Transactivation of the human immunodeficiency virus promoter by human herpesvirus 6 (HHV-6) strains GS and Z-29 in primary human T lymphocytes and identification of transactivating HHV-6(GS) gene fragments.

Human herpesvirus 6 (HHV-6) can activate the human immunodeficiency virus (HIV) promoter and accelerate cytopathic effects in HIV-infected human T cells. This study examines the regions of the HIV promoter required for HHV-6 transactivation in a heterogeneous population of primary human T lymphocytes with or without antigenic stimulation. Two different strains of HHV-6, GS and Z29, transactivated the HIV promoter. The GS strain transactivated the promoter in both stimulated and resting T cells, while the Z29 strain increased HIV promoter activity only in stimulated T cells. Three DNA clones containing HHV-6(GS) genomic fragments transactivated the HIV promoter in cotransfected T cells. A 21.4-kb DNA clone, pZVB70, showed the highest transactivating ability, while two other DNA fragments, pZVB10 (6.2 kb) and pZVH14 (8.7 kb), showed lower activity. One of these clones, pZVH14, activated the HIV promoter construct containing a mutation in the NF kappa B site. However, this mutated NF kappa B promoter was not transactivated during HHV-6(GS) infection or after cotransfection with pZVB70 or pZVB10. These data indicate that the NF kappa B sites of the HIV promoter are essential for its transactivation during HHV-6(GS) infection. By increasing HIV promoter activity in primary T lymphocytes, HHV-6 may consequently increase HIV replication, leading to an increase in the cytopathic effect on coinfected human T cells.     

Pyrrolines as Prodrugs of γ-Aminobutyric Acid Analogues

Abstract: Δ'-Pyrroline, S-methyl-Δ'-pyrroline, and 5.5-dimethyl-Δ'-pyrroline have been identified as substances metabolized to γ-aminobutyric acid (GABA), 4-aminopentanoic acid (raethyl GABA), and 4-amino-4-methylpen-tanoic acid (dimethyl GABA), respectively. An enzyme system residing in the soluble fraction of rabbit liver catalyzes the conversion of Δ'-pyrroline to GABA and its lactam, 2-pyrrolidinone. Acetaldehyde, allopurinol, and cyanide inhibited the reaction. Incubation of deuterium-labeled Δ'-pyrroline with mouse brain homogenates produced deuterated GABA. Mouse liver 10,000 g supernatant and mouse brain homogenates converted S-methyl-Δ'-pyrroline to methyl GABA, and 5,5-dimethyl-Δ'-pyrroline to dimethyl GABA. Four hours after intraperitoneal injection of 5-methyl-Δ'-pyrroline (200 mg/kg), methyl GABA was detected in mouse brain (0.27 μ-mol/g). Dimethyl GABA (1.21 μmol/g) was determined in mouse brain 30 min after intraperitoneal administration of 5.5-dimethyl-Δ'-pyrroline (200 mg/kg). Neither methyl GABA nor dimethyl GABA penetrated into the central nervous system when administered in the periphery. The present studies suggest that pyrrolines may represent a chemical class of brain-penetrating precursors of pharmacologically active analogues of GABA.     

