growth rates of the PSP decreased, while the effect of exceptionally dry or wet periods was retained, indicating that at least a part of the temporal variability arose from reversible shrinkage-related bias. We also suggest that the efficacy of the bias correction could be improved by measuring the proxy on trees that belong to different size classes and census timing, but not necessarily to different species. Our approach allows for reanalysis - and possible reinterpretation of temporal trends in tree growth, above ground biomass change, or carbon fluxes in forests, and their relationships with resource availability in the context of climate change. (C) 2014 Elsevier B.V. All rights reserved.

An estimate of the number of tropical tree species

The high species richness of tropical forests has long been recognized, yet there remains substantial uncertainty regarding the actual number of tropical tree species. Using a pantropical tree inventory database from closed canopy forests, consisting of 657,630 trees belonging to 11,371 species, we use a fitted value of Fisher's alpha and an approximate pantropical stem total to estimate the minimum number of tropical forest tree species to fall between similar to 40,000 and similar to 53,000, i.e., at the high end of previous estimates. Contrary to common assumption, the Indo-Pacific region was found to be as species-rich as the Neotropics, with both regions having a minimum of similar to 19,000-25,000 tree species. Continental Africa is relatively depauperate with a minimum of similar to 4,500-6,000 tree species. Very few species are shared among the African, American, and the Indo-Pacific regions. We provide a methodological framework for estimating species richness in trees that may help refine species richness estimates of tree-dependent taxa.

An approximation to the QoS aware throughput region of a tree network under IEEE 802.15.4 CSMA/CA with application to wireless sensor network design

In the context of wireless sensor networks, we are motivated by the design of a tree network spanning a set of source nodes that generate packets, a set of additional relay nodes that only forward packets from the sources, and a data sink. We assume that the paths from the sources to the sink have bounded hop count, that the nodes use the IEEE 802.15.4 CSMA/CA for medium access control, and that there are no hidden terminals. In this setting, starting with a set of simple fixed point equations, we derive explicit conditions on the packet generation rates at the sources, so that the tree network approximately provides certain quality of service (QoS) such as end-to-end delivery probability and mean delay. The structures of our conditions provide insight on the dependence of the network performance on the arrival rate vector, and the topological properties of the tree network. Our numerical experiments suggest that our approximations are able to capture a significant part of the QoS aware throughput region (of a tree network), that is adequate for many sensor network applications. Furthermore, for the special case of equal arrival rates, default backoff parameters, and for a range of values of target QoS, we show that among all path-length-bounded trees (spanning a given set of sources and the data sink) that meet the conditions derived in the paper, a shortest path tree achieves the maximum throughput. (C) 2015 Elsevier B.V. All rights reserved.

Fruit Trait Preference in Rhesus Macaques (Macaca mulatta) and its Implications for Seed Dispersal

Primates constitute 25-40 % of the frugivore biomass of tropical forests. Primate fruit preference, as a determinant of seed dispersal, can therefore have a significant impact on these ecosystems. Although the traits of fruits included in primate diets have been described, fruit trait preference has been less studied with respect to fruit availability. We examined fruit trait preference and its implications for seed dispersal in the rhesus macaque (Macaca mulatta), a dietarily flexible species and important seed disperser, at the Buxa Tiger Reserve, India. Over a year, we monitored the phenology of selected trees in the study area, observed the feeding behavior of rhesus macaques using scans and focal animal sampling, and documented morphological traits of the fruits/seeds consumed. Using generalized linear modeling, we found that the kind of edible tissue was the chief determinant of fruit consumption, with M. mulatta feeding primarily on fruits with juicy-soft pulp and acting as seed predators for those with no discernible pulp. Overall, the preferred traits were external covers that could be easily pierced by a fingernail, medium to large seeds, true stone-like seeds, and juicy-soft edible tissue, thereby implying that fruit taxa with these traits had a higher probability of being dispersed. Macaques were more selective during the high fruit availability period than the low fruit availability period, preferentially feeding on soft-skinned fruits with juicy-soft pulp. We suggest that further studies be conducted across habitats and time to understand the consistency of interactions between primates and fruits with specific traits to determine the degree of selective pressure (if any) that is exerted by primates on fruit traits.