Key pointsr We recently found that feeding healthy mice a diet with reduced levels of branched-chain amino acids (BCAAs), which are associated with insulin resistance in both humans and rodents, modestly improves glucose tolerance and slows fat mass gain.r In the present study, we show that a reduced BCAA diet promotes rapid fat mass loss without calorie restriction in obese mice.r Selective reduction of dietary BCAAs also restores glucose tolerance and insulin sensitivity to obese mice, even as they continue to consume a high-fat, high-sugar diet.r A low BCAA diet transiently induces FGF21 (fibroblast growth factor 21) and increases energy expenditure.r We suggest that dietary protein quality (i.e. the precise macronutrient composition of dietary protein) may impact the effectiveness of weight loss diets.Abstract Obesity and diabetes are increasing problems around the world, and although even moderate weight loss can improve metabolic health, reduced calorie diets are notoriously difficult to sustain. Branched-chain amino acids (BCAAs; leucine, isoleucine and valine) are elevated in the blood of obese, insulin-resistant humans and rodents. We recently demonstrated that specifically reducing dietary levels of BCAAs has beneficial effects on the metabolic health of young, growing mice, improving glucose tolerance and modestly slowing fat mass gain. In the present study, we examine the hypothesis that reducing dietary BCAAs will promote weight loss, reduce adiposity, and improve blood glucose control in diet-induced obese mice with pre-existing metabolic syndrome. We find that specifically reducing dietary BCAAs rapidly reverses diet-induced obesity and improves glucoregulatory control in diet-induced obese mice. Most dramatically, mice eating an otherwise unhealthy high-calorie, high-sugar Western diet with reduced levels of BCAAs lost weight and fat mass rapidly until regaining a normal weight. Importantly, this normalization of weight was mediated not by caloric restriction or increased activity, but by increased energy expenditure, and was accompanied by a transient induction of the energy balance regulating hormone FGF21 (fibroblast growth factor 21). Consumption of a Western diet reduced in BCAAs was also accompanied by a dramatic improvement in glucose tolerance and insulin resistance. Our results link dietary BCAAs with the regulation of metabolic health and energy balance in obese animals, and suggest that specifically reducing dietary BCAAs may represent a highly translatable option for the treatment of obesity and insulin resistance.

Objective. To identify and characterize a fully human antibody directed against B lymphocyte stimulator (BLyS), a tumor necrosis factor-related cytokine that plays a critical role in the regulation of B cell maturation and development. Elevated levels of BLyS have been implicated in the pathogenesis of autoimmune diseases.Methods. A human phage display library was screened for antibodies against human BLyS. A human monoclonal antibody, LymphoStat-B, specific for human BLyS was obtained from the library screening and subsequent affinity optimization mutagenesis. The antibody was tested for inhibition of human BLyS in vitro and in an in vivo murine model. Additionally, the consequences of BLyS inhibition were tested in vivo by administration of LymphoStat-B to cynomolgus monkeys.Results. LymphoStat-B bound with high affinity to human BLyS and inhibited the binding of BLyS to its

