AMOD: a morpholino oligonucleotide selection tool

AMOD is a web-based program that aids in the functional evaluation of nucleotide sequences through sequence characterization and antisense morpholino oligonucleotide (target site) selection. Submitted sequences are analyzed by translation initiation site prediction algorithms and sequence-to-sequence comparisons; results are used to characterize sequence features required for morpholino design. Within a defined subsequence, base composition and homodimerization values are computed for all putative morpholino oligonucleotides. Using these properties, morpholino candidates are selected and compared with genomic and transcriptome databases with the goal to identify target-specific enriched morpholinos. AMOD has been used at the University of Minnesota to design ∼200 morpholinos for a functional genomics screen in zebrafish. The AMOD web server and a tutorial are freely available to both academic and commercial users at

Storm-induced upwelling of high pCO2 waters onto the continental shelf of the western Arctic Ocean and implications for carbonate mineral saturation states

Author Posting. © American Geophysical Union, 2012. This article is posted here by permission of American Geophysical Union for personal use, not for redistribution. The definitive version was published in Geophysical Research Letters 39 (2012): L07606, doi:10.1029/2012GL051574.The carbon system of the western Arctic Ocean is undergoing a rapid transition as sea ice extent and thickness decline. These processes are dynamically forcing the region, with unknown consequences for CO2 fluxes and carbonate mineral saturation states, particularly in the coastal regions where sensitive ecosystems are already under threat from multiple stressors. In October 2011, persistent wind-driven upwelling occurred in open water along the continental shelf of the Beaufort Sea in the western Arctic Ocean. During this time, cold (32.4) halocline water—supersaturated with respect to atmospheric CO2 (pCO2 > 550 μatm) and undersaturated in aragonite (Ωaragonite < 1.0) was transported onto the Beaufort shelf. A single 10-day event led to the outgassing of 0.18–0.54 Tg-C and caused aragonite undersaturations throughout the water column over the shelf. If we assume a conservative estimate of four such upwelling events each year, then the annual flux to the atmosphere would be 0.72–2.16 Tg-C, which is approximately the total annual sink of CO2 in the Beaufort Sea from primary production. Although a natural process, these upwelling events have likely been exacerbated in recent years by declining sea ice cover and changing atmospheric conditions in the region, and could have significant impacts on regional carbon budgets. As sea ice retreat continues and storms increase in frequency and intensity, further outgassing events and the expansion of waters that are undersaturated in carbonate minerals over the shelf are probable.Funding for this work was provided by the National Science Foundation (ARC1041102 – JTM, OPP0856244-RSP, and ARC1040694- LWJ), the National Oceanic and Atmospheric Administration (CIFAR11021- RHB) and the West Coast & Polar Regions Undersea Research Center (POFP00983 – CLM and JM).2012-10-1

Toward Improved Observing of the Rapidly Changing Arctic Ocean

Arctic Observing Summit (April 30 – May 2, 2013, Vancouver, Canada); AON statementIn order to observe and understand the Arctic Ocean and its response to climate change, the traditional approach of acquiring observations when and where the Arctic is accessible has to be enhanced with multi-faceted measurement systems operating autonomously to provide year-round information in real time. The major goal of such a network of autonomous sensors is to measure and monitor physical, chemical and biological parameters in the atmosphere, sea ice and ocean on at least daily intervals

Generalized calculation of magnetic coupling constants for Mott-Hubbard insulators: Application to ferromagnetic Cr compounds

Using a Rayleigh-Schr\"odinger perturbation expansion of multi-band Hubbard models, we present analytic expressions for the super-exchange coupling constants between magnetic transition metal ions of arbitrary separation in Mott-Hubbard insulators. The only restrictions are i) all ligand ions are closed shell anions and ii) all contributing interaction paths are of equal length. For short paths, our results essentially confirm the Goodenough-Kanamori-Anderson rules, yet in general there does not exist any simple rule to predict the sign of the magnetic coupling constants. The most favorable situation for ferromagnetic coupling is found for ions with less than half filled d shells, the (relative) tendency to ferromagnetic coupling increases with increasing path length. As an application, the magnetic interactions of the Cr compounds Rb$_2$CrCl$_4$, CrCl$_3$, CrBr$_3$ and CrI$_3$ are investigated, all of which except CrCl$_3$ are ferromagnets.Comment: 13 pages, 6 eps figures, submitted to Phys Rev