Cytotoxicity studies of chromium(III) complexes on human dermal fibroblasts

The cytotoxicity of certain Cr(III) complexes, such as [Cr(salen)(H(2)O)(2)](+), [Cr(edta)(H(2)O)](-), [Cr(en)(3)](3+), [Cr(ox)(3)](3-), [Cr(pic)(3)], and CrCl(3), which differ in ionic character and ligand environment in human dermal skin fibroblasts, has been studied. After 72 h of exposure to 100 microM doses of chromium(III) complexes, the order in which the complexes had an inhibitory effect on cell viability was [Cr(en)(3)](3+) > [Cr(salen)(H(2)O)(2)](+) > [Cr(ox)(3)](3-) > [Cr(edta)(H(2)O)](-) > [Cr(pic)(3)] > CrCl(3). Based on viability studies it was confirmed that [Cr(en)(3)](3+), a triply charged cation, inhibits cell proliferation, and therefore, it was chosen to carry out further investigations. [Cr(en)(3)](3+), at a dose of 50 microM, was found to bring about surface morphological changes, evidenced by cellular blebbing and spike formation accompanied by nuclear damage. TEM analysis revealed substantial intracellular damage to fibroblasts in terms of the formation of apoptotic bodies and chromatin condensation, thus reflecting cell death. FACS analysis further revealed DNA damage by formation of a sub-G(1) peak with 84.2% DNA as aneuploid DNA and arrest of the G(2) / M phase of the cell cycle. Cellular DNA damage was confirmed by agarose gel electrophoresis with the characteristic appearance of a DNA streak in DNA isolated from [Cr(en)(3)](3+)-treated fibroblasts. The proposed mechanism suggests the plausible role of Cr(V), formed as a result of oxidation of Cr(III) by cellular oxidative enzymes, in the cytotoxic response. Consequently, any Cr(III) complex that is absorbed by cells and can be oxidized to Cr(V) must be considered a potential carcinogen. This has potential implications for the increased use of Cr(III) complexes as dietary supplements and highlights the need to consider the cytotoxicity and genotoxicity of a variety of Cr(III) complexes and to understand the potential hazards of Cr(III) complexes encountered in research laboratories.     

Strict preparation and evaluation of water-soluble hat-stacked carbon nanofibers for biomedical application and their high biocompatibility: influence of nanofiber-surface functional groups on cytotoxicity

Water-soluble H-CNFs modified with a carboxyl group possessed the ability to induce TNF-alpha, whereas CHAPS-treated H-CNFs possessed significantly greater activity and were also found to activate NF-kappaB reporter activity, to a significantly greater level than H-CNFs; furthermore the functional group modified or coated on the surface of H-CNFs was a significant cytotoxic factor that affected cell activation.     

Two new black fly species of Simulium (Simulium) (Diptera: Simuliidae) from South India

Two new black fly species, Simulium (Simulium) pothigaiense sp. n. and Simulium (Simulium) valparaiense sp. n., are described based on females, males, pupae and larvae. Simulium (Simulium) pothigaiense sp. n. is characterized by a claw simple without subbasal tooth and the base of radial vein with hair tuft in the females, maxillary palp with small sensory vesicle in the males, pupal gill with 10 short slender filaments and very short common basal stalk in the pupae and 5 hypostomal bristles per side lying parallel to lateral margin in the larvae. Simulium (Simulium) valparaiense sp. n. is characterized by hind basitarsus 6.6 times as long as its greatest width and claw with small distinct subbasal tooth in the females, gonostyle in medial view 2.3 times longer than coxite in the males, common basal stalk of pupal gill 0.31 times as long as interspiracular trunk, and larval thoracic and abdominal cuticle with pair of dorsolateral protuberances. Taxonomic notes are given to distinguish these two new species from closely related species. www.zoobank.org/urn:lsid:zoobank.org:pub:7285F6BC-607D-4F76-B6EE-9CAC67CDE651     

Diagnosis of the Earliest Strain-Specific Interactions between Tobacco Mosaic Virus and Chloroplasts of Tobacco Leaves in Vivo by Means of Chlorophyll Fluorescence Imaging

Fluorescence imaging was used to diagnose early stages of the strain-specific interactions between tobacco mosaic virus (strain PV230) and chloroplasts following infection of tobacco leaves (Nicotiana tabacum cv Xanthi). The earliest indication of interaction in tissues that ultimately become chlorotic was a reduction in chlorophyll fluorescence, and there was little fluorescence quenching compared with adjacent healthy tissues. Subsequently, fluorescence increased but remained unquenched. In the late stages fluorescence declined again in chlorotic regions as the chloroticmosaic symptoms developed. These in vivo data showing altered fluorescence yields confirm strain-specific interaction of virus coat protein with photosystem II (PSII) components in vitro, leading to photoinhibition and photooxidation of chlorophyll in infected cells and the development of visible chlorotic-mosaic symptoms. Although mechanisms leading to the low, unquenched fluorescence condition are not known, the intermediate high, unquenched fluorescence condition is consistent with impaired PSII electron transport as measured in vitro. Fluorescence lesions appear more rapidly and develop more extensively in high light, consistent with the faster and larger extent of symptom formation in high-light-grown leaves than in low-light-grown leaves.     

