Short-lived alpha-helical intermediates in the folding of beta-sheet proteins.

Several lines of evidence point strongly toward the importance of highly alpha-helical intermediates in the folding of all globular proteins, regardless of their native structure. However, experimental refolding studies demonstrate no observable alpha-helical intermediate during refolding of some beta-sheet proteins and have dampened enthusiasm for this model of protein folding. In this study, beta-sheet proteins were hypothesized to have potential to form amphiphilic helices at a period of <3.6 residues/turn that matches or exceeds the potential at 3.6 residues/turn. Hypothetically, such potential is the basis for an effective and unidirectional mechanism by which highly alpha-helical intermediates might be rapidly disassembled during folding and potentially accounts for the difficulty in detecting highly alpha-helical intermediates during the folding of some proteins. The presence of this potential was confirmed, indicating that a model entailing ubiquitous formation of alpha-helical intermediates during the folding of globular proteins predicts previously unrecognized features of primary structure. Further, the folding of fatty acid binding protein, a predominantly beta-sheet protein that exhibits no apparent highly alpha-helical intermediate during folding, was dramatically accelerated by 2,2,2-trifluoroethanol, a solvent that stabilizes alpha-helical structure. This observation suggests that formation of an alpha-helix can be a rate-limiting step during folding of a predominantly beta-sheet protein and further supports the role of highly alpha-helical intermediates in the folding of all globular proteins.

Developmental origins of precocial forelimbs in marsupial neonates.

Marsupial mammals are born in an embryonic state, as compared with their eutherian counterparts, yet certain features are accelerated. The most conspicuous of these features are the precocial forelimbs, which the newborns use to climb unaided from the opening of the birth canal to the teat. The developmental mechanisms that produce this acceleration are unknown. Here we show that heterochronic and heterotopic changes early in limb development contribute to forelimb acceleration. Using Tbx5 and Tbx4 as fore- and hindlimb field markers, respectively, we have found that, compared with mouse, both limb fields arise notably early during opossum development. Patterning of the forelimb buds is also accelerated, as Shh expression appears early relative to the outgrowth of the bud itself. In addition, the forelimb fields and forelimb myocyte allocation are increased in size and number, respectively, and migration of the spinal nerves into the forelimb bud has been modified. This shift in the extent of the forelimb field is accompanied by shifts in Hox gene expression along the anterior-posterior axis. Furthermore, we found that both fore- and hindlimb fields arise gradually during gastrulation and extension of the embryonic axis, in contrast to the appearance of the limb fields in their entirety in all other known cases. Our results show a surprising evolutionary flexibility in the early limb development program of amniotes and rule out the induction of the limb fields by mature structures such as the somites or mesonephros.

Morphological and genomic characterization of Filobasidiella depauperata: a homothallic sibling species of the pathogenic cryptococcus species complex.

The fungal species Cryptococcus neoformans and Cryptococcus gattii cause respiratory and neurological disease in animals and humans following inhalation of basidiospores or desiccated yeast cells from the environment. Sexual reproduction in C. neoformans and C. gattii is controlled by a bipolar system in which a single mating type locus (MAT) specifies compatibility. These two species are dimorphic, growing as yeast in the asexual stage, and producing hyphae, basidia, and basidiospores during the sexual stage. In contrast, Filobasidiella depauperata, one of the closest related species, grows exclusively as hyphae and it is found in association with decaying insects. Examination of two available strains of F. depauperata showed that the life cycle of this fungal species shares features associated with the unisexual or same-sex mating cycle in C. neoformans. Therefore, F. depauperata may represent a homothallic and possibly an obligately sexual fungal species. RAPD genotyping of 39 randomly isolated progeny from isolate CBS7855 revealed a new genotype pattern in one of the isolated basidiospores progeny, therefore suggesting that the homothallic cycle in F. depauperata could lead to the emergence of new genotypes. Phylogenetic analyses of genes linked to MAT in C. neoformans indicated that two of these genes in F. depauperata, MYO2 and STE20, appear to form a monophyletic clade with the MATa alleles of C. neoformans and C. gattii, and thus these genes may have been recruited to the MAT locus before F. depauperata diverged. Furthermore, the ancestral MATa locus may have undergone accelerated evolution prior to the divergence of the pathogenic Cryptococcus species since several of the genes linked to the MATa locus appear to have a higher number of changes and substitutions than their MATalpha counterparts. Synteny analyses between C. neoformans and F. depauperata showed that genomic regions on other chromosomes displayed conserved gene order. In contrast, the genes linked to the MAT locus of C. neoformans showed a higher number of chromosomal translocations in the genome of F. depauperata. We therefore propose that chromosomal rearrangements appear to be a major force driving speciation and sexual divergence in these closely related pathogenic and saprobic species.

