Proliferation-dependent differential regulation of the dolichol pathway genes in Saccharomyces cerevisiae

The dolichol pathway serves in the synthesis of the dolichol-linked oligosaccharide precursor for protein N-glycosylation. Recently, we reported that mRNAs of genes that function at the early steps in the dolichol pathway in yeast, ALG7, ALG1 and ALG2, were co-ordinately induced following growth stimulation of G0-arrested cells in a manner similar to that of the transcripts of the early growth response genes (Kukuruzinska,M.A. and Lennon,K. Glycobiology, 4, 437-443, 1994). To determine whether the entire dolichol pathway was co-ordinately regulated with growth, we examined the expression of genes functioning late in the pathway, including two genes encoding oligosaccharyltransferase subunits, at two critical control points in the G1 phase of cell cycle: G0/G1 and START. We show that early in G1, at the G0/G1 transition point, the late ALG genes and the two oligosaccharyltransferase-encoding genes examined were regulated co-ordinately with the early ALG genes: they were downregulated upon exit from the mitotic cell cycle into G0, and they were induced following growth stimulation in the absence of de novo protein synthesis. All the dolichol pathway genes produced transcripts with short, half-lives that were rapidly stabilized in the presence of cycloheximide. In contrast, cell division arrest late in G1, at START, was accompanied by a selective downregulation of only the first dolichol pathway gene, ALG7, and not of the genes functioning later in the pathway. These results indicate that, depending on their position in G1, cells either co-ordinately or differentially regulate the dolichol pathway gene

Exploring and developing student midwives' experiences (ESME) : an appreciative inquiry study

Background: There is increased focus on investing in midwifery students as our future workforce. Inquiring into what helps to support an enriched learning experience for student midwives in clinical placements is timely. Aim: To work collaboratively with key stakeholders (student midwives, midwives) in clinical placements to generate an experience-based understanding of what works well in relation to the student midwife experience and from this understanding, co-create ways to enhance students’ experiences. Design: An appreciative inquiry approach was used to discover what matters and what works well at present in the student midwife experience from the perspective of student midwives, midwives, and midwifery managers and to use this knowledge to create enhanced experiences in the future. Data were generated across four local health districts in New South Wales, Australia. Data were analysed using immersion crystallisation and then mapped to the ‘Senses Framework’. Setting: Four midwifery units in tertiary teaching public hospitals in NSW. Participants: There were 124 participants in this study: 45 midwifery students and 76 employed midwives. Measurements and findings: The data culminated in the refinement of the ‘Senses Framework’ for use in the midwifery learning context. Student midwives and midwives valued experiences that helped them to feel safe, to feel that they belong, to experience continuity in their learning and work, to have a sense of purpose, to have their achievements and their contributions to be recognised and to feel that they matter. Furthermore, the midwives themselves valued the experience of these senses in supporting them to be facilitators of learning in the workplace. The relational framework for learning together in the work-place has the potential to support achievement of the sense of security, belonging, continuity, purpose, achievement and significance for all involved. Key conclusions: There is much to celebrate in what is being achieved currently in promoting excellence in learning experiences in the midwifery context. In particular this study has made conscious the contribution that student midwives and midwives can and do make to enable the positive and reciprocal relationships that develop in the student midwife experience that support the nurturing of enriched learning environments. This study emphasised that learning in the workplace is a relational endeavour, rooted in the day to day engagement between student midwives, midwives and others. By mapping these positive processes to the senses framework these processes are made more explicit and provide guidance for enhancing the learning experience in the midwifery context. Implications for practice: The framework and related inquiry tools developed from the study may be useful in other settings to further test out the impact of this relational approach to learning for student midwives

Determination of the mass of the neutron star in SMC X-1, LMC X-4 and Cen X-3 with VLT/UVES

