Effect of population size on the estimation of QTL: a test using resistance to barley stripe rust

The limited population sizes used in many quantitative trait locus (QTL) detection experiments can lead to underestimation of QTL number, overestimation of QTL effects, and failure to quantify QTL interactions. We used the barley/barley stripe rust pathosystem to evaluate the effect of population size on the estimation of QTL parameters. We generated a large (n=409) population of doubled haploid lines derived from the cross of two inbred lines, BCD47 and Baronesse. This population was evaluated for barley stripe rust severity in the Toluca Valley, Mexico, and in Washington State, USA, under field conditions. BCD47 was the principal donor of resistance QTL alleles, but the susceptible parent also contributed some resistance alleles. The major QTL, located on the long arm of chromosome 4H, close to the Mlo gene, accounted for up to 34% of the phenotypic variance. Subpopulations of different sizes were generated using three methods—resampling, selective genotyping, and selective phenotyping—to evaluate the effect of population size on the estimation of QTL parameters. In all cases, the number of QTL detected increased with population size. QTL with large effects were detected even in small populations, but QTL with small effects were detected only by increasing population size. Selective genotyping and/or selective phenotyping approaches could be effective strategies for reducing the costs associated with conducting QTL analysis in large populations. The method of choice will depend on the relative costs of genotyping versus phenotyping. Electronic Supplementary Material Supplementary material is available for this article at     

Gene expression changes reflect clinical response in a placebo-controlled randomized trial of abatacept in patients with diffuse cutaneous systemic sclerosis

IntroductionSystemic sclerosis is an autoimmune disease characterized by inflammation and fibrosis of the skin and internal organs. We sought to assess the clinical and molecular effects associated with response to intravenous abatacept in patients with diffuse cutaneous systemic.MethodsAdult diffuse cutaneous systemic sclerosis patients were randomized in a 2:1 double-blinded fashion to receive abatacept or placebo over 24 weeks. Primary outcomes were safety and the change in modified Rodnan Skin Score (mRSS) at week 24 compared with baseline. Improvers were defined as patients with a decrease in mRSS of ≥30 % post-treatment compared to baseline. Skin biopsies were obtained for differential gene expression and pathway enrichment analyses and intrinsic gene expression subset assignment.ResultsTen subjects were randomized to abatacept (n = 7) or placebo (n = 3). Disease duration from first non-Raynaud’s symptom was significantly longer (8.8 ± 3.8 years vs. 2.4 ± 1.6 years, p = 0.004) and median mRSS was higher (30 vs. 22, p = 0.05) in the placebo compared to abatacept group. Adverse events were similar in the two groups. Five out of seven patients (71 %) randomized to abatacept and one out of three patients (33 %) randomized to placebo experienced ≥30 % improvement in skin score. Subjects receiving abatacept showed a trend toward improvement in mRSS at week 24 (−8.6 ± 7.5, p = 0.0625) while those in the placebo group did not (−2.3 ± 15, p = 0.75). After adjusting for disease duration, mRSS significantly improved in the abatacept compared with the placebo group (abatacept vs. placebo mRSS decrease estimate −9.8, 95 % confidence interval −16.7 to −3.0, p = 0.0114). In the abatacept group, the patients in the inflammatory intrinsic subset showed a trend toward greater improvement in skin score at 24 weeks compared with the patients in the normal-like intrinsic subset (−13.5 ± 3.1 vs. −4.5 ± 6.4, p = 0.067). Abatacept resulted in decreased CD28 co-stimulatory gene expression in improvers consistent with its mechanism of action. Improvers mapped to the inflammatory intrinsic subset and showed decreased gene expression in inflammatory pathways, while non-improver and placebos showed stable or reverse gene expression over 24 weeks.ConclusionsClinical improvement following abatacept therapy was associated with modulation of inflammatory pathways in skin.

Trial registration

ClinicalTrials.gov {"type":"clinical-trial","attrs":{"text":"NCT00442611","term_id":"NCT00442611"}}NCT00442611. Registered 1 March 2007.

Electronic supplementary material

The online version of this article (doi:10.1186/s13075-015-0669-3) contains supplementary material, which is available to authorized users.     

Using the allergic immune system to target cancer: activity of IgE antibodies specific for human CD20 and MUC1

Monoclonal antibodies are widely used in the treatment of many B cell lymphomas and certain solid tumors. All currently approved therapeutic monoclonal antibodies are of the immunoglobulin G (IgG) isotype. We hypothesized that tumor-specific monoclonal antibodies of the IgE isotype may serve as effective cancer therapeutics. To test this hypothesis, we produced mouse?Chuman chimeric IgE antibodies specific for the human B cell antigen CD20 and the epithelial antigen MUC1. We demonstrate here that anti-hCD20 IgE antibodies have in vitro cytotoxic activity when used with purified allergic effector cells derived from umbilical cord blood. At an effector-tumor ratio of 2:1, mast cells and tumor-specific IgE induced a 2.5-fold increase in tumor cell death, as compared to control IgE. Similar results were observed when eosinophils were used as effector cells. In an in vivo murine model of breast carcinoma, administration of anti-hMUC1 IgE reduced the growth of MUC1⁺ tumors by 25?C30?% in hFc??RI transgenic mice. In contrast, local production of IgE and cytokines chemotactic for macrophages, eosinophils and mast cells led to complete tumor eradication. These results suggest that allergic effector cells activated by IgE and cell surface antigens have the capacity to induce tumor cell death in vitro and in vivo. The use of chimeric antibodies and hFc??RI transgenic mice will greatly enhance investigations in the nascent field of allergo-oncology.     