Growth hormone mitigates against lethal irradiation and enhances hematologic and immune recovery in mice and nonhuman primates.

Medications that can mitigate against radiation injury are limited. In this study, we investigated the ability of recombinant human growth hormone (rhGH) to mitigate against radiation injury in mice and nonhuman primates. BALB/c mice were irradiated with 7.5 Gy and treated post-irradiation with rhGH intravenously at a once daily dose of 20 microg/dose for 35 days. rhGH protected 17 out of 28 mice (60.7%) from lethal irradiation while only 3 out of 28 mice (10.7%) survived in the saline control group. A shorter course of 5 days of rhGH post-irradiation produced similar results. Compared with the saline control group, treatment with rhGH on irradiated BALB/c mice significantly accelerated overall hematopoietic recovery. Specifically, the recovery of total white cells, CD4 and CD8 T cell subsets, B cells, NK cells and especially platelets post radiation exposure were significantly accelerated in the rhGH-treated mice. Moreover, treatment with rhGH increased the frequency of hematopoietic stem/progenitor cells as measured by flow cytometry and colony forming unit assays in bone marrow harvested at day 14 after irradiation, suggesting the effects of rhGH are at the hematopoietic stem/progenitor level. rhGH mediated the hematopoietic effects primarily through their niches. Similar data with rhGH were also observed following 2 Gy sublethal irradiation of nonhuman primates. Our data demonstrate that rhGH promotes hematopoietic engraftment and immune recovery post the exposure of ionizing radiation and mitigates against the mortality from lethal irradiation even when administered after exposure.

The evolutionary origin of Indian Ocean tortoises (Dipsochelys).

Today, the only surviving wild population of giant tortoises in the Indian Ocean occurs on the island of Aldabra. However, giant tortoises once inhabited islands throughout the western Indian Ocean. Madagascar, Africa, and India have all been suggested as possible sources of colonization for these islands. To address the origin of Indian Ocean tortoises (Dipsochelys, formerly Geochelone gigantea), we sequenced the 12S, 16S, and cyt b genes of the mitochondrial DNA. Our phylogenetic analysis shows Dipsochelys to be embedded within the Malagasy lineage, providing evidence that Indian Ocean giant tortoises are derived from a common Malagasy ancestor. This result points to Madagascar as the source of colonization for western Indian Ocean islands by giant tortoises. Tortoises are known to survive long oceanic voyages by floating with ocean currents, and thus, currents flowing northward towards the Aldabra archipelago from the east coast of Madagascar would have provided means for the colonization of western Indian Ocean islands. Additionally, we found an accelerated rate of sequence evolution in the two Malagasy Pyxis species examined. This finding supports previous theories that shorter generation time and smaller body size are related to an increase in mitochondrial DNA substitution rate in vertebrates.

Wee1-regulated apoptosis mediated by the crk adaptor protein in Xenopus egg extracts.

Many of the biochemical reactions of apoptotic cell death, including mitochondrial cytochrome c release and caspase activation, can be reconstituted in cell-free extracts derived from Xenopus eggs. In addition, because caspase activation does not occur until the egg extract has been incubated for several hours on the bench, upstream signaling processes occurring before full apoptosis are rendered accessible to biochemical manipulation. We reported previously that the adaptor protein Crk is required for apoptotic signaling in egg extracts (Evans, E.K., W. Lu, S.L. Strum, B.J. Mayer, and S. Kornbluth. 1997. EMBO (Eur. Mol. Biol. Organ.) J. 16:230-241). Moreover, we demonstrated that removal of Crk Src homology (SH)2 or SH3 interactors from the extracts prevented apoptosis. We now report the finding that the relevant Crk SH2-interacting protein, important for apoptotic signaling in the extract, is the well-known cell cycle regulator, Wee1. We have demonstrated a specific interaction between tyrosine-phosphorylated Wee1 and the Crk SH2 domain and have shown that recombinant Wee1 can restore apoptosis to an extract depleted of SH2 interactors. Moreover, exogenous Wee1 accelerated apoptosis in egg extracts, and this acceleration was largely dependent on the presence of endogenous Crk protein. As other Cdk inhibitors, such as roscovitine and Myt1, did not act like Wee1 to accelerate apoptosis, we propose that Wee1-Crk complexes signal in a novel apoptotic pathway, which may be unrelated to Wee1's role as a cell cycle regulator.