SUMO chain formation is required for response to replication arrest in S. pombe

SUMO is a ubiquitin-like protein that is post-translationally attached to one or more lysine residues on target proteins. Despite having only 18% sequence identity with ubiquitin, SUMO contains the conserved betabetaalphabetabetaalphabeta fold present in ubiquitin. However, SUMO differs from ubiquitin in having an extended N-terminus. In S. pombe the N-terminus of SUMO/Pmt3 is significantly longer than those of SUMO in S. cerevisiae, human and Drosophila. Here we investigate the role of this N-terminal region. We have used two dimensional gel electrophoresis to demonstrate that S. pombe SUMO/Pmt3 is phosphorylated, and that this occurs on serine residues at the extreme N-terminus of the protein. Mutation of these residues (in pmt3-1) results in a dramatic reduction in both the levels of high Mr SUMO-containing species and of total SUMO/Pmt3, indicating that phosphorylation of SUMO/Pmt3 is required for its stability. Despite the significant reduction in high Mr SUMO-containing species, pmt3-1 cells do not display an aberrant cell morphology or sensitivity to genotoxins or stress. Additionally, we demonstrate that two lysine residues in the N-terminus of S. pombe SUMO/Pmt3 (K14 and K30) can act as acceptor sites for SUMO chain formation in vitro. Inability to form SUMO chains results in aberrant cell and nuclear morphologies, including stretched and fragmented chromatin. SUMO chain mutants are sensitive to the DNA synthesis inhibitor, hydroxyurea (HU), but not to other genotoxins, such as UV, MMS or CPT. This implies a role for SUMO chains in the response to replication arrest in S. pomb

A Conditional Yeast E1 Mutant Blocks the Ubiquitin–Proteasome Pathway and Reveals a Role for Ubiquitin Conjugates in Targeting Rad23 to the Proteasome

E1 ubiquitin activating enzyme catalyzes the initial step in all ubiquitin-dependent processes. We report the isolation of uba1-204, a temperature-sensitive allele of the essential Saccharomyces cerevisiae E1 gene, UBA1. Uba1-204 cells exhibit dramatic inhibition of the ubiquitin–proteasome system, resulting in rapid depletion of cellular ubiquitin conjugates and stabilization of multiple substrates. We have employed the tight phenotype of this mutant to investigate the role ubiquitin conjugates play in the dynamic interaction of the UbL/UBA adaptor proteins Rad23 and Dsk2 with the proteasome. Although proteasomes purified from mutant cells are intact and proteolytically active, they are depleted of ubiquitin conjugates, Rad23, and Dsk2. Binding of Rad23 to these proteasomes in vitro is enhanced by addition of either free or substrate-linked ubiquitin chains. Moreover, association of Rad23 with proteasomes in mutant and wild-type cells is improved upon stabilizing ubiquitin conjugates with proteasome inhibitor. We propose that recognition of polyubiquitin chains by Rad23 promotes its shuttling to the proteasome in vivo

Understanding the Role of the Josephin Domain in the PolyUb Binding and Cleavage Properties of Ataxin-3

