cHerpes simplex virus 2 (HSV-2) is an important human pathogen that is the major cause of genital herpes infections and a significant contributor to the epidemic spread of human immunodeficiency virus infections. The UL21 gene is conserved throughout the Alphaherpesvirinae subfamily and encodes a tegument protein that is dispensable for HSV-1 and pseudorabies virus replication in cultured cells; however, its precise functions have not been determined. To investigate the role of UL21 in the HSV-2 replicative cycle, we constructed a UL21 deletion virus (HSV-2 ⌬UL21) using an HSV-2 bacterial artificial chromosome, pYEbac373. HSV-2 ⌬UL21 was unable to direct the production of infectious virus in noncomplementing cells, whereas the repaired HSV-2 ⌬UL21 strain grew to wild-type (WT) titers, indicating that UL21 is essential for virus propagation. Cells infected with HSV-2 ⌬UL21 demonstrated a 2-h delay in the kinetics of immediate early viral gene expression. However, this delay in gene expression was not responsible for the inability of cells infected with HSV-2 ⌬UL21 to produce virus insofar as late viral gene products accumulated to WT levels by 24 h postinfection (hpi). Electron and fluorescence microscopy studies indicated that DNA-containing capsids formed in the nuclei of ⌬UL21-infected cells, while significantly reduced numbers of capsids were located in the cytoplasm late in infection. Taken together, these data indicate that HSV-2 UL21 has an early function that facilitates viral gene expression as well as a late essential function that promotes the egress of capsids from the nucleus.

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To cite this article: Feric NT, Boffa MB, Johnston SM, Koschinsky ML. Apolipoprotein(a) inhibits the conversion of Glu-plasminogen to Lys-plasminogen: a novel mechanism for lipoprotein(a)-mediated inhibition of plasminogen activation. J Thromb Haemost 2008; 6: 2113-20.Summary. Background: Elevated plasma concentrations of lipoprotein(a) [Lp(a)] are associated with an increased risk for thrombotic disorders. Lp(a) is a unique lipoprotein consisting of a low-density lipoprotein-like moiety covalently linked to apolipoprotein(a) [apo(a)], a homologue of the fibrinolytic proenzyme plasminogen. Several in vitro and in vivo studies have shown that Lp(a)/apo(a) can inhibit tissue-type plasminogen activator-mediated plasminogen activation on fibrin surfaces, although the mechanism of inhibition by apo(a) remains controversial. Essential to fibrin clot lysis are a number of plasmin-dependent positive feedback reactions that enhance the efficiency of plasminogen activation, including the plasminmediated conversion of Glu-plasminogen to Lys-plasminogen. Objective: Using acid-urea gel electrophoresis to resolve the two forms of radiolabeled plasminogen, we determined whether apo(a) is able to inhibit Glu-plasminogen to Lys-plasminogen conversion. Methods: The assays were performed in the absence or presence of different recombinant apo(a) species, including point mutants, deletion mutants and variants that represent greater than 90% of the known apo(a) isoform sizes. Results: Apo(a) substantially suppressed Glu-plasminogen conversion. Critical roles were identified for the kringle IV types 5-9 and kringle V; contributory roles for sequences within the aminoterminal half of the molecule were also observed. Additionally, with the exception of the smallest naturally-occurring isoform of apo(a), isoform size was found not to contribute to the inhibitory capacity of apo(a). Conclusion: These findings underscore a novel contribution to the understanding of Lp(a)/apo(a)-mediated inhibition of plasminogen activation: the ability of the apo(a) component of Lp(a) to inhibit the key positive feedback step of plasmin-mediated Glu-plasminogen to Lys-plasminogen conversion.

The Us2 gene encodes a tegument protein that is conserved in most members of the Alphaherpesvirinae. Previous studies on the pseudorabies virus (PRV) Us2 ortholog indicated that it is prenylated, associates with membranes, and spatially regulates the enzymatic activity of the MAP (mitogen-activated protein) kinase ERK (extracellular signal-related kinase) through direct binding and sequestration of ERK at the cytoplasmic face of the plasma membrane. Here we present an analysis of the herpes simplex virus 2 (HSV-2) Us2 ortholog and demonstrate that, like PRV Us2, HSV-2 Us2 is a virion component and that, unlike PRV Us2, it does not interact with ERK in yeast two-hybrid assays. HSV-2 Us2 lacks prenylation signals and other canonical membrane-targeting motifs yet is tightly associated with detergent-insoluble membranes and localizes predominantly to recycling endosomes. Experiments to identify cellular proteins that facilitate HSV-2 Us2 membrane association were inconclusive; however, these studies led to the identification of HSV-2 Us2 as a ubiquitin-interacting protein, providing new insight into the functions of HSV-2 Us2.

To investigate the value of externally available computer‐based bibliographic retrieval services in agriculture, 100 retrospective searches in response to real requests for information were carried out by computer. Seventy‐five of these were paralleled using conventional manual methods and the results were analysed and compared for efficiency, cost and staff time required. The databases most frequently used were CAIN (now AGRICOLA), BIOSIS and Chemical Abstracts Condensates (searched online using Lockheed Dialog), and Medline. Online searches took one‐sixth of the staff time required for manual searching and cost much the same. Online searches tended to have higher relative recall and lower precision than manual searches of the same database. The differences in performance are due to the different entry points available in the printed and machine‐readable forms of a database. Detailed knowledge of these will enable a choice to be made of the more efficient method of searching when both manual and online searching of a database is possible. Online searches were well received by users and their use could increase the output of library and information staff engaged in retrospective searching.