Inhibition of the anaphase-promoting complex by the Xnf7 ubiquitin ligase.

Degradation of specific protein substrates by the anaphase-promoting complex/cyclosome (APC) is critical for mitotic exit. We have identified the protein Xenopus nuclear factor 7 (Xnf7) as a novel APC inhibitor able to regulate the timing of exit from mitosis. Immunodepletion of Xnf7 from Xenopus laevis egg extracts accelerated the degradation of APC substrates cyclin B1, cyclin B2, and securin upon release from cytostatic factor arrest, whereas excess Xnf7 inhibited APC activity. Interestingly, Xnf7 exhibited intrinsic ubiquitin ligase activity, and this activity was required for APC inhibition. Unlike other reported APC inhibitors, Xnf7 did not associate with Cdc20, but rather bound directly to core subunits of the APC. Furthermore, Xnf7 was required for spindle assembly checkpoint function in egg extracts. These data suggest that Xnf7 is an APC inhibitor able to link spindle status to the APC through direct association with APC core components.

Genes, race, and the ethics of belief.

A Troublesome Inheritance, by Nicholas Wade, should be read by anyone interested in race and recent human evolution. Wade deserves credit for challenging the popular dog-ma that biological differences between groups either don't exist or cannot ex-plain the relative success of different groups at different tasks. Wade's work should be read alongside another re-cent book, The 10,000 Year Explosion: How Civilization Accelerated Human Evolution, by Gregory Cochran and Henry Harpending. Together, these books represent a ma-jor turning point in the public debate about the speed with which relatively isolated groups can evolve: both books suggest that small genetic differences between members of different groups can have large impacts on their abilities and propensities, which in turn affect the outcomes of the societies in which they live.

Is chronic asthma associated with shorter leukocyte telomere length at midlife?

RATIONALE: Asthma is prospectively associated with age-related chronic diseases and mortality, suggesting the hypothesis that asthma may relate to a general, multisystem phenotype of accelerated aging. OBJECTIVES: To test whether chronic asthma is associated with a proposed biomarker of accelerated aging, leukocyte telomere length. METHODS: Asthma was ascertained prospectively in the Dunedin Multidisciplinary Health and Development Study cohort (n = 1,037) at nine in-person assessments spanning ages 9-38 years. Leukocyte telomere length was measured at ages 26 and 38 years. Asthma was classified as life-course-persistent, childhood-onset not meeting criteria for persistence, and adolescent/adult-onset. We tested associations between asthma and leukocyte telomere length using regression models. We tested for confounding of asthma-leukocyte telomere length associations using covariate adjustment. We tested serum C-reactive protein and white blood cell counts as potential mediators of asthma-leukocyte telomere length associations. MEASUREMENTS AND MAIN RESULTS: Study members with life-course-persistent asthma had shorter leukocyte telomere length as compared with sex- and age-matched peers with no reported asthma. In contrast, leukocyte telomere length in study members with childhood-onset and adolescent/adult-onset asthma was not different from leukocyte telomere length in peers with no reported asthma. Adjustment for life histories of obesity and smoking did not change results. Study members with life-course-persistent asthma had elevated blood eosinophil counts. Blood eosinophil count mediated 29% of the life-course-persistent asthma-leukocyte telomere length association. CONCLUSIONS: Life-course-persistent asthma is related to a proposed biomarker of accelerated aging, possibly via systemic eosinophilic inflammation. Life histories of asthma can inform studies of aging.

Human-chimpanzee differences in a FZD8 enhancer alter cell-cycle dynamics in the developing neocortex.

