Two regulatory isozymes of glutamine synthetase from Bacillus caldolyticus, an extreme thermophile.

Two glutamine synthetases (EC 6.3.1.2) have been purified to homogeneity from $\text{B. caldolyticus}$ strain YTP, grown at 70° on a minimal, defined medium. The enzymes are virtually identical in size and molecular weight (12 sub-units of MW 50,000), but differ in their isoelectric points, electrophoretic mobility, net charge, inherent thermal stability, affinity for substrates, and activity responses to metal ions and pH. Of primary interest is the observation that the more acidic form (pI = 5.2), E_I, is strongly feedback-regulated by certain amino acids derived from glutamine (Gly, L-Ala, L- and D-ser) but not by L-Glu or AMP, whereas the less acidic form (pI = 5.5), E_II, is inhibited most strongly by L-Gln and AMP, but not by the above amino acids. Both enzymes are inhibited strongly by ADP, CTP, NAD, glucosamine-6-P, less strongly by nucleotide diphosphates and L-Trp, and are activated by nucleotide monophosphates other than AMP. These results suggest that overall regulation of glutamine synthetase by the full spectrum of end product metabolites derived from L-Gln is accomplished by regulatory isozymes in this extremely thermophilic organism.     

The Epstein-Barr virus BRLF1 immediate-early gene product transactivates the human immunodeficiency virus type 1 long terminal repeat by a mechanism which is enhancer independent.

The Epstein-Barr virus (EBV) immediate-early gene product, BRLF1, transactivates the human immunodeficiency virus type 1 (HIV-1) long terminal repeat. BRLF1-induced transactivation of HIV-1 promoter constructs is accompanied by an increase in plasmid mRNA and is reporter gene independent. Previously, BRLF1 transactivation of EBV promoters has been mapped to regions which function as enhancer elements. Deletional analysis demonstrates that BRLF1 transactivation of the HIV-1 promoter does not require the HIV-1 enhancer. Thus, the EBV BRLF1 gene product may transactivate by at least two different mechanisms, one mechanism involving certain enhancer elements and another mechanism which is enhancer independent.     

Coexisting angiomyolipoma and renal cell carcinoma in a kidney of an elderly woman: case report and review of the literature

Angiomyolipoma is a well described but relatively uncommon benign renal neoplasm composed of varying admixtures of mature adipose tissue, smooth muscle, and thick-walled blood vessels. The incidence of angiomyolipoma is about 0.3% overall. It frequently occurs in patients with tuberous sclerosis. Even more uncommon is the simultaneous occurrence of angiomyolipoma and renal cell cancer in the same kidney in a patient without tuberous sclerosis.     

Evidence for an active site arginine in UDP-glucuronyltransferase

2,3-Butanedione inactivates the pure form of UDP-glucuronyltransferase used in these experiments (GT2P) (EC 2.4.1.17) purified from pig liver microsomes. The kinetics of the reaction indicates that 2,3-butanedione reacts with two amino acids that affect activity. A rapid, partial inactivation is followed by a slower rate of inactivation that leads eventually to completely inactive enzyme. UDP-glucuronic acid and glucuronic acid, as compared with UDP, are effective as protectors against the slow, secondary phase of inactivation; no ligand tested protected against the rapid phase of inactivation. The lipid environment of GT2P was a determinant of the pseudo-first order rate constant for the slow phase of inactivation, but did not affect the rate of the rapid phase of inactivation. The data suggest that GT2P contains an active site arginine that interacts with the -COO- at C-6 of the glucuronic acid moiety of UDP-glucuronic acid.     

Hormone-regulated expression and distribution of versican in mouse uterine tissues

Background  

Remodeling of the extracellular matrix is one of the most striking features observed in the uterus during the estrous cycle and after hormone replacement. Versican (VER) is a hyaluronan-binding proteoglycan that undergoes RNA alternative splicing, generating four distinct isoforms. This study analyzed the synthesis and distribution of VER in mouse uterine tissues during the estrous cycle, in ovariectomized (OVX) animals and after 17beta-estradiol (E2) and medroxyprogesterone (MPA) treatments, either alone or in combination.     

Dual miRNA Targeting Restricts Host Range and Attenuates Neurovirulence of Flaviviruses

Mosquito-borne flaviviruses are among the most significant arboviral pathogens worldwide. Vaccinations and mosquito population control programs remain the most reliable means for flavivirus disease prevention, and live attenuated viruses remain one of the most attractive flavivirus vaccine platforms. Some live attenuated viruses are capable of infecting principle mosquito vectors, as demonstrated in the laboratory, which in combination with their intrinsic genetic instability could potentially lead to a vaccine virus reversion back to wild-type in nature, followed by introduction and dissemination of potentially dangerous viral strains into new geographic locations. To mitigate this risk we developed a microRNA-targeting approach that selectively restricts replication of flavivirus in the mosquito host. Introduction of sequences complementary to a mosquito-specific mir-184 and mir-275 miRNAs individually or in combination into the 3’NCR and/or ORF region resulted in selective restriction of dengue type 4 virus (DEN4) replication in mosquito cell lines and adult Aedes mosquitos. Moreover a combined targeting of DEN4 genome with mosquito-specific and vertebrate CNS-specific mir-124 miRNA can silence viral replication in two evolutionally distant biological systems: mosquitoes and mouse brains. Thus, this approach can reinforce the safety of newly developed or existing vaccines for use in humans and could provide an additional level of biosafety for laboratories using viruses with altered pathogenic or transmissibility characteristics.