Mutants in a conserved region near the carboxy-terminus of HIV-1 Rev identify functionally important residues and exhibit a dominant negative phenotype

The rev protein (Rev) of human immunodeficiency virus increases the cytoplasmic expression of viral structural gene mRNAs. We had previously reported the existence of a region (residues 73-98) near the carboxy-terminus in HIV-1 Rev essential for its function. To further define the structural elements in this region, we examined the effects of substitution mutations in highly conserved residues in this region, between amino acids 75-81, on Rev function. Mutations in Pro76-77 and Arg80 retained Rev function, whereas those in Leu75 and Leu81 abolished Rev activity and exhibited trans-dominant suppression of wt Rev function. The Leu81 mutation, in particular, exhibited an efficient dominant negative phenotype. Leu75 and Leu81 thus appear to define residues essential to the Rev "effector" function.     

Statistical Optimization of Bioprocess Parameters for Improved Production of L-Asparaginase from Lactobacillus plantarum

Proceedings of the National Academy of Sciences, India Section B: Biological Sciences - L-asparaginase (ASNase), a tetrameric enzyme, holds comprehensive applications in food industries as a...     

Evidence that a second tumor antigen coded by adenovirus early gene region E1a is required for efficient cell transformation.

The expression of the adenovirus (Ad) early coding region 1a (E1a) is required for virus-induced cell transformation and for the activation of other viral early genes and some cellular genes. Two overlapping early mRNAs of 13S and 12S that are transcribed from this region code for a 289-amino acid protein and a 243-amino acid protein, respectively. Earlier studies have shown that the 289-amino acid protein is essential for cell transformation. We have constructed an Ad type 2 (Ad2) deletion mutant (dl231) in which the intervening sequence for the 13S mRNA is precisely removed. Mutant dl231 is completely viable in human KB cells and produces normal amounts of 13S mRNA but much reduced amounts of a defective 12S mRNA. Mutant dl231 induces focal transformation of established rat embryo fibroblasts at a frequency one-fifth to one-half that of wild-type virus. However, the transformed cells are defective in their ability to form anchorage-independent colonies on semisolid medium. Therefore, our results demonstrate that the 243-amino acid protein is required for full transformation of rat embryo cells.     

Pangolin Indexing System: implications in forensic surveillance of large seizures

International Journal of Legal Medicine - Demand for pangolin scales in East Asia has increased dramatically in the past two decades, raising concern to the pangolin survival and bringing them to...     

Singapore Neonatal Resuscitation Guidelines 2021

Neonatal resuscitation is a coordinated, team-based series of timed sequential steps that focuses on a transitional physiology to improve perinatal and neonatal outcomes. The practice of neonatal resuscitation has evolved over time and continues to be shaped by emerging evidence as well as key opinions. We present the revised Neonatal Resuscitation Guidelines for Singapore 2021. The recommendations from the International Liaison Committee on Resuscitation Neonatal Task Force Consensus on Science and Treatment Recommendations (2020) and guidelines from the American Heart Association and European Resuscitation Council were compared with existing guidelines. The recommendations of the Neonatal Subgroup of the Singapore Resuscitation and First Aid Council were derived after the work group discussed and appraised the current available evidence and their applicability to local clinical practice.     

Functional relatedness between the E1a and E1b regions of group C and group D human adenoviruses

The functional relatedness of the transforming genes (E1a and E1b) of adenovirus type 9 (group D) which induces mammary tumors in rats and those of the non-tumorigenic adenoviruses, Ad2 and Ad5 (group C) was examined. Transfection of established rat embryo cells with a DNA segment containing the E1a and E1b regions of Ad9 resulted in efficient transformation; similar results have been shown for group A, B and C Ads. In contrast to Ads of group A, B and C, Ad9 DNA containing the E1 region or the entire viral genome was unable to transform primary baby rat kidney (BRK) cells. The functional relatedness of genes encoded within the E1 region was compared using a mutant complementation assay in which various group C mutants defective in the entire E1 region or in the E1a or E1b regions alone as well as mutants defective exclusively within the 19K or 58K T antigens coding regions of E1b were coinfected with wild type (wt) Ad9 and tested for group C mutant DNA replication, virus production, or expression of early and late genes. These studies have shown that a defect in the entire E1 region of Ad2 could only be complemented poorly by Ad9; our earlier studies have shown that coinfection with Ad12 (group A) or Ad7 (group B) resulted in efficient complementation (Brusca and Chinnadurai (1981) J. Virol. 39, 300-305). Further analysis indicated that a defect in the E1a region could be complemented by the group D E1a region. The level of E1a complementation as judged by mutant DNA replication and activation of expression of mutant early viral genes was about one-fourth to one-fifth the level in 293 cells that constitutively express Ad5 E1a and E1b regions. Our results indicate that a defect in the E1b 19K T antigen, which leads to degradation of intracellular DNA in infected cells, could be complemented by the group D protein. However, a defect in the E1b 58K T antigen could not be efficiently complemented by the group D protein. Coinfection of group C mutants defective in the 58K T antigen and Ad9 wt did not lead to an increase in the mutant viral production. Furthermore, in cells coinfected with the 58K T antigen mutants and Ad9 wt there was a large reduction in the accumulation of group C late cytoplasmic RNA. The observed complementation defect of Ad9 in supporting multiplication of group C mutants defective in the entire E1 region may therefore be a cumulative effect of both E1a and E1b regions.     

