bluuebunny/crossref_metadata_2025 huggingface dataset:
Novel Endogenous Type C Retrovirus in Baboons: Complete Sequence, Providing Evidence for Baboon Endogenous Virus
gag-pol
Ancestry
Authors: Mang Rui , Goudsmit Jaap , van der Kuyl Antoinette C.
Published: December 2019

A complete endogenous type C viral genome has been isolated from a baboon genomic library. The provirus,
Papio cynocephalus
endogenous retrovirus (PcEV), is 8,572 nucleotides long, and 38 to 59 proviral copies per baboon genome are found. The PcEV provirus possesses the typical simple retroviral gene organization, including two long terminal repeats and genes encoding gag, pol, and env proteins. The open reading frames for
gag-pol
and
env
are complete but have premature stop codons or frameshift mutations. The primer binding site of PcEV is complementary to tRNA
Gly
. The
gag
and
pol
genes of PcEV are closely related to those of the baboon endogenous virus (BaEV). The
env
coding region of PcEV is related to the
env
genes of type C retroviruses. This suggests that PcEV is one of the ancestors of BaEV contributing the type C
gag-pol
genome fragment to the type C/D recombinant virus BaEV. Earlier it was shown that another endogenous type D virus (simian endogenous retrovirus) provided the
env
gene for BaEV (A. C. van der Kuyl et al., J. Virol. 71:3666–3676, 1997).
Cytomegalovirus (CMV) DNA Load Is an Independent Predictor of CMV Disease and Survival in Advanced AIDS
Authors: Spector Stephen A. , Hsia Karen , Crager Michael , Pilcher Marshall , Cabral Sheila , Stempien Mary Jean
Published: December 2019

The impact of cytomegalovirus (CMV) on human immunodeficiency virus type 1 (HIV-1) disease progression has been controversial. In this study, we sought to determine if CMV viral load is independent of HIV-1 viral load in predicting CMV disease and survival. Our findings indicate that in patients with advanced AIDS, CMV DNA load is an independent marker of CMV disease and survival and is more predictive than HIV-1 RNA load. Moreover, patients who respond to preemptive therapy with oral ganciclovir, with resulting undetectable levels of CMV DNA, in their plasma, have a significantly lower risk of developing CMV disease and higher rates of survival, despite stable or increasing HIV-1 RNA loads. These data provide support for CMV as an independent risk factor for mortality in persons with advanced AIDS and further suggest that effective preemptive therapy for CMV can improve patient survival rates.
E4ORF3 Requirement for Achieving Long-Term Transgene Expression from the Cytomegalovirus Promoter in Adenovirus Vectors
Authors: Armentano Donna , Smith Michael P. , Sookdeo Cathleen C. , Zabner Joseph , Perricone Michael A. , St. George Judith A. , Wadsworth Samuel C. , Gregory Richard J.
Published: December 2019

Analysis of transgene expression under the control of the cytomegalovirus (CMV) promoter from adenovirus vectors in which the E4 region was modified indicated that E4ORF3 is required for long-term expression in the murine lung. CMV promoter truncation led to the persistence of expression in the absence of E4, thus eliminating the ORF3 requirement.
Mass Determination of Rous Sarcoma Virus Virions by Scanning Transmission Electron Microscopy
Authors: Vogt Volker M. , Simon Martha N.
Published: December 2019

The internal structural protein of retroviruses, Gag, comprises most of the mass of the virion, and Gag itself can give rise to virus-like particles when expressed in appropriate cells. Previously the stoichiometry of Gag in virions was inferred from indirect measurements carried out 2 decades ago. We now have directly determined the masses of individual particles of the prototypic avian retrovirus, Rous sarcoma virus (RSV), by using scanning transmission electron microscopy. In this technique, the number of scattered electrons in the dark-field image integrated over an individual freeze-dried virus particle on a grid is directly proportional to its mass. The RSV virions had a mean mass of 2.5 × 10
8
Da, corresponding to about 1,500 Gag molecules per virion. The population of virions was not homogeneous, with about one-third to two-thirds of the virions deviating from the mean by more than 10% of the mass in two respective preparations. The mean masses for virions carrying genomes of 7.4 or 9.3 kb were indistinguishable, suggesting that mass variability is not due to differences in RNA incorporation.
Broad, Intense Anti-Human Immunodeficiency Virus (HIV) Ex Vivo CD8+Responses in HIV Type 1-Infected Patients: Comparison with Anti-Epstein-Barr Virus Responses and Changes during Antiretroviral Therapy
Authors: Dalod Marc , Dupuis Marion , Deschemin Jean-Christophe , Sicard Didier , Salmon Dominique , Delfraissy Jean-Francois , Venet Alain , Sinet Martine , Guillet Jean-Gerard
Published: December 2019