Ataxin-3, the disease protein in the neurodegenerative disorder Spinocerebellar Ataxia Type 3 or Machado Joseph disease, is a cysteine protease implicated in the ubiquitin proteasome pathway. It contains multiple ubiquitin binding sites through which it anchors polyubiquitin chains of different linkages that are then cleaved by the N-terminal catalytic (Josephin) domain. The properties of the ubiquitin interacting motifs (UIMs) in the C-terminus of ataxin-3 are well established. Very little is known, however, about how two recently identified ubiquitin-binding sites in the Josephin domain contribute to ubiquitin chain binding and cleavage. In the current study, we sought to define the specific contribution of the Josephin domain to the catalytic properties of ataxin-3 and assess how the topology and affinity of these binding sites modulate ataxin-3 activity. Using NMR we modeled the structure of diUb/Josephin complexes and showed that linkage preferences are imposed by the topology of the two binding sites. Enzymatic studies further helped us to determine a precise hierarchy between the sites. We establish that the structure of Josephin dictates specificity for K48-linked chains. Site 1, which is close to the active site, is indispensable for cleavage. Our studies open the way to understand better the cellular function of ataxin-3 and its link to pathology

Remote climate forcing of decadal-scale regime shifts in Northwest Atlantic shelf ecosystems

Author Posting. © Association for the Sciences of Limnology and Oceanography, 2013. This article is posted here by permission of Association for the Sciences of Limnology and Oceanography for personal use, not for redistribution. The definitive version was published in Association for the Sciences of Limnology and Oceanography, doi:10.4319/lo.2013.58.3.0803.Decadal-scale regime shifts in Northwest Atlantic shelf ecosystems can be remotely forced by climate-associated atmosphere–ocean interactions in the North Atlantic and Arctic Ocean Basins. This remote climate forcing is mediated primarily by basin- and hemispheric-scale changes in ocean circulation. We review and synthesize results from process-oriented field studies and retrospective analyses of time-series data to document the linkages between climate, ocean circulation, and ecosystem dynamics. Bottom-up forcing associated with climate plays a prominent role in the dynamics of these ecosystems, comparable in importance to that of top-down forcing associated with commercial fishing. A broad perspective, one encompassing the effects of basin- and hemispheric-scale climate processes on marine ecosystems, will be critical to the sustainable management of marine living resources in the Northwest Atlantic.Funding for this research was provided by the National Science Foundation as part of the Regional and Pan-Regional Synthesis Phases of the U.S. Global Ocean Ecosystem (GLOBEC) Program

Functional Analysis of Human Hematopoietic Stem Cell Gene Expression Using Zebrafish

Although several reports have characterized the hematopoietic stem cell (HSC) transcriptome, the roles of HSC-specific genes in hematopoiesis remain elusive. To identify candidate regulators of HSC fate decisions, we compared the transcriptome of human umbilical cord blood and bone marrow CD34(+)CD33(−)CD38(−)Rho(lo)c-kit(+) cells, enriched for hematopoietic stem/progenitor cells with CD34(+)CD33(−)CD38(−)Rho(hi) cells, enriched in committed progenitors. We identified 277 differentially expressed transcripts conserved in these ontogenically distinct cell sources. We next performed a morpholino antisense oligonucleotide (MO)-based functional screen in zebrafish to determine the hematopoietic function of 61 genes that had no previously known function in HSC biology and for which a likely zebrafish ortholog could be identified. MO knock down of 14/61 (23%) of the differentially expressed transcripts resulted in hematopoietic defects in developing zebrafish embryos, as demonstrated by altered levels of circulating blood cells at 30 and 48 h postfertilization and subsequently confirmed by quantitative RT-PCR for erythroid-specific hbae1 and myeloid-specific lcp1 transcripts. Recapitulating the knockdown phenotype using a second MO of independent sequence, absence of the phenotype using a mismatched MO sequence, and rescue of the phenotype by cDNA-based overexpression of the targeted transcript for zebrafish spry4 confirmed the specificity of MO targeting in this system. Further characterization of the spry4-deficient zebrafish embryos demonstrated that hematopoietic defects were not due to more widespread defects in the mesodermal development, and therefore represented primary defects in HSC specification, proliferation, and/or differentiation. Overall, this high-throughput screen for the functional validation of differentially expressed genes using a zebrafish model of hematopoiesis represents a major step toward obtaining meaningful information from global gene profiling of HSCs