The human neocortex differs from that of other great apes in several notable regards, including altered cell cycle, prolonged corticogenesis, and increased size [1-5]. Although these evolutionary changes most likely contributed to the origin of distinctively human cognitive faculties, their genetic basis remains almost entirely unknown. Highly conserved non-coding regions showing rapid sequence changes along the human lineage are candidate loci for the development and evolution of uniquely human traits. Several studies have identified human-accelerated enhancers [6-14], but none have linked an expression difference to a specific organismal trait. Here we report the discovery of a human-accelerated regulatory enhancer (HARE5) of FZD8, a receptor of the Wnt pathway implicated in brain development and size [15, 16]. Using transgenic mice, we demonstrate dramatic differences in human and chimpanzee HARE5 activity, with human HARE5 driving early and robust expression at the onset of corticogenesis. Similar to HARE5 activity, FZD8 is expressed in neural progenitors of the developing neocortex [17-19]. Chromosome conformation capture assays reveal that HARE5 physically and specifically contacts the core Fzd8 promoter in the mouse embryonic neocortex. To assess the phenotypic consequences of HARE5 activity, we generated transgenic mice in which Fzd8 expression is under control of orthologous enhancers (Pt-HARE5::Fzd8 and Hs-HARE5::Fzd8). In comparison to Pt-HARE5::Fzd8, Hs-HARE5::Fzd8 mice showed marked acceleration of neural progenitor cell cycle and increased brain size. Changes in HARE5 function unique to humans thus alter the cell-cycle dynamics of a critical population of stem cells during corticogenesis and may underlie some distinctive anatomical features of the human brain.

Clinical Research Environment in India: Challenges and Proposed Solutions.

India has compelling need and keen aspirations for indigenous clinical research. Notwithstanding this need and previously reported growth the expected expansion of Indian clinical research has not materialized. We reviewed the scientific literature, lay press reports, and ClinicalTrials.gov data for information and commentary on projections, progress, and impediments associated with clinical trials in India. We also propose targeted solutions to identified challenges. The Indian clinical trial sector grew by (+) 20.3% CAGR (compound annual growth rate) between 2005 and 2010 and contracted by (-) 14.6% CAGR between 2010 and 2013. Phase-1 trials grew by (+) 43.5% CAGR from 2005-2013, phase-2 trials grew by (+) 19.8% CAGR from 2005-2009 and contracted by (-) 12.6% CAGR from 2009-2013, and phase-3 trials grew by (+) 13.0% CAGR from 2005-2010 and contracted by (-) 28.8% CAGR from 2010-2013. This was associated with a slowing of the regulatory approval process, increased media coverage and activist engagement, and accelerated development of regulatory guidelines and recuperative initiatives. We propose the following as potential targets for restorative interventions: Regulatory overhaul (leadership and enforcement of regulations, resolution of ambiguity in regulations, staffing, training, guidelines, and ethical principles [e.g., compensation]).Education and training of research professionals, clinicians, and regulators.Public awareness and empowerment. After a peak in 2009-2010, the clinical research sector in India appears to be experiencing a contraction. There are indications of challenges in regulatory enforcement of guidelines; training of clinical research professionals; and awareness, participation, partnership, and the general image amongst the non-professional media and public. Preventative and corrective principles and interventions are outlined with the goal of realizing the clinical research potential in India.

Huntingtin is required for normal excitatory synapse development in cortical and striatal circuits.

Huntington's disease (HD) is a neurodegenerative disease caused by the expansion of a poly-glutamine (poly-Q) stretch in the huntingtin (Htt) protein. Gain-of-function effects of mutant Htt have been extensively investigated as the major driver of neurodegeneration in HD. However, loss-of-function effects of poly-Q mutations recently emerged as potential drivers of disease pathophysiology. Early synaptic problems in the excitatory cortical and striatal connections have been reported in HD, but the role of Htt protein in synaptic connectivity was unknown. Therefore, we investigated the role of Htt in synaptic connectivity in vivo by conditionally silencing Htt in the developing mouse cortex. When cortical Htt function was silenced, cortical and striatal excitatory synapses formed and matured at an accelerated pace through postnatal day 21 (P21). This exuberant synaptic connectivity was lost over time in the cortex, resulting in the deterioration of synapses by 5 weeks. Synaptic decline in the cortex was accompanied with layer- and region-specific reactive gliosis without cell loss. To determine whether the disease-causing poly-Q mutation in Htt affects synapse development, we next investigated the synaptic connectivity in a full-length knock-in mouse model of HD, the zQ175 mouse. Similar to the cortical conditional knock-outs, we found excessive excitatory synapse formation and maturation in the cortices of P21 zQ175, which was lost by 5 weeks. Together, our findings reveal that cortical Htt is required for the correct establishment of cortical and striatal excitatory circuits, and this function of Htt is lost when the mutant Htt is present.