The ex vivo antiviral CD8+repertoires of 34 human immunodeficiency virus (HIV)-seropositive patients with various CD4+T-cell counts and virus loads were analyzed by gamma interferon enzyme-linked immunospot assay, using peptides derived from HIV type 1 and Epstein-Barr virus (EBV). Most patients recognized many HIV peptides, with markedly high frequencies, in association with all the HLA class I molecules tested. We found no correlation between the intensity of anti-HIV CD8+responses and the CD4+counts or virus load. In contrast, the polyclonality of anti-HIV CD8+responses was positively correlated with the CD4+counts. The anti-EBV responses were significantly less intense than the anti-HIV responses and were positively correlated with the CD4+counts. Longitudinal follow-up of several patients revealed the remarkable stability of the anti-HIV and anti-EBV CD8+responses in two patients with stable CD4+counts, while both antiviral responses decreased in two patients with obvious progression toward disease. Last, highly active antiretroviral therapy induced marked decreases in the number of anti-HIV CD8+T cells, while the anti-EBV responses increased. These findings emphasize the magnitude of the ex vivo HIV-specific CD8+responses at all stages of HIV infection and suggest that the CD8+hyperlymphocytosis commonly observed in HIV infection is driven mainly by virus replication, through intense, continuous activation of HIV-specific CD8+T cells until ultimate progression toward disease. Nevertheless, highly polyclonal anti-HIV CD8+responses may be associated with a better clinical status. Our data also suggest that a decrease of anti-EBV CD8+responses may occur with depletion of CD4+T cells, but this could be restored by highly active antiretroviral treatment.
Modulation of Hepatitis C Virus NS5A Hyperphosphorylation by Nonstructural Proteins NS3, NS4A, and NS4B
Authors: Oliver Koch Jan , Bartenschlager Ralf
Published: December 2019

NS5A of the hepatitis C virus (HCV) is a highly phosphorylated protein involved in resistance against interferon and required most likely for replication of the viral genome. Phosphorylation of this protein is mediated by a cellular kinase(s) generating multiple proteins with different electrophoretic mobilities. In the case of the genotype 1b isolate HCV-J, in addition to the basal phosphorylated NS5A (designated pp56), a hyperphosphorylated form (pp58) was found on coexpression of NS4A (T. Kaneko, Y. Tanji, S. Satoh, M. Hijikata, S. Asabe, K. Kimura, and K. Shimotohno, Biochem. Biophys. Res. Commun. 205:320–326, 1994). Using a comparative analysis of two full-length genomes of genotype 1b, competent or defective for NS5A hyperphosphorylation, we investigated the requirements for this NS5A modification. We found that hyperphosphorylation occurs when NS5A is expressed as part of a continuous NS3-5A polyprotein but not when it is expressed on its own or
trans
complemented with one or several other viral proteins. Results obtained with chimeras of both genomes show that single amino acid substitutions within NS3 that do not affect polyprotein cleavage can enhance or reduce NS5A hyperphosphorylation. Furthermore, mutations in the central or carboxy-terminal NS4A domain as well as small deletions in NS4B can also reduce or block hyperphosphorylation without affecting polyprotein processing. These requirements most likely reflect the formation of a highly ordered NS3-5A multisubunit complex responsible for the differential phosphorylation of NS5A and probably also for modulation of its biological activities.
Studies of the Genomic RNA of Leukosis Viruses: Implications for RNA Dimerization
Authors: Ortiz-Conde Betty A. , Hughes Stephen H.
Published: December 2019

Retroviral particles contain two positive-strand genomic RNAs linked together by noncovalent bonds that can be dissociated under mild conditions. We studied genomic RNAs of wild-type and mutant avian leukosis viruses (ALVs) in an attempt to (i) better understand the site(s) of RNA dimerization, (ii) examine whether the primer binding site (PBS) and tRNA primer are involved in dimerization, and (iii) determine the structure of genomic RNA in protease-deficient (PR
−
) mutants. We showed that extensively nicked wild-type ALV genomic RNAs melt cooperatively. This implies a complex secondary and/or tertiary structure for these RNAs that extends well beyond the 5′ dimerization site. To investigate the role of the PBS-tRNA complex in dimerization, we analyzed genomic RNAs from mutant viruses in which the tRNA
Trp
PBS had been replaced with sequences homologous to the 3′ end of six other chicken tRNAs. We found the genomic RNAs of these viruses are dimers that dissociate at the same temperature as wild-type viral RNA, which suggests that the identity of the PBS and the tRNA primer do not affect dimer stability. We studied two ALV PR
−
mutants: one containing a large (>1.9-kb) inversion spanning the 3′ end of
gag
and much of
pol
, rendering it deficient in PR, reverse transcriptase, and integrase, and another with a point mutation in PR. In both of these mutant viruses, the genomic RNA appears to be either primarily or exclusively monomeric. These data suggest that ALV can package its RNA as monomers that subsequently dimerize.
Picornavirus Receptor Down-Regulation by Plasminogen Activator Inhibitor Type 2
Authors: Shafren D. R. , Gardner J. , Mann V. H. , Antalis T. M. , Suhrbier A.
Published: December 2019

Therapeutic interference with virus-cell surface receptor interactions represents a viable antiviral strategy. Here we demonstrate that cytoplasmic expression of the serine protease inhibitor (serpin), plasminogen activator inhibitor type 2 (PAI-2), affords a high level of protection from lytic infection by multiple human picornaviruses. The antiviral action of PAI-2 was mediated primarily through transcriptional down-regulation of the following virus receptors: intercellular adhesion molecule 1 (ICAM-1, a cellular receptor for the major group of rhinoviruses), decay-accelerating factor (a cellular receptor for echoviruses and coxsackieviruses), and to a lesser extent the coxsackie-adenovirus receptor protein (a cellular receptor for group B coxsackieviruses and group C adenoviruses). Expression of related cell surface receptors, including membrane cofactor protein and the poliovirus receptor, remained unaffected. These findings suggest that PAI-2 and/or related serpins may form the basis of novel antiviral strategies against picornavirus infections and also therapeutic interventions against ICAM-1-mediated respiratory inflammation.
Sequence Requirements for the Assembly of Simian Virus 40 T Antigen and the T-Antigen Origin Binding Domain on the Viral Core Origin of Replication
Authors: Kim Henry Y. , Barbaro Brett A. , Joo Woo S. , Prack Andrea E. , Sreekumar K. R. , Bullock Peter A.
Published: December 2019