We present the results of a spectroscopic monitoring campaign of the OB-star companions to the eclipsing X-ray pulsars SMC X-1, LMC X-4 and Cen X-3. High-resolution optical spectra obtained with UVES on the ESO Very Large Telescope are used to determine the radial-velocity orbit of the OB (super)giants with high precision. The excellent quality of the spectra provides the opportunity to measure the radial-velocity curve based on individual lines, and to study the effect of possible distortions of the line profiles due to e.g. X-ray heating on the derived radial-velocity amplitude. Several spectral lines show intrinsic variations with orbital phase. The magnitude of these variations depends on line strength, and thus provides a criterion to select lines that do not suffer from distortions. The undistorted lines show a larger radial-velocity amplitude than the distorted lines, consistent with model predictions. Application of our line-selection criteria results in a mean radial-velocity amplitude K(Opt) of 20.2 +/- 1.1, 35.1 +/- 1.5, and 27.5 +/- 2.3 km/s (1 sigma errors), for the OB companion to SMC X-1, LMC X-4 and Cen X-3, respectively. Adding information on the projected rotational velocity of the OB companion (derived from our spectra), the duration of X-ray eclipse and orbital parameters of the X-ray pulsar (obtained from literature), we arrive at a neutron star mass of 1.06^{+0.11}_{-0.10}, 1.25^{+0.11}_{-0.10} and 1.34^{+0.16}_{-0.14} M{sun} for SMC X-1, LMC X-4 and Cen X-3, respectively. The mass of SMC X-1 is near the minimum mass (~1 M{sun}) expected for a neutron star produced in a supernova. We discuss the implications of the measured mass distribution on the neutron-star formation mechanism, in relation to the evolutionary history of the massive binaries.Comment: Accepted for publication in Astronomy and Astrophysic

The Near-Earth Object Surveyor Mission

The Near-Earth Object (NEO) Surveyor mission is a NASA observatory designed to discover and characterize near-Earth asteroids and comets. The mission's primary objective is to find the majority of objects large enough to cause severe regional impact damage ($>$140 m in effective spherical diameter) within its five-year baseline survey. Operating at the Sun-Earth L1 Lagrange point, the mission will survey to within 45 degrees of the Sun in an effort to find the objects in the most Earth-like orbits. The survey cadence is optimized to provide observational arcs long enough to reliably distinguish near-Earth objects from more distant small bodies that cannot pose an impact hazard. Over the course of its survey, NEO Surveyor will discover $\sim$200,000 - 300,000 new NEOs down to sizes as small as $\sim$10 m and thousands of comets, significantly improving our understanding of the probability of an Earth impact over the next century.Comment: accepted to PS

A framework for human microbiome research

A variety of microbial communities and their genes (the microbiome) exist throughout the human body, with fundamental roles in human health and disease. The National Institutes of Health (NIH)-funded Human Microbiome Project Consortium has established a population-scale framework to develop metagenomic protocols, resulting in a broad range of quality-controlled resources and data including standardized methods for creating, processing and interpreting distinct types of high-throughput metagenomic data available to the scientific community. Here we present resources from a population of 242 healthy adults sampled at 15 or 18 body sites up to three times, which have generated 5,177 microbial taxonomic profiles from 16S ribosomal RNA genes and over 3.5 terabases of metagenomic sequence so far. In parallel, approximately 800 reference strains isolated from the human body have been sequenced. Collectively, these data represent the largest resource describing the abundance and variety of the human microbiome, while providing a framework for current and future studies