SMN-mediated assembly of RNPs: a complex story

Although many RNA-protein complexes or ribonucleoproteins (RNPs) assemble spontaneously in vitro, little is known about how they form in the environment of a living cell. Insight into RNP assembly has come unexpectedly from functional analyses of the survival motor neuron (SMN) protein, a gene product that is affected in the neuromuscular disease spinal muscular atrophy. These studies show that the assembly of spliceosomal U-rich small nuclear RNPs in vivo depends on the activity of two large protein complexes, one of which contains the SMN protein. These complexes might also facilitate the assembly of other cellular RNPs.     

Protein and peptide array analysis of autoimmune disease

Molecular cloning, sequencing of the human genome, and other major advances in biomedical research have contributed substantially to our understanding of autoimmune disease. Nevertheless, to date, such advances have failed to reveal the etiology of or yield curative therapies for autoimmune disease. New approaches are needed. Proteomics, the large-scale study of expression and function of proteins that compose our tissues and mediate disease, represents a powerful and promising strategy. We developed protein and peptide arrays to profile autoantibody responses in autoimmune disease. Protein and peptide array analysis of autoimmune samples is revealing human and pathogen proteins involved in initiation and perpetuation of autoimmunity. Proteomic determination of autoantibody profiles can be utilized for diagnosis, prognostication, and guiding tolerizing therapy for autoimmune disease.     

To manage or not? Successful native tree seedling restoration despite a dense,invasive shrub,Berberis thunbergii

Plant Ecology - Invasive plant species suppress native trees through a variety of mechanisms. A non-native shrub, Berberis thunbergii, has been shown to depress native tree seedling densities in...     

Hybrid maize breeding with doubled haploids: I. One-stage versus two-stage selection for testcross performance

Optimum allocation of resources is of fundamental importance for the efficiency of breeding programs. The objectives of our study were to (1) determine the optimum allocation for the number of lines and test locations in hybrid maize breeding with doubled haploids (DHs) regarding two optimization criteria, the selection gain ΔG _k and the probability P _k of identifying superior genotypes, (2) compare both optimization criteria including their standard deviations (SDs), and (3) investigate the influence of production costs of DHs on the optimum allocation. For different budgets, number of finally selected lines, ratios of variance components, and production costs of DHs, the optimum allocation of test resources under one- and two-stage selection for testcross performance with a given tester was determined by using Monte Carlo simulations. In one-stage selection, lines are tested in field trials in a single year. In two-stage selection, optimum allocation of resources involves evaluation of (1) a large number of lines in a small number of test locations in the first year and (2) a small number of the selected superior lines in a large number of test locations in the second year, thereby maximizing both optimization criteria. Furthermore, to have a realistic chance of identifying a superior genotype, the probability P _k of identifying superior genotypes should be greater than 75%. For budgets between 200 and 5,000 field plot equivalents, P _k > 75% was reached only for genotypes belonging to the best 5% of the population. As the optimum allocation for P _k (5%) was similar to that for ΔG _k , the choice of the optimization criterion was not crucial. The production costs of DHs had only a minor effect on the optimum number of locations and on values of the optimization criteria. C. Friedrich H. Longin and H. Friedrich Utz contributed equally to this work.     

Identification of residues in the human guanylate-binding protein 1 critical for nucleotide binding and cooperative GTP hydrolysis

The guanylate-binding proteins (GBPs) form a group of interferon-gamma inducible GTP-binding proteins which belong to the family of dynamin-related proteins. Like other members of this family, human guanylate-binding protein 1 (hGBP1) shows nucleotide-dependent oligomerisation that stimulates the GTPase activity of the protein. A unique feature of the GBPs is their ability to hydrolyse GTP to GDP and GMP. In order to elucidate the relationship between these findings, we designed point mutants in the phosphate-binding loop (P-loop) as well as in the switch I and switch II regions of the protein based on the crystal structure of hGBP1. These mutant proteins were analysed for their interaction with guanine nucleotides labeled with a fluorescence dye and for their ability to hydrolyse GTP in a cooperative manner. We identified mutations of amino acid residues that decrease GTPase activity by orders of magnitude a part of which are conserved in GTP-binding proteins. In addition, mutants in the P-loop were characterized that strongly impair binding of nucleotide. In consequence, together with altered GTPase activity and given cellular nucleotide concentrations this results in hGBP1 mutants prevailingly resting in the nucleotide-free (K51A and S52N) or the GTP bound form (R48A), respectively. Using size-exclusion chromatography and analytical ultracentrifugation we addressed the impact on protein oligomerisation. In summary, mutants of hGBP1 were identified and biochemically characterized providing hGBP1 locked in defined states in order to investigate their functional role in future cell biology studies.