The regions of the simian virus 40 (SV40) core origin that are required for stable assembly of virally encoded T antigen (T-ag) and the T-ag origin binding domain (T-ag-obd
131–260
) have been determined. Binding of the purified T-ag-obd
131–260
is mediated by interactions with the central region of the core origin, site II. In contrast, T-ag binding and hexamer assembly requires a larger region of the core origin that includes both site II and an additional fragment of DNA that may be positioned on either side of site II. These studies indicate that in the context of T-ag, the origin binding domain can engage the pentanucleotides in site II only if a second region of T-ag interacts with one of the flanking sequences. The requirements for T-ag double-hexamer assembly are complex; the nucleotide cofactor present in the reaction modulates the sequence requirements for oligomerization. Nevertheless, these experiments provide additional evidence that only a subset of the SV40 core origin is required for assembly of T-ag double hexamers.
The E7 Oncoprotein of Human Papillomavirus Type 16 Stabilizes p53 through a Mechanism Independent of p19
ARF
Authors: Seavey Scott E. , Holubar Marisa , Saucedo Leslie J. , Perry Mary Ellen
Published: December 2019

High-risk human papillomaviruses are causally associated with cervical cancer. Two viral oncogenes, E6 and E7, are expressed in most cervical cancers, and these genes cause cancer when expressed in experimental animals. The E6 protein targets the p53 tumor suppressor for degradation, while the E7 protein inactivates the retinoblastoma susceptibility protein (pRb), in part by stimulating its degradation. In contrast, expression of E7 in the absence of E6 leads to stabilization of p53. Here we show that E7 stabilizes p53 in mouse embryo fibroblasts lacking p19
ARF
. The stable p53 is active as a transcriptional activator, as evidenced by the increased expression of the p53-responsive
mdm2
gene. Normally, MDM2 protein inhibits p53 function in an autoregulatory loop. Regulation of p53 by MDM2 is required for murine development as well as for proliferation of cultured human fibroblasts. However, E7-expressing human fibroblasts continue to divide even though E7 abrogates the ability of MDM2 and p53 to bind. Furthermore, E7-expressing cells are not more sensitive to UV light, an agent that has been reported to induce apoptosis mediated by p53. These results indicate that in addition to inhibiting the ability of MDM2 to regulate p53, E7 must block signaling steps downstream of p53 to allow cell division.
A Recombinant Hepatitis C Virus RNA-Dependent RNA Polymerase Capable of Copying the Full-Length Viral RNA
Authors: Oh Jong-Won , Ito Takayoshi , Lai Michael M. C.
Published: December 2019

All of the previously reported recombinant RNA-dependent RNA polymerases (RdRp), the NS5B enzymes, of hepatitis C virus (HCV) could function only in a primer-dependent and template-nonspecific manner, which is different from the expected properties of the functional viral enzymes in the cells. We have now expressed a recombinant NS5B that is able to synthesize a full-length HCV genome in a template-dependent and primer-independent manner. The kinetics of RNA synthesis showed that this RdRp can initiate RNA synthesis de novo and yield a full-length RNA product of genomic size (9.5 kb), indicating that it did not use the copy-back RNA as a primer. This RdRp was also able to accept heterologous viral RNA templates, including poly(A)- and non-poly(A)-tailed RNA, in a primer-independent manner, but the products in these cases were heterogeneous. The RdRp used some homopolymeric RNA templates only in the presence of a primer. By using the 3′-end 98 nucleotides (nt) of HCV RNA, which is conserved in all genotypes of HCV, as a template, a distinct RNA product was generated. Truncation of 21 nt from the 5′ end or 45 nt from the 3′ end of the 98-nt RNA abolished almost completely its ability to serve as a template. Inclusion of the 3′-end variable sequence region and the U-rich tract upstream of the X region in the template significantly enhanced RNA synthesis. The 3′ end of minus-strand RNA of HCV genome also served as a template, and it required a minimum of 239 nt from the 3′ end. These data defined the
cis
-acting sequences for HCV RNA synthesis at the 3′ end of HCV RNA in both the plus and minus senses. This is the first recombinant HCV RdRp capable of copying the full-length HCV RNA in the primer-independent manner expected of the functional HCV RNA polymerase.
A New Group of Hepadnaviruses Naturally Infecting Orangutans (
Pongo pygmaeus
)
Authors: Warren Kristin S. , Heeney Jonathan L. , Swan Ralph A. , Heriyanto  , Verschoor Ernst J.
Published: December 2019