Structure, function and diversity of the healthy human microbiome

Author Posting. © The Authors, 2012. This article is posted here by permission of Nature Publishing Group. The definitive version was published in Nature 486 (2012): 207-214, doi:10.1038/nature11234.Studies of the human microbiome have revealed that even healthy individuals differ remarkably in the microbes that occupy habitats such as the gut, skin and vagina. Much of this diversity remains unexplained, although diet, environment, host genetics and early microbial exposure have all been implicated. Accordingly, to characterize the ecology of human-associated microbial communities, the Human Microbiome Project has analysed the largest cohort and set of distinct, clinically relevant body habitats so far. We found the diversity and abundance of each habitat’s signature microbes to vary widely even among healthy subjects, with strong niche specialization both within and among individuals. The project encountered an estimated 81–99% of the genera, enzyme families and community configurations occupied by the healthy Western microbiome. Metagenomic carriage of metabolic pathways was stable among individuals despite variation in community structure, and ethnic/racial background proved to be one of the strongest associations of both pathways and microbes with clinical metadata. These results thus delineate the range of structural and functional configurations normal in the microbial communities of a healthy population, enabling future characterization of the epidemiology, ecology and translational applications of the human microbiome.This research was supported in part by National Institutes of Health grants U54HG004969 to B.W.B.; U54HG003273 to R.A.G.; U54HG004973 to R.A.G., S.K.H. and J.F.P.; U54HG003067 to E.S.Lander; U54AI084844 to K.E.N.; N01AI30071 to R.L.Strausberg; U54HG004968 to G.M.W.; U01HG004866 to O.R.W.; U54HG003079 to R.K.W.; R01HG005969 to C.H.; R01HG004872 to R.K.; R01HG004885 to M.P.; R01HG005975 to P.D.S.; R01HG004908 to Y.Y.; R01HG004900 to M.K.Cho and P. Sankar; R01HG005171 to D.E.H.; R01HG004853 to A.L.M.; R01HG004856 to R.R.; R01HG004877 to R.R.S. and R.F.; R01HG005172 to P. Spicer.; R01HG004857 to M.P.; R01HG004906 to T.M.S.; R21HG005811 to E.A.V.; M.J.B. was supported by UH2AR057506; G.A.B. was supported by UH2AI083263 and UH3AI083263 (G.A.B., C. N. Cornelissen, L. K. Eaves and J. F. Strauss); S.M.H. was supported by UH3DK083993 (V. B. Young, E. B. Chang, F. Meyer, T. M. S., M. L. Sogin, J. M. Tiedje); K.P.R. was supported by UH2DK083990 (J. V.); J.A.S. and H.H.K. were supported by UH2AR057504 and UH3AR057504 (J.A.S.); DP2OD001500 to K.M.A.; N01HG62088 to the Coriell Institute for Medical Research; U01DE016937 to F.E.D.; S.K.H. was supported by RC1DE0202098 and R01DE021574 (S.K.H. and H. Li); J.I. was supported by R21CA139193 (J.I. and D. S. Michaud); K.P.L. was supported by P30DE020751 (D. J. Smith); Army Research Office grant W911NF-11-1-0473 to C.H.; National Science Foundation grants NSF DBI-1053486 to C.H. and NSF IIS-0812111 to M.P.; The Office of Science of the US Department of Energy under Contract No. DE-AC02-05CH11231 for P.S. C.; LANL Laboratory-Directed Research and Development grant 20100034DR and the US Defense Threat Reduction Agency grants B104153I and B084531I to P.S.C.; Research Foundation - Flanders (FWO) grant to K.F. and J.Raes; R.K. is an HHMI Early Career Scientist; Gordon&BettyMoore Foundation funding and institutional funding fromthe J. David Gladstone Institutes to K.S.P.; A.M.S. was supported by fellowships provided by the Rackham Graduate School and the NIH Molecular Mechanisms in Microbial Pathogenesis Training Grant T32AI007528; a Crohn’s and Colitis Foundation of Canada Grant in Aid of Research to E.A.V.; 2010 IBM Faculty Award to K.C.W.; analysis of the HMPdata was performed using National Energy Research Scientific Computing resources, the BluBioU Computational Resource at Rice University

Clinical Sequencing Exploratory Research Consortium: Accelerating Evidence-Based Practice of Genomic Medicine