Previous studies of the interactions of NO with human hemoglobin have implied the predominance of reaction channels that alternatively eliminate NO by converting it to nitrate, or tightly complex it on the ␣ subunit ferrous hemes. Both channels could effectively quench NO bioactivity. More recent work has raised the idea that NO groups can efficiently transfer from the hemes to cysteine thiols within the ␤ subunit (cys␤-93) to form bioactive nitrosothiols. The regulation of NO function, through its chemical position in the hemoglobin, is supported by response to oxygen and to redox agents that modulate the molecular and electronic structure of the protein. In this article, we focus on reactions in which Fe(III) hemes could provide the oxidative requirements of this NO-group transfer chemistry. We report a detailed investigation of the reductive nitrosylation of human met-Hb, in which we demonstrate the production of S-nitroso (SNO)-Hb through a heme-Fe(III)NO intermediate. The production of SNO-Hb is strongly favored (over nitrite) when NO is gradually introduced in limited total quantities; in this situation, moreover, heme nitrosylation occurs primarily within the ␤ subunits of the hemoglobin tetramer. SNO-Hb can similarly be produced when Fe(II)NO hemes are subjected to mild oxidation. The reaction of deoxygenated hemoglobin with limited quantities of nitrite leads to the production of ␤ subunit Fe(II)NO hemes, with SNO-Hb produced on subsequent oxygenation. The common theme of these reactions is the effective coupling of heme-iron and NO redox chemistries. Collectively, they establish a connectivity between hemes and thiols in Hb, through which NO is readily dislodged from storage on the heme to form bioactive SNO-Hb.T he transfer of NO groups within human hemoglobin from hemes to cys(␤-93) thiols to form a bioactive nitrosothiol represents a novel intramolecular biochemistry that is both of fundamental interest and has considerable implications for understanding the physiological effects of NO in the regulation of vascular tension and blood f low. A requirement of this transfer, common to biological S-nitrosylation (1), is the redox activation of the NO group (2). In this article, we report the results of experiments that probe the idea that heme-iron valence change can support the oxidative requirements of NO-group transfer and thus efficiently lead to the production of S-nitroso (SNO)-Hb. As a model of the reaction between ferric hemes and NO, the reductive nitrosylation of human methemoglobin is examined in detail. Product distribution assays reveal that SNO-Hb is formed as a nitrosation product, which, moreover, is substantially favored over NO 2 Ϫ when NO is gradually introduced as a limiting reagent; furthermore, in this situation, heme nitrosylation occurs primarily within the ␤ subunits of the Hb tetramer. A kinetic analysis unambiguously reveals the intermediacy of heme-Fe(III)NO in this reaction. To extend our observations to reactions that could mimic this chemistry but do not require an accumula...

Protein restricted (PR) diets promote health and longevity in many species. While the precise components of a PR diet that mediate the beneficial effects to longevity have not been defined, we recently showed that many metabolic effects of PR can be attributed to reduced dietary levels of the branched-chain amino acids (BCAAs) leucine, isoleucine, and valine. Here, we demonstrate that restricting dietary BCAAs increases the survival of two different progeroid mouse models, delays frailty and promotes the metabolic health of wild-type C57BL/6J mice when started in midlife, and leads to a 30% increase in lifespan and a reduction in frailty in male, but not female, wild-type mice when fed lifelong. Our results demonstrate that restricting dietary BCAAs can increase healthspan and longevity in mice, and suggest that reducing dietary BCAAs may hold potential as a translatable intervention to promote healthy aging.

Background and Aims The breadfruit genus (Artocarpus, Moraceae) includes valuable underutilized fruit tree crops with a centre of diversity in Southeast Asia. It belongs to the monophyletic tribe Artocarpeae, whose only other members include two small neotropical genera. This study aimed to reconstruct the phylogeny, estimate divergence dates and infer ancestral ranges of Artocarpeae, especially Artocarpus, to better understand spatial and temporal evolutionary relationships and dispersal patterns in a geologically complex region.Methods To investigate the phylogeny and biogeography of Artocarpeae, this study used Bayesian and maximum likelihood approaches to analyze DNA sequences from six plastid and two nuclear regions from 75% of Artocarpus species, both neotropical Artocarpeae genera, and members of all other Moraceae tribes. Six fossil-based calibrations within the Moraceae family were used to infer divergence times. Ancestral areas and estimated dispersal events were also inferred.Key Results Artocarpeae, Artocarpus and four monophyletic Artocarpus subgenera were well supported. A late Cretaceous origin of the Artocarpeae tribe in the Americas is inferred, followed by Eocene radiation of Artocarpus in Asia, with the greatest diversification occurring during the Miocene. Borneo is reconstructed as the ancestral range of Artocarpus, with dozens of independent in situ diversification events inferred there, as well as dispersal events to other regions of Southeast Asia. Dispersal pathways of Artocarpus and its ancestors are proposed.Conclusions Borneo was central in the diversification of the genus Artocarpus and probably served as the centre from which species dispersed and diversified in several directions. The greatest amount of diversification is inferred to have occurred during the Miocene, when sea levels fluctuated and land connections frequently existed between Borneo, mainland Asia, Sumatra and Java. Many species found in these areas have extant overlapping ranges, suggesting that sympatric speciation may have occurred. By contrast, Artocarpus diversity east of Borneo (where many of the islands have no historical connections to the landmasses of the Sunda and Sahul shelves) is unique and probably the product of over water long-distance dispersal events and subsequent diversification in allopatry. This work represents the most comprehensive Artocarpus phylogeny and biogeography study to date and supports Borneo as an evolutionary biodiversity hotspot.