A high prevalence (42.6%) of hepatitis B virus (HBV) infection was suspected in 195 formerly captive orangutans due to a large number of serum samples which cross-reacted with human HBV antigens. It was assumed that such viral infections were contracted from humans during captivity. However, two wild orangutans were identified which were HBV surface antigen positive, indicating that HBV or related viruses may be occurring naturally in the orangutan populations. Sequence analyses of seven isolates revealed that orangutans were infected with hepadnaviruses but that these were clearly divergent from the six known human HBV genotypes and those of other nonhuman hepadnaviruses reported. Phylogenetic analyses revealed geographic clustering with Southeast Asian genotype C viruses and gibbon ape HBV. This implies a common origin of infection within this geographic region, with cross-species transmission of hepadnaviruses among hominoids.
Induction of Antiviral Antibodies by DNA Immunization Requires neither Perforin-Mediated nor CD8+-T-Cell-Mediated Lysis of Antigen-Expressing Cells
Authors: Hassett Daniel E. , Zhang Jie , Whitton J. Lindsay
Published: December 2019

DNA immunization induces antibodies to the encoded protein, which indicates that the protein must gain access to the extracellular milieu, allowing it to interact with naı̈ve B lymphocytes. It has been suggested that antigen release may be effected by cytotoxic-T-lymphocyte-mediated lysis of transfected antigen-expressing cells; this might be particularly important for the induction of responses to a noncytopathic, cytosolic protein. Here we show that the induction of antibody responses to one such DNA-encoded protein required neither perforin nor CD8+T cells. In addition, there was no skewing of the immunoglobulin G isotypes in the absence of perforin.
Rhesus Rhadinovirus Establishes a Latent Infection in B Lymphocytes In Vivo
Authors: Bergquam Eric P. , Avery Nancy , Shiigi Stanley M. , Axthelm Michael K. , Wong Scott W.
Published: December 2019

Recent DNA sequence analysis indicates that rhesus rhadinovirus (RRV) is a member of the lymphotropic gamma-2 herpesvirus family. To determine if RRV is lymphotropic, peripheral blood mononuclear cells from naturally infected monkeys were separated by immunomagnetic bead depletion and analyzed for the presence of RRV by virus isolation and nested PCR. The recovery and consistent detection of RRV in the CD20
+
-enriched fraction clearly demonstrates that B lymphocytes are a major site of virus persistence.
Molecular Characterization of Proteolytic Processing of the Gag Proteins of Human Spumavirus
Authors: Pfrepper Klaus-Ingmar , Löchelt Martin , Rackwitz Hans-Richard , Schnölzer Martina , Heid Hans , Flügel Rolf M.
Published: December 2019

Spumaviruses, or foamy viruses, express Gag proteins that are incompletely processed by the viral protease in cell cultures. To delineate the proteolytic cleavage sites between potential Gag subdomains, recombinant human spumaretrovirus (HSRV) Gag proteins of different lengths were expressed, purified by affinity chromatography, and subjected to HSRV protease assays. HSRV-specific proteolytic cleavage products were isolated and characterized by Western blotting. Peptides spanning potential cleavage sites, as deduced from the sizes of the proteolytic cleavage products, were chemically synthesized and assayed with HSRV protease. The cleaved peptides were then subjected to mass spectrometry. In control experiments, HSRV protease-deficient mutant proteins were used to rule out unspecific processing by nonviral proteases. The cleavage site junctions identified and the calculated sizes of the cleavage products were in agreement with those of the authentic cleavage products of the HSRV Gag proteins detectable in viral proteins from purified HSRV particles and in virus-infected cells. The biological significance of the data was confirmed by mutational analysis of the cleavage sites in a recombinant Gag protein and in the context of the infectious HSRV DNA provirus.
Cell Wall and Secreted Proteins ofCandida albicans: Identification, Function, and Expression
Authors: Chaffin W. Lajean , López-Ribot José Luis , Casanova Manuel , Gozalbo Daniel , Martínez José P.
Published: December 2019

The cell wall is essential to nearly every aspect of the biology and pathogenicity of Candida albicans. Although it was intially considered an almost inert cellular structure that protected the protoplast against osmotic offense, more recent studies have demonstrated that it is a dynamic organelle. The major components of the cell wall are glucan and chitin, which are associated with structural rigidity, and mannoproteins. The protein component, including both mannoprotein and nonmannoproteins, comprises some 40 or more moieties. Wall proteins may differ in their expression, secretion, or topological location within the wall structure. Proteins may be modified by glycosylation (primarily addition of mannose residues), phosphorylation, and ubiquitination. Among the secreted enzymes are those that are postulated to have substrates within the cell wall and those that find substrates in the extracellular environment. Cell wall proteins have been implicated in adhesion to host tissues and ligands. Fibrinogen, complement fragments, and several extracellular matrix components are among the host proteins bound by cell wall proteins. Proteins related to the hsp70 and hsp90 families of conserved stress proteins and some glycolytic enzyme proteins are also found in the cell wall, apparently as bona fide components. In addition, the expression of some proteins is associated with the morphological growth form of the fungus and may play a role in morphogenesis. Finally, surface mannoproteins are strong immunogens that trigger and modulate the host immune response during candidiasis.
Growth Kinetics of Suspended Microbial Cells: From Single-Substrate-Controlled Growth to Mixed-Substrate Kinetics
Authors: Kovárová-Kovar Karin , Egli Thomas
Published: December 2019