Temporal Stability of Molecular Diversity Measures in Natural Populations of Drosophila pseudoobscura and Drosophila persimilis

Many molecular ecological and evolutionary studies sample wild populations at a single point in time, failing to consider that data they collect represents genetic variation from a potentially unrepresentative snapshot in time. Variation across time in genetic parameters may occur quickly in species that produce multiple generations of offspring per year. However, many studies of rapid contemporary microevolution examine phenotypic trait divergence as opposed to molecular evolutionary divergence. Here, we compare genetic diversity in wild caught populations of Drosophila persimilis and D. pseudoobscura collected 16 years apart at the same time of year and same site at four X-linked and two mitochondrial loci to assess genetic stability. We found no major changes in nucleotide diversity in either species, but we observed a drastic shift in Tajima’s D between D. pseudoobscura timepoints at one locus associated with the increased abundance of a set of related haplotypes. Our data also suggests that D. persimilis may have recently accelerated its demographic expansion. While the changes we observed were modest, this study reinforces the importance of considering potential temporal variation in genetic parameters within single populations over short evolutionary timescales.

Global value chains in a post-Washington Consensus world

Contemporary globalization has been marked by significant shifts in the organization and governance of global industries. In the 1970s and 1980s, one such shift was characterized by the emergence of buyer-driven and producer-driven commodity chains. In the early 2000s, a more differentiated typology of governance structures was introduced, which focused on new types of coordination in global value chains (GVCs). Today the organization of the global economy is entering another phase, with transformations that are reshaping the governance structures of both GVCs and global capitalism at various levels: (1) the end of the Washington Consensus and the rise of contending centers of economic and political power; (2) a combination of geographic consolidation and value chain concentration in the global supply base, which, in some cases, is shifting bargaining power from lead firms in GVCs to large suppliers in developing economies; (3) new patterns of strategic coordination among value chain actors; (4) a shift in the end markets of many GVCs accelerated by the economic crisis of 2008-09, which is redefining regional geographies of investment and trade; and (5) a diffusion of the GVC approach to major international donor agencies, which is prompting a reformulation of established development paradigms. © 2013 © 2013 Taylor & Francis.

Oldest known cranium of a juvenile New World monkey (Early Miocene, Patagonia, Argentina): implications for the taxonomy and the molar eruption pattern of early platyrrhines.

A juvenile cranium of Homunculus patagonicus Ameghino, 1891a from the late Early Miocene of Santa Cruz Province (Argentina) provides the first evidence of developing cranial anatomy for any fossil platyrrhine. The specimen preserves the rostral part of the cranium with deciduous and permanent alveoli and teeth. The dental eruption sequence in the new specimen and a reassessment of eruption patterns in living and fossil platyrrhines suggest that the ancestral platyrrhine pattern of tooth replacement was for the permanent incisors to erupt before M(1), not an accelerated molar eruption (before the incisors) as recently proposed. Two genera and species of Santacrucian monkeys are now generally recognized: H. patagonicus Ameghino, 1891a and Killikaike blakei Tejedor et al., 2006. Taxonomic allocation of Santacrucian monkeys to these species encounters two obstacles: 1) the (now lost) holotype and a recently proposed neotype of H. patagonicus are mandibles from different localities and different geologic members of the Santa Cruz Formation, separated by approximately 0.7 million years, whereas the holotype of K. blakei is a rostral part of a cranium without a mandible; 2) no Santacrucian monkey with associated cranium and mandible has ever been found. Bearing in mind these uncertainties, our examination of the new specimen as well as other cranial specimens of Santacrucian monkeys establishes the overall dental and cranial similarity between the holotype of Killikaike blakei, adult cranial material previously referred to H. patagonicus, and the new juvenile specimen. This leads us to conclude that Killikaike blakei is a junior subjective synonym of H. patagonicus.