Protective effect of Cardiospermum halicacabum leaf extract on glycoprotein components on STZ–induced hyperglycemic rats

ObjectiveTo investigate the protective role of Cardiospermum halicacabum (C. halicacabum) leaf extract on glycoprotein metabolism in streptozotocin (STZ)-induced diabetic rats.MethodsDiabetes was induced in male albino Wistar rats by intraperitonial administration of STZ. The C. halicacabum leaf extract (CHE) was administered orally to normal and STZ–diabetic rats for 45 days. The effects of C. halicacabum leaf extract (CHE) on plasma and tissue glycoproteins (hexose, hexosamine, fucose and sialic acid) were determined.ResultsThe levels of plasma and tissues glycoproteins containing hexose, hexosamine and fucose were significantly increased in STZ–induced diabetic rats. In addition, the level of sialic acid significantly increased in plasma and liver while decreased in kidney of STZ–induced diabetic rats. After administration of CHE to diabetic rats, the metabolic alteration of glycoprotein reverted towards normal levels.ConclusionsThe present study indicates that the CHE possesses a protective effect on abnormal glycoprotein metabolism in addition to its antihyperglycemic activity.     

Functional characterisation of bioactive peptide derived from terrestrial snail Cryptozona bistrialis and its wound‐healing property in normal and diabetic‐induced Wistar albino rats

A peptide might be an exciting biomaterial or template for the development of novel wound‐healing agents. In this report, it was isolated from the terrestrial snail Cryptozona bistrialis by enzymatic digestion and was evaluated for its in vitro wound‐healing activity in NIH/3T3 mouse fibroblasts cell line and in vivo wound‐healing activity in normal and diabetic‐induced Wistar albino rats. The C. bistrialis protein was digested by the papain enzyme, and 21.79 kDa peptide (Cb‐peptide) was purified by reversed‐phase high‐performance liquid chromatography and identified by MALDI (matrix‐assisted laser desorption/ionization)‐TOF analysis. The isolated Cb‐peptide was characterised by various analytical methods. The peptide demonstrated a capacity to prevent the development of pathogenic bacterial and fungal cultures and proved that it promotes significant wound‐healing activity in the wound scratch assay method by rapid cell migration and closure of wound. Isolated Cb‐peptide was lyophilised and formulated to ointment and analysed for in vivo wound‐healing activity in normal and diabetic (alloxan monohydrate)‐induced Wistar albino rats. Cb‐peptide ointment‐treated groups showed a greater degree of wound healing and early and complete period of epithelialisation in normal and diabetic‐induced Wistar albino rats. Cb‐peptide ointment‐treated groups showed significant excision and incision wound‐healing activity. A conclusion was reached that the peptide isolated from C. bistrialis showed greater wound‐healing activity compared with vehicle control and standard control.     

PLDLS-dependent interaction of E1A with CtBP: regulation of CtBP nuclear localization and transcriptional functions

C-terminal binding proteins (CtBPs) are cellular corepressors that are targeted by adenovirus E1A. A conserved motif of E1A (PLDLS) interacts with an N-terminal hydrophobic cleft of CtBPs. Many cellular cofactors also interact with CtBPs through PLDLS-like motifs. E1A interaction with CtBP2 changed the composition of the CtBP2 protein complex and enhanced CtBP2 acetylation. We have identified a mutant of CtBP2 (M48A) that fails to interact with cellular cofactors while interacting normally with E1A. Other cleft mutations in CtBP2 affected interaction of both cellular cofactors and E1A. The M48A mutant did not repress the cellular E-cadherin promoter but inhibited transactivation mediated by the E1A N-terminal region through interaction with the E1A PLDLS motif. In vitro, E1A enhanced CtBP2 acetylation by p300 via a mechanism involving dissociation of acetylated CtBP2 from p300. E1A enhanced nuclear localization of CtBP1 as well as a cytoplasmically localized acetylation-deficient mutant of CtBP2 (3KR-CtBP2) through PLDLS-dependent interaction. Chromatin immunoprecipitation assays revealed presence of CtBP2 on E-cadherin and c-fos promoters. While E1A did not significantly alter targeting of CtBP2 to the E-cadherin and c-fos promoters, it dramatically enhanced promoter targeting of 3KR-CtBP2. Our results raise a possibility that E1A may gain access to cellular promoters through PLDLS-dependent interaction with CtBPs.     

Adenovirus E1a as a tumor-suppressor gene.

The E1a gene of group C adenoviruses is one of the most studied transforming genes of DNA tumor viruses. These transforming genes have been conventionally considered as dominant oncogenes since they transform cells in vitro and many of the resulting transformed cell lines induce tumors in experimental animals. It now appears that, in addition to its well-known transforming activities, E1a possesses activities which suppress transformation, tumorigenesis and malignant progression (metastasis) of tumor cells. Thus, E1a appears to meet the definition of both a dominant oncogene and a tumor-suppressor gene.