Growth kinetics, i.e., the relationship between specific growth rate and the concentration of a substrate, is one of the basic tools in microbiology. However, despite more than half a century of research, many fundamental questions about the validity and application of growth kinetics as observed in the laboratory to environmental growth conditions are still unanswered. For pure cultures growing with single substrates, enormous inconsistencies exist in the growth kinetic data reported. The low quality of experimental data has so far hampered the comparison and validation of the different growth models proposed, and only recently have data collected from nutrient-controlled chemostat cultures allowed us to compare different kinetic models on a statistical basis. The problems are mainly due to (i) the analytical difficulty in measuring substrates at growth-controlling concentrations and (ii) the fact that during a kinetic experiment, particularly in batch systems, microorganisms alter their kinetic properties because of adaptation to the changing environment. For example, for Escherichia coli growing with glucose, a physiological long-term adaptation results in a change in K
S
for glucose from some 5 mg liter
−1
to ca. 30 μg liter
−1
. The data suggest that a dilemma exists, namely, that either “intrinsic” K
S
(under substrate-controlled conditions in chemostat culture) or μ
max
(under substrate-excess conditions in batch culture) can be measured but both cannot be determined at the same time. The above-described conventional growth kinetics derived from single-substrate-controlled laboratory experiments have invariably been used for describing both growth and substrate utilization in ecosystems. However, in nature, microbial cells are exposed to a wide spectrum of potential substrates, many of which they utilize simultaneously (in particular carbon sources). The kinetic data available to date for growth of pure cultures in carbon-controlled continuous culture with defined mixtures of two or more carbon sources (including pollutants) clearly demonstrate that simultaneous utilization results in lowered residual steady-state concentrations of all substrates. This should result in a competitive advantage of a cell capable of mixed-substrate growth because it can grow much faster at low substrate concentrations than one would expect from single-substrate kinetics. Additionally, the relevance of the kinetic principles obtained from defined culture systems with single, mixed, or multicomponent substrates to the kinetics of pollutant degradation as it occurs in the presence of alternative carbon sources in complex environmental systems is discussed. The presented overview indicates that many of the environmentally relevant apects in growth kinetics are still waiting to be discovered, established, and exploited.
Chlorophyll Fluorescence Analysis of Cyanobacterial Photosynthesis and Acclimation
Authors: Campbell Douglas , Hurry Vaughan , Clarke Adrian K. , Gustafsson Petter , Öquist Gunnar
Published: December 2019

Cyanobacteria are ecologically important photosynthetic prokaryotes that also serve as popular model organisms for studies of photosynthesis and gene regulation. Both molecular and ecological studies of cyanobacteria benefit from real-time information on photosynthesis and acclimation. Monitoring in vivo chlorophyll fluorescence can provide noninvasive measures of photosynthetic physiology in a wide range of cyanobacteria and cyanolichens and requires only small samples. Cyanobacterial fluorescence patterns are distinct from those of plants, because of key structural and functional properties of cyanobacteria. These include significant fluorescence emission from the light-harvesting phycobiliproteins; large and rapid changes in fluorescence yield (state transitions) which depend on metabolic and environmental conditions; and flexible, overlapping respiratory and photosynthetic electron transport chains. The fluorescence parameters F
V
/F
M
, F
V
′/F
M
′,q
p
,q
N
, NPQ, and φPS II were originally developed to extract information from the fluorescence signals of higher plants. In this review, we consider how the special properties of cyanobacteria can be accommodated and used to extract biologically useful information from cyanobacterial
in vivo
chlorophyll fluorescence signals. We describe how the pattern of fluorescence yield versus light intensity can be used to predict the acclimated light level for a cyanobacterial population, giving information valuable for both laboratory and field studies of acclimation processes. The size of the change in fluorescence yield during dark-to-light transitions can provide information on respiration and the iron status of the cyanobacteria. Finally, fluorescence parameters can be used to estimate the electron transport rate at the acclimated growth light intensity.
Linkage Map of
Escherichia coli
K-12, Edition 10: The Physical Map
Authors: Rudd Kenneth E.
Published: December 2019

A physical map, EcoMap10, of the now completely sequenced Escherichia coli chromosome is presented. Calculated genomic positions for the eight restriction enzymes BamHI, HindIII, EcoRI, EcoRV, BglI, KpnI, PstI, and PvuII are depicted. Both sequenced and unsequenced Kohara/Isono miniset clones are aligned to this calculated restriction map. DNA sequence searches identify the precise locations of insertion sequence elements and repetitive extragenic palindrome clusters. EcoGene10, a revised set of genes and functionally uncharacterized open reading frames (ORFs), is also depicted on EcoMap10. The complete set of unnamed ORFs in EcoGene10 are assigned provisional names beginning with the letter “y” by using a systematic nomenclature.
Regulation of Cdc28 Cyclin-Dependent Protein Kinase Activity during the Cell Cycle of the Yeast
Saccharomyces cerevisiae
Authors: Mendenhall Michael D. , Hodge Amy E.
Published: December 2019