Marker-assisted introgression of the major bacterial blight resistance gene, <Emphasis Type="Italic">Xa21</Emphasis> and blast resistance gene, <Emphasis Type="Italic">Pi54</Emphasis> into RPHR-1005, the restorer line of the popular rice hybrid,DRRH3

The elite Indian rice hybrid, DRRH3 is highly susceptible to two major diseases, bacterial blight (BB) and blast, which limit its productivity significantly. In the present study, we have introgressed two major genes, viz., Xa21 and Pi54 conferring resistance against BB and blast, respectively into RPHR-1005, the male parent of DRRH3 through marker-assisted backcross breeding (MABB) and analyzed the backcross derived plants for their resistance against BB and blast. RPBio Patho-2 was used as a donor for both the resistance genes. Gene-specific markers were used for the foreground selection of Xa21 and Pi54 at each stage of backcrossing and markers specific for the major fertility restorer genes, Rf3 and Rf4 were used only at BC₁F₁ generation for foreground selection. Background selection was done using 62 polymorphic SSR markers and marker-assisted backcrossing was continued till BC₃ generation. At BC₃F₄, through intensive phenotype-based selections 15 promising lines (ABLs) possessing high level of resistance against BB and blast, high yield, fine-grain type, complete fertility restoration along with better panicle exsertion and taller plant type as compared to RPHR-1005 were identified and test crossed with APMS 6 A, the female parent of DRRH3. The newly derived hybrids (i.e. improved versions of DRRH3) were observed to possess high level of resistance against BB and blast along with medium-slender grain type and yield level better than or equivalent to that of DRRH3. Our study exemplifies the utility of MABB for targeted improvement of multiple traits in hybrid rice.     

Antigenic cross-reactions among herpes simplex virus types 1 and 2, Epstein-Barr virus, and cytomegalovirus.

Polyvalent rabbit antisera against herpes simplex virus type 1 and 2 (HSV-1 and HSV-2), cytomegalovirus (CMV), and Epstein-Barr virus (EBV), monospecific antisera against affinity-purified HSV-2 glycoproteins gB and gG, and a panel of monoclonal antibodies against HSV and EBV proteins were used to analyze cross-reactive molecules in cells infected with the four herpesviruses. A combination of immunoprecipitation and Western blotting with these reagents was used to determine that all four viruses coded for a glycoprotein that cross-reacted with HSV-1 gB. CMV coded for proteins that cross-reacted with HSV-2 gC, gD, and gE. Both CMV and EBV coded for proteins that cross-reacted with HSV-2 gG. Antigenic counterparts to the p45 nucleocapsid protein of HSV-2 were present in HSV-1 and CMV, and counterparts of the major DNA-binding protein and the ribonucleotide reductase of HSV-1 were present in all the viruses. The EBV virion glycoprotein gp85 was immunoprecipitated by antisera to HSV-1, HSV-2, and CMV. Antisera to CMV and EBV neutralized the infectivity of both HSV-1 and HSV-2 at high concentrations. This suggests that cross-reactivity between these four human herpesviruses may have pathogenic as well as evolutionary significance.     

Detection by monoclonal antibodies of an early membrane protein induced by Epstein-Barr virus.

Two monoclonal antibodies, E8B3 and E8D2, were raised against Epstein-Barr virus (EBV)-producing cells and were shown to immunoprecipitate a protein with an approximate molecular weight of 105,000 (p105). The protein was detectable only in EBV-containing cells which were supporting the virus lytic cycle, and its synthesis increased after cells were induced with phorbol esters. The molecule was radiolabeled and immunoprecipitated from virus-producing cells that had been extrinsically labeled with 125I, and the antibodies E8B3 and E8D2 reacted in immunofluorescence assays with infected cells; the molecule was also associated with virion particles. Synthesis of p105 was not blocked by phosphonoacetic acid and could be induced in Raji cells by superinfection with virus derived from P3HR1 cells. These data support the conclusion that p105 is an EBV-specific early membrane protein.