Despite rapid technical progress and demonstrable effectiveness for some types of diagnosis and therapy, much remains to be learned about clinical genome and exome sequencing (CGES) and its role within the practice of medicine. The Clinical Sequencing Exploratory Research (CSER) consortium includes 18 extramural research projects, one National Human Genome Research Institute (NHGRI) intramural project, and a coordinating center funded by the NHGRI and National Cancer Institute. The consortium is exploring analytic and clinical validity and utility, as well as the ethical, legal, and social implications of sequencing via multidisciplinary approaches; it has thus far recruited 5,577 participants across a spectrum of symptomatic and healthy children and adults by utilizing both germline and cancer sequencing. The CSER consortium is analyzing data and creating publically available procedures and tools related to participant preferences and consent, variant classification, disclosure and management of primary and secondary findings, health outcomes, and integration with electronic health records. Future research directions will refine measures of clinical utility of CGES in both germline and somatic testing, evaluate the use of CGES for screening in healthy individuals, explore the penetrance of pathogenic variants through extensive phenotyping, reduce discordances in public databases of genes and variants, examine social and ethnic disparities in the provision of genomics services, explore regulatory issues, and estimate the value and downstream costs of sequencing. The CSER consortium has established a shared community of research sites by using diverse approaches to pursue the evidence-based development of best practices in genomic medicine

Finishing the euchromatic sequence of the human genome

The sequence of the human genome encodes the genetic instructions for human physiology, as well as rich information about human evolution. In 2001, the International Human Genome Sequencing Consortium reported a draft sequence of the euchromatic portion of the human genome. Since then, the international collaboration has worked to convert this draft into a genome sequence with high accuracy and nearly complete coverage. Here, we report the result of this finishing process. The current genome sequence (Build 35) contains 2.85 billion nucleotides interrupted by only 341 gaps. It covers ∼99% of the euchromatic genome and is accurate to an error rate of ∼1 event per 100,000 bases. Many of the remaining euchromatic gaps are associated with segmental duplications and will require focused work with new methods. The near-complete sequence, the first for a vertebrate, greatly improves the precision of biological analyses of the human genome including studies of gene number, birth and death. Notably, the human enome seems to encode only 20,000-25,000 protein-coding genes. The genome sequence reported here should serve as a firm foundation for biomedical research in the decades ahead

The dolichol pathway of protein N-glycosylation: molecular genetics and functional significance

Thesis (D.Sc.D.)--Boston University, Henry M. Goldman School of Dental Medicine, 1998 (Oral Biology).Includes bibliographical references: (leaves 110-176).Protein N-glycosylation is metabolic process that is obligatory for viability in all eukaryotes. It results in the modification of appropriate asparagine residues of polypeptides with oligosaccharide structures, influencing their properties and bioactivities. N-glycosylation is initiated, via the dolichol pathway, with the synthesis of the lipid-linked oligosaccharide precursor that is later transferred to protein. This synthesis occurs in the endoplasmic reticulum and involves the expression of a set of evolutionarily conserved ALG genes. Contrary to the previous dogma, recent studies suggested that the early dolichol pathway genes are highly regulated, early growth response genes (Kukuruzinska and Lennon, 1994). RNA blotting assays presented here indicate that all the ALG genes, as well as two oligosaccharyltransferase (OT) subunits, are coordinately regulated with growth early in G1, at the GJG1 transition point. In contrast, at a second point in the cell cycle critical to proliferation, late in G1, the genes of the dolichol pathway and OT are differentially regulated. Specifically, ALG7 expression is selectively downregulated in response to cell cycle arrest. This proliferation­ dependent expression of the first dolichol pathway gene, ALG7, is also demonstrated in a higher vertebrate system, zebrafish, via immunohistochemistry and Western blotting. The mechanisms involved in the regulation of ALG7 expression in yeast were assessed with RNA blotting assays. Evidence is provided that this gene is controlled on transcriptional and post-transcriptional levels. Moreover, data from in vitro transcription/translation studies of the mammalian ALG7 gene suggest the possibility of additional control of this gene's expression, on the level of translation. Finally, the functional significance of the early dolichol pathway genes, and ALG7 in particular, is demonstrated through the use of disparate methodologies that include RNA blotting, flow cytometry, fluorescence and light microscopy. The results suggest that the early ALG genes have developmental relevance, since unwarranted changes in their expression cause defects in proliferation, differentiation and cytoskeletal organization. Moreover, these studies reveal potential downstream genetic targets for the ALG genes, and protein N-­glycosylation in general, that include G1 cyclins and proteins involved in organization of the actin cytoskeleton