The cyclin-dependent protein kinase (CDK) encoded by CDC28 is the master regulator of cell division in the budding yeast Saccharomyces cerevisiae. By mechanisms that, for the most part, remain to be delineated, Cdc28 activity controls the timing of mitotic commitment, bud initiation, DNA replication, spindle formation, and chromosome separation. Environmental stimuli and progress through the cell cycle are monitored through checkpoint mechanisms that influence Cdc28 activity at key cell cycle stages. A vast body of information concerning how Cdc28 activity is timed and coordinated with various mitotic events has accrued. This article reviews that literature. Following an introduction to the properties of CDKs common to many eukaryotic species, the key influences on Cdc28 activity—cyclin-CKI binding and phosphorylation-dephosphorylation events—are examined. The processes controlling the abundance and activity of key Cdc28 regulators, especially transcriptional and proteolytic mechanisms, are then discussed in detail. Finally, the mechanisms by which environmental stimuli influence Cdc28 activity are summarized.
Antisense RNA: Function and Fate of Duplex RNA in Cells of Higher Eukaryotes
Authors: Kumar Madhur , Carmichael Gordon G.
Published: December 2019

There is ample evidence that cells of higher eukaryotes express double-stranded RNA molecules (dsRNAs) either naturally or as the result of viral infection or aberrant, bidirectional transcriptional readthrough. These duplex molecules can exist in either the cytoplasmic or nuclear compartments. Cells have evolved distinct ways of responding to dsRNAs, depending on the nature and location of the duplexes. Since dsRNA molecules are not thought to exist naturally within the cytoplasm, dsRNA in this compartment is most often associated with viral infections. Cells have evolved defensive strategies against such molecules, primarily involving the interferon response pathway. Nuclear dsRNA, however, does not induce interferons and may play an important posttranscriptional regulatory role. Nuclear dsRNA appears to be the substrate for enzymes which deaminate adenosine residues to inosine residues within the polynucleotide structure, resulting in partial or full unwinding. Extensively modified RNAs are either rapidly degraded or retained within the nucleus, whereas transcripts with few modifications may be transported to the cytoplasm, where they serve to produce altered proteins. This review summarizes our current knowledge about the function and fate of dsRNA in cells of higher eukaryotes and its potential manipulation as a research and therapeutic tool.
Protein Phylogenies and Signature Sequences: A Reappraisal of Evolutionary Relationships among Archaebacteria, Eubacteria, and Eukaryotes
Authors: Gupta Radhey S.
Published: December 2019

The presence of shared conserved insertion or deletions (indels) in protein sequences is a special type of signature sequence that shows considerable promise for phylogenetic inference. An alternative model of microbial evolution based on the use of indels of conserved proteins and the morphological features of prokaryotic organisms is proposed. In this model, extant archaebacteria and gram-positive bacteria, which have a simple, single-layered cell wall structure, are termed monoderm prokaryotes. They are believed to be descended from the most primitive organisms. Evidence from indels supports the view that the archaebacteria probably evolved from gram-positive bacteria, and I suggest that this evolution occurred in response to antibiotic selection pressures. Evidence is presented that diderm prokaryotes (i.e., gram-negative bacteria), which have a bilayered cell wall, are derived from monoderm prokaryotes. Signature sequences in different proteins provide a means to define a number of different taxa within prokaryotes (namely, low G+C and high G+C gram-positive, Deinococcus-Thermus, cyanobacteria, chlamydia-cytophaga related, and two different groups of Proteobacteria) and to indicate how they evolved from a common ancestor. Based on phylogenetic information from indels in different protein sequences, it is hypothesized that all eukaryotes, including amitochondriate and aplastidic organisms, received major gene contributions from both an archaebacterium and a gram-negative eubacterium. In this model, the ancestral eukaryotic cell is a chimera that resulted from a unique fusion event between the two separate groups of prokaryotes followed by integration of their genomes.
Bacillus subtilis
Spore Coat
Authors: Driks Adam
Published: December 2019

In response to starvation, bacilli and clostridia undergo a specialized program of development that results in the production of a highly resistant dormant cell type known as the spore. A proteinacious shell, called the coat, encases the spore and plays a major role in spore survival. The coat is composed of over 25 polypeptide species, organized into several morphologically distinct layers. The mechanisms that guide coat assembly have been largely unknown until recently. We now know that proper formation of the coat relies on the genetic program that guides the synthesis of spore components during development as well as on morphogenetic proteins dedicated to coat assembly. Over 20 structural and morphogenetic genes have been cloned. In this review, we consider the contributions of the known coat and morphogenetic proteins to coat function and assembly. We present a model that describes how morphogenetic proteins direct coat assembly to the specific subcellular site of the nascent spore surface and how they establish the coat layers. We also discuss the importance of posttranslational processing of coat proteins in coat morphogenesis. Finally, we review some of the major outstanding questions in the field.
Surface Proteins of Gram-Positive Bacteria and Mechanisms of Their Targeting to the Cell Wall Envelope
Authors: Navarre William Wiley , Schneewind Olaf
Published: December 2019

