Detection of linkage between marker loci and loci affecting quantitative traits in crosses between segregating populations

Summary By making use of pedigree information and information on marker-genotypes of the parent and F-1 individuals crossed to form an F-2 population, it is possible to carry out a linkage analysis between marker loci and loci affecting quantitative traits in a cross between segregating parent populations that are at fixation for alternative alleles at the QTL, but share the same alleles at the marker loci. For two-allele systems, depending on marker allele frequencies in the parent populations, 2–4 times as many F-2 offspring will have to be raised and scored for markers and quantitative traits in order to provide power equivalent to that obtained in a cross between fully inbred lines. Major savings in number of F-2 offspring raised can be achieved by scoring each parent pair for a large number of markers in each chromosomal region and scoring F-1 and F-2 offspring only for those markers for which the parents were homozygous for alternative alleles. For multiple allele systems, particularly when dealing with hypervariable loci, only 10%–20% additional F-2 offspring will have to be raised and scored to provide power equivalent to that obtained in a cross between inbred lines. When a resource population contains novel favorable alleles at quantitative trait loci that are not present (or rare) in a commercial population, analyses of this sort will enable the loci of interest to be identified, mapped and manipulated effectively in breeding programs.Contribution no. 2124-E, 1987 series from The Agricultural Research Organization, The Volcani Center, Bet Dagan, Israel     

Chromosomal localization of ARSB,the gene for human N-acetylgalactosamine-4-sulphatase

Summary A deficiency of N-acetylgalactosamine-4-sulphatase (G4S, gene symbol ARSB), results in the accumulation of undegraded substrate and the lysosomal storage disorder, Maroteaux-Lamy syndrome (mucopolysaccharidosis type VI). In situ hybridization using an ³H-labelled human G4S genomic DNA fragment to human metaphase chromosomes localized ARSB to chromosome 5q13–5q14. This location is consistent with, an refines, previous chromosomal assignments based on the expression of human G4S in somatic cell hybrids.     

Opportunities and challenges for digital morphology

Abstract  

Advances in digital data acquisition, analysis, and storage have revolutionized the work in many biological disciplines such as genomics, molecular phylogenetics, and structural biology, but have not yet found satisfactory acceptance in morphology. Improvements in non-invasive imaging and three-dimensional visualization techniques, however, permit high-throughput analyses also of whole biological specimens, including museum material. These developments pave the way towards a digital era in morphology. Using sea urchins (Echinodermata: Echinoidea), we provide examples illustrating the power of these techniques. However, remote visualization, the creation of a specialized database, and the implementation of standardized, world-wide accepted data deposition practices prior to publication are essential to cope with the foreseeable exponential increase in digital morphological data.     

The Syk Kinase SmTK4 of Schistosoma mansoni Is Involved in the Regulation of Spermatogenesis and Oogenesis

The signal transduction protein SmTK4 from Schistosoma mansoni belongs to the family of Syk kinases. In vertebrates, Syk kinases are known to play specialized roles in signaling pathways in cells of the hematopoietic system. Although Syk kinases were identified in some invertebrates, their role in this group of animals has not yet been elucidated. Since SmTK4 is the first Syk kinase from a parasitic helminth, shown to be predominantly expressed in the testes and ovary of adult worms, we investigated its function. To unravel signaling cascades in which SmTK4 is involved, yeast two-/three-hybrid library screenings were performed with either the tandem SH2-domain, or with the linker region including the tyrosine kinase domain of SmTK4. Besides the Src kinase SmTK3 we identified a new Src kinase (SmTK6) acting upstream of SmTK4 and a MAPK-activating protein, as well as mapmodulin acting downstream. Their identities and colocalization studies pointed to a role of SmTK4 in a signaling cascade regulating the proliferation and/or differentiation of cells in the gonads of schistosomes. To confirm this decisive role we performed biochemical and molecular approaches to knock down SmTK4 combined with a novel protocol for confocal laser scanning microscopy for morphological analyses. Using the Syk kinase-specific inhibitor Piceatannol or by RNAi treatment of adult schistosomes in vitro, corresponding phenotypes were detected in the testes and ovary. In the Xenopus oocyte system it was finally confirmed that Piceatannol suppressed the activity of the catalytic kinase domain of SmTK4. Our findings demonstrate a pivotal role of SmTK4 in gametogenesis, a new function for Syk kinases in eukaryotes.     

The SARS‐unique domain (SUD) of SARS‐CoV and SARS‐CoV‐2 interacts with human Paip1 to enhance viral RNA translation

The ongoing outbreak of severe acute respiratory syndrome (SARS) coronavirus 2 (SARS‐CoV‐2) demonstrates the continuous threat of emerging coronaviruses (CoVs) to public health. SARS‐CoV‐2 and SARS‐CoV share an otherwise non‐conserved part of non‐structural protein 3 (Nsp3), therefore named as “SARS‐unique domain” (SUD). We previously found a yeast‐2‐hybrid screen interaction of the SARS‐CoV SUD with human poly(A)‐binding protein (PABP)‐interacting protein 1 (Paip1), a stimulator of protein translation. Here, we validate SARS‐CoV SUD:Paip1 interaction by size‐exclusion chromatography, split‐yellow fluorescent protein, and co‐immunoprecipitation assays, and confirm such interaction also between the corresponding domain of SARS‐CoV‐2 and Paip1. The three‐dimensional structure of the N‐terminal domain of SARS‐CoV SUD (“macrodomain II”, Mac2) in complex with the middle domain of Paip1, determined by X‐ray crystallography and small‐angle X‐ray scattering, provides insights into the structural determinants of the complex formation. In cellulo, SUD enhances synthesis of viral but not host proteins via binding to Paip1 in pBAC‐SARS‐CoV replicon‐transfected cells. We propose a possible mechanism for stimulation of viral translation by the SUD of SARS‐CoV and SARS‐CoV‐2.     