Summary Ecological restoration is critical for mitigating habitat loss and providing ecosystem services. However, restorations often have lower diversity than remnant, reference sites. Phylogenetic diversity is an important component of biodiversity and ecosystem function that has only recently been used to evaluate restoration outcomes. To move towards prediction in the restoration of biodiversity, it is necessary to understand how phylogenetic diversity of restorations compares with that of reference sites, and where deficits are found, to evaluate factors constraining phylogenetic diversity. We quantified plant taxonomic and phylogenetic diversity in eastern tallgrass prairie, one of the most endangered ecosystems on earth. We measured diversity at large (site) and small (plot) scales in 19 restored prairies and compared patterns with those from 41 remnant prairies. To evaluate how environmental conditions and management actions influence outcomes, we tested the effects of soil properties and seed mix composition on diversity of restorations. Restored prairies were less phylogenetically diverse than remnants at both spatial scales. On the other hand, the total species richness of remnant and restored prairies did not significantly differ, but remnants had higher native richness. Restored communities were taxonomically and phylogenetically distinct from remnants. Soil properties (moisture and pH) influenced phylogenetic diversity and composition. There were positive relationships between the taxonomic and phylogenetic diversity of seed mixes and resulting diversity of planted assemblages (excluding volunteer species). Species in seed mixes were more closely related than expected by chance, and several clades found in remnant prairies were missing from seed mixes. Synthesis and applications. Restored tallgrass prairies had lower phylogenetic diversity than remnant prairies, which may contribute to the widely observed phenomenon of restorations not being functionally equivalent to reference sites. It is encouraging for restoration efforts that seed mix phylogenetic diversity predicted phylogenetic diversity of planted assemblages. This indicates that designing phylogenetically diverse seed mixes for restoration is beneficial. In addition, clades found in reference sites that are missing from restoration seed mixes could be added to new or existing restorations to reduce gaps in phylogenetic diversity. Further work on the effects of management on phylogenetic diversity is needed to advance restoration of biodiversity.

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Objective-We first showed that absence of p53 accelerates atherosclerosis development in apoE-deficient mice. In this study, we investigated how macrophage-specific loss of p53 function might modulate atherosclerosis development in LDL receptor-deficient mice, a model for familial hypercholesterolemia. Methods and Results-We transferred bone marrow cells isolated from p53ϩ/ϩ and p53 Ϫ/Ϫ mice to lethally irradiated LDL receptor Ϫ/Ϫ mice and evaluated the aortic atherosclerotic lesion areas in the recipients at different times afterward. At 15 weeks and again at 20 weeks, we found larger aortic lesion size in mice receiving p53Ϫ/Ϫ cells compared with those that received p53 ϩ/ϩ cells. By measuring the rate of bromodeoxyuridine incorporation, we found that the absence of p53 in macrophages stimulates cellular proliferation. In contrast, the rate of apoptosis in the atheromatous lesion was similar in the two groups of mice. Furthermore, we found that the absence of macrophage-specific p53 expression was associated with vulnerable-appearing lesions marked by increased tissue necrosis and reduced collagen deposition. Conclusions-p53 plays a crucial role in atherosclerosis lesion development and remodeling, and macrophage-specific p53 deficiency stimulates cellular proliferation leading to a