The cell wall envelope of gram-positive bacteria is a macromolecular, exoskeletal organelle that is assembled and turned over at designated sites. The cell wall also functions as a surface organelle that allows gram-positive pathogens to interact with their environment, in particular the tissues of the infected host. All of these functions require that surface proteins and enzymes be properly targeted to the cell wall envelope. Two basic mechanisms, cell wall sorting and targeting, have been identified. Cell well sorting is the covalent attachment of surface proteins to the peptidoglycan via a C-terminal sorting signal that contains a consensus LPXTG sequence. More than 100 proteins that possess cell wall-sorting signals, including the M proteins of Streptococcus pyogenes, protein A of Staphylococcus aureus, and several internalins of Listeria monocytogenes, have been identified. Cell wall targeting involves the noncovalent attachment of proteins to the cell surface via specialized binding domains. Several of these wall-binding domains appear to interact with secondary wall polymers that are associated with the peptidoglycan, for example teichoic acids and polysaccharides. Proteins that are targeted to the cell surface include muralytic enzymes such as autolysins, lysostaphin, and phage lytic enzymes. Other examples for targeted proteins are the surface S-layer proteins of bacilli and clostridia, as well as virulence factors required for the pathogenesis of L. monocytogenes (internalin B) and Streptococcus pneumoniae (PspA) infections. In this review we describe the mechanisms for both sorting and targeting of proteins to the envelope of gram-positive bacteria and review the functions of known surface proteins.
Transposon Tn21, Flagship of the Floating Genome
Authors: Liebert Cynthia A. , Hall Ruth M. , Summers Anne O.
Published: December 2019

The transposon Tn21 and a group of closely related transposons (the Tn21 family) are involved in the global dissemination of antibiotic resistance determinants in gram-negative facultative bacteria. The molecular basis for their involvement is carriage by the Tn21 family of a mobile DNA element (the integron) encoding a site-specific system for the acquisition of multiple antibiotic resistance genes. The paradigm example, Tn21, also carries genes for its own transposition and a mercury resistance (mer) operon. We have compiled the entire 19,671-bp sequence of Tn21 and assessed the possible origins and functions of the genes it contains. Our assessment adds molecular detail to previous models of the evolution of Tn21 and is consistent with the insertion of the integron In2 into an ancestral Tn501-like mer transposon. Codon usage analysis indicates distinct host origins for the ancestral mer operon, the integron, and the gene cassette and two insertion sequences which lie within the integron. The sole gene of unknown function in the integron, orf5, resembles a puromycin-modifying enzyme from an antibiotic producing bacterium. A possible seventh gene in the mer operon (merE), perhaps with a role in Hg(II) transport, lies in the junction between the integron and the mer operon. Analysis of the region interrupted by insertion of the integron suggests that the putative transposition regulator, tnpM, is the C-terminal vestige of a tyrosine kinase sensor present in the ancestral mer transposon. The extensive dissemination of the Tn21 family may have resulted from the fortuitous association of a genetic element for accumulating multiple antibiotic resistances (the integron) with one conferring resistance to a toxic metal at a time when clinical, agricultural, and industrial practices were rapidly increasing the exposure to both types of selective agents. The compendium offered here will provide a reference point for ongoing observations of related elements in multiply resistant strains emerging worldwide.
More than 50% of hip replacements seem to last 25 years
Authors: Cook Rob , Davidson Peter , Martin Rosie
Published: December 2019

The study

How long does a hip replacement last? A systematic review and meta-analysis of case series and national registry reports with more than 15 years of follow-up

Evans JT, Evans JP, Walker R, Blom AW, Whitehouse MR, Sayers A

Published on 16 February 2019 Lancet 2019;393:647-54

This study was supported by the NIHR Biomedical Research Centre at the University Hospitals Bristol NHS Foundation Trust and the University of Bristol.

To read the full NIHR Signal, go to: https://discover.dc.nihr.ac.uk/content/signal-000781/more-than-50-of-hip-replacements-appear-to-last-25-years
Diagnostic property of direct referral from general practitioners to contrast-enhanced thoracoabdominal CT in patients with serious but non-specific symptoms or signs of cancer: a retrospective cohort study on cancer prevalence after 12 months
Authors: Møller Marie , Juvik Bue , Olesen Stine Chabert , Sandstrøm Hanne , Laxafoss Erling , Reuter Simon Bertram , Bodtger Uffe
Published: December 2019

Objectives

To describe the diagnostic properties of thoracoabdominal contrast-enhanced CT (ceCT), when general practitioners (GPs) managed referral to ceCT through the non-specific symptoms or signs of cancer-cancer patient pathway (NSSC-CPP).

Design

Retrospective cohort study including patients from a part of Denmark.

Setting

Department of Internal Medicine at a university hospital.

Participants

In total, 529 patients underwent ceCT.

Primary and secondary outcomes

Our primary objective was to estimate the negative and positive likelihood ratios for being diagnosed with cancer within 1 year after ceCT. Our secondary outcomes were prevalence and final diagnoses of malignancy (including temporal trends since implementation of NSSC-CPP in 2012), the prevalence of revision of CT scans and referral patterns based on ceCT results.

Results

In total, 529 subjects underwent ceCT and malignancy was identified in 104 (19.7%) patients; 101 (97.1%) during initial workup and 3 patients during the subsequent 12 months follow-up.

Eleven patients had a false-negative ceCT, and revision classified the ceCT as ‘probable/possible malignancy’ in eight (73%) patients. The negative predictive value was 98% and positive predictive value 63%. Negative and positive likelihood ratios for malignancy was 0.1 and 7.9, respectively.

Conclusion

Our study shows that ceCT as part of GP-coordinated workup has a low negative likelihood ratio for identifying malignancy; this is important since identifying patients for further workup is vital.
Providing care to refugees through mainstream general practice in the southern health region of New Zealand: a qualitative study of primary healthcare professionals’ perspectives
Authors: Richard Lauralie , Richardson Georgia , Jaye Chrystal , Stokes Tim
Published: December 2019

Objective

To explore the perspectives of primary healthcare (PHC) professionals providing care to refugees through mainstream general practice.