The C‐terminal regions of YidC from Rhodopirellula baltica and Oceanicaulis alexandrii bind to ribosomes and partially substitute for SRP receptor function in Escherichia coli

The marine Gram‐negative bacteria Rhodopirellula baltica and Oceanicaulis alexandrii have, in contrast to Escherichia coli, membrane insertases with extended positively charged C‐terminal regions similar to the YidC homologues in mitochondria and Gram‐positive bacteria. We have found that chimeric forms of E. coli YidC fused to the C‐terminal YidC regions from the marine bacteria mediate binding of YidC to ribosomes and therefore may have a functional role for targeting a nascent protein to the membrane. Here, we show in E. coli that an extended C‐terminal region of YidC can compensate for a loss of SRP‐receptor function in vivo. Furthermore, the enhanced affinity of the ribosome to the chimeric YidC allows the isolation of a ribosome nascent chain complex together with the C‐terminally elongated YidC chimera. This complex was visualized at 8.6 Å by cryo‐electron microscopy and shows a close contact of the ribosome and a YidC monomer.     

Measurement of dietary exposure: a challenging problem which may be overcome thanks to metabolomics?

The diet is an important environmental exposure, and its measurement is an essential component of much health-related research. However, conventional tools for measuring dietary exposure have significant limitations being subject to an unknown degree of misreporting and dependent upon food composition tables to allow estimation of intakes of energy, nutrients and non-nutrient food constituents. In addition, such tools may be inappropriate for use with certain groups of people. As an alternative approach, the recent techniques of metabolite profiling or fingerprinting, which allows simultaneous monitoring of multiple and dynamic components of biological fluids, may provide metabolic signals indicative of food intake. Samples can be analysed through numerous analytical platforms, followed by multivariate data analysis. In humans, metabolomics has been applied successfully in pharmacology, toxicology and medical screening, but nutritional metabolomics is still in its infancy. Biomarkers of a small number of specific foods and nutrients have been developed successfully but less targeted and more high-throughput methods, that do not need prior knowledge of which signals might be discriminatory, and which may allow a more global characterisation of dietary intake, remain to be tested. A proof a principle project (the MEDE Study) is currently underway in our laboratories to test the hypothesis that high-throughput, non-targeted metabolite fingerprinting using flow injection electrospray mass spectrometry can be applied to human biofluids (blood and urine) to characterise dietary exposure in humans.     

Effects on the male endocrine system of long-term treatment with gonadotropin-releasing hormone agonists and estrogens in male-to-female transsexuals.

We studied hormonal changes resulting from long-term treatment with gonadotropin-releasing hormone agonist and 17beta estradiol valerate in 40 healthy middle-aged male-to-female transsexuals over a period of two years. All of the participants received injections of 3.8 mg goserelin acetate every four weeks in combination with 6 mg oral 17beta estradiol valerate per day for cross-sex hormone treatment for male-to-female transsexuals. There was a significant reduction in the levels of serum luteinizing hormone and follicle-stimulating hormone to the hypogonadal stage. Mean testosterone levels decreased by 97% to 0.52 and 0.59 nmol/l after 12 months and 24 months, respectively. There was a significant reduction in dehydroepiandrosterone sulfate by 37% after 12 months and 43% after 24 months, and androstendione by 29% after 12 months and 27% after 24 months, respectively. Cortisol levels were reduced by 43% and 50%, respectively. Estrogen levels were significantly increased from 77.51 to 677 after 12 months and 661 pmol/l after 24 months. Sex hormone-binding globulin and corticoid-binding globulin levels were significantly increased after 12 and 24 months. There was a significant decrease in all measured androgen fractions and cortisol during long-term treatment with gonadotropin-releasing hormone agonist and 17beta estradiol valerate. Apart from suppression of testicular hormone production, one possible interpretation is that treatment with long-term gonadotropin-releasing hormone agonist and 17beta estradiol valerate influences adrenal hormone levels in healthy middle-aged male-to-female transsexuals. Cortisol serum levels may be decreased due to estrogen-induced increase in corticoid-binding globulin.     

Mitotic requirement for aurora A kinase is bypassed in the absence of aurora B kinase

We investigated why treatment of cells with dual aurora A and B kinase inhibitors produces phenotypes identical to inactivation of aurora B. We found that dual aurora kinase inhibitors in fact potently inhibit cellular activities of both kinases, indicating that inactivation of aurora B bypasses aurora A in mitosis. RNAi experiments further established that inactivation of aurora B indeed bypasses the requirement for aurora A and leads to polyploidy. Inactivation of aurora A activates checkpoint kinase BubR1 in an aurora B-dependent manner. Our results thus show that aurora B is responsible for mitotic arrest in the absence of aurora A.