Design

Qualitative exploratory design with semistructured interviews subjected to inductive thematic analysis.

Setting and participants

Nine general practices enrolled in the Dunedin Refugee Resettlement Programme, in New Zealand (NZ)’s southern health region. Participants included nine general practitioners and six practice nurses.

Results

Three analytical constructs were identified: relational engagement with refugees, refugee healthcare delivery and providers’ professional role shaped by complexity. Building meaningful relational connections involved acknowledging refugees’ journeys by getting to know them as people. This was instrumental for the development of an empathetic understanding of the complex human trajectories that characterise refugees’ journeys to NZ. Participants encountered challenges in providing care to refugees with respect to time-limited consultations, variable use of interpreter services, fragmentation of care between agencies and need for improved health infrastructure to ensure a fluid interface between PHC, secondary care and community support services. The current business model of NZ general practice was perceived to interfere with value-driven care and discouraged tailoring of care to specific patient groups, raising concerns about the ‘fit’ of mainstream general practice to address the complex healthcare needs of refugees. Meeting the needs of refugees across the social determinants of health involved a lot of ‘behind the scenes work’ particularly in the absence of shared information systems and the lack of well-established referral pathways to connect refugees to services beyond the health sector. This led to providers feeling overwhelmed and uncertain about their ability to provide appropriate care to refugees.

Conclusions

This study provides rich context-specific findings that enhance PHC responsiveness to the needs of refugees in NZ.
Directed intervention to improve the rate of admission medication reconciliation in an acute care hospital
Authors: Kyi Htay Htay , Sundus Saira , Marcus Huda , Sotzen Jason , Suit Parker , Cranford James , Bachuwa Ghassan , McDonald Philip J
Published: December 2019

Introduction

Electronic medication reconciliation systems are known to reduce medication errors. We hypothesised that refinement of the electronic medical record (EMR) and provider education could improve adherence to completion of admission medication reconciliation, thereby potentially limiting prescribing errors. Our goal was to improve the percentage of patients with medication reconciliation completed within 24 hours of admission to at least 90%.

Methods

A prospective interventional study was conducted at a university-affiliated community hospital between 1 January 2017 and 30 September 2018. We determined the baseline percentage of medication reconciliations performed within 24 hours of admission, and those completed at any time prior to discharge from the hospital. Three plan-do-study-act cycles were then performed, with interventions including live and email reminders to complete medication reconciliation and addition of a column to EMR patient lists indicating whether reconciliation had been completed.

Results

The percentage of medication reconciliations completed within 24 hours of admission was lowest for the pre-intervention cycle (62.4%) and was highest for Cycle 3 (80.9%). The percentage of reconciliations completed any time prior to discharge was higher and increased in a similar stepwise fashion from 71.1% to 88.4% through Cycle 3. There was a post-intervention trend toward a higher rate of reconciliation completion for patients aged 18–40. Male patients were also more likely to have their admission medication reconciliations completed prior to discharge.

Conclusion

Our interventions resulted in a statistically significant 18.5% increase in the rate of admission reconciliation completion. Though this increase fell short of our goal, this study demonstrates that provider education and optimisation of the EMR can sustainably improve adherence with medication reconciliation, thereby fostering improved patient care. Further improvement could be achieved by focusing on the medication lists of our older patients and female patients.
Sleep in elite swimmers: prevalence of sleepiness, obstructive sleep apnoea and poor sleep quality
Authors: Surda Pavol , Putala Matus , Siarnik Pavel , Walker Abigail , De Rome Katherine , Amin Nikul , Sangha Miljyot Singh , Fokkens Wytske
Published: December 2019

Objectives

Limited data suggest that swimmers might be affected by poor quality of sleep significantly. The aim was to explore the prevalence of sleep disturbances in swimmers and possible link between rhinitis and sleep disturbance.

Methods

Study 1 was an observational case–control, questionnaire-based study involving 157 elite and non-elite swimmers, 36 non-swimming athletes and 50 controls. In study 2, we measured sleep quality and duration using actigraphy in 20 elite swimmers. We also looked for presence of sleep-disordered breathing using overnight pulse oximetry monitor.

Results

In study 1, we observed a significant difference in prevalence of excessive daytime sleepiness between groups of elite swimmers and controls. Pittsburgh Sleep Quality Index (PSQI) scores do not suggest that quality of sleep in group of swimmers is impaired. In study 2, we found that prevalence of obstructive sleep apnoea (OSA) in elite swimmers defined as oxygen desaturation index ≥5 was 30%. Analysis of actigraphy data revealed that on nights prior to training days, ‘going to’ bed time was significantly earlier and total sleep time was significantly reduced.

Conclusion

Swimmers and non-swimming athletes suffer significantly more with excessive daytime sleepiness than healthy controls. In elite swimmers, this is likely linked to high prevalence of OSA. PSQI scores do not suggest that quality of sleep in group of swimmers is impaired, but actigraphy shows great variations between sleep pattern preceding training and rest day. This seems to be associated with early-morning sessions which can be a disruptive element of weekly sleep patterns.