ABSTRACT
cIAP1 is an important member of the inhibitor of apoptosis family of proteins and is involved in the regulation of the NF-kappaB-signalling pathway downstream of the TNF receptor. We report here that UV irradiation leads to downregulation of cIAP1 expression because of enhanced cIAP1 mRNA destabilization. An AU-rich element located within the 3' untranslated region of cIAP1 mRNA is sufficient to mediate cIAP1 mRNA instability. Furthermore, we have identified hnRNP A1 as a cIAP1 3'UTR-binding protein. hnRNP A1 is a primarily nuclear protein, but accumulates in the cytoplasm after exposure of cells to UV irradiation. Indeed, we find that hnRNP A1 enhances the destabilization of cIAP1 mRNA during UV irradiation. Moreover, siRNA-mediated knockdown of hnRNP A1 restores cIAP1 levels and prevents UV irradiation-induced activation of the NF-kappaB signal transduction pathway, suggesting that hnRNP A1 is an essential post-transcriptional modulator of cIAP1 expression, and thus cIAP1 activity.
Subject(s)
Heterogeneous-Nuclear Ribonucleoprotein Group A-B/metabolism , Inhibitor of Apoptosis Proteins/genetics , NF-kappa B/metabolism , RNA Stability , RNA, Messenger/metabolism , 3' Untranslated Regions/metabolism , Cell Line , Heterogeneous Nuclear Ribonucleoprotein A1 , Humans , Inhibitor of Apoptosis Proteins/metabolism , Inhibitor of Apoptosis Proteins/radiation effects , RNA, Messenger/radiation effects , RNA, Small Interfering , Receptors, Tumor Necrosis Factor/metabolism , Signal Transduction , Ultraviolet RaysABSTRACT
The sources of the induced activity from the d(48.5)+Be fast neutron therapy beam of the Harper Hospital superconducting cyclotron have been investigated. The distribution of activity in the treatment room was measured, and the levels of dose equivalent to the staff were established. Activation spectra were measured with a high purity RE-Ge detector. Peaks corresponding to 28Al, 56Mn, 24Na, 64Cu, 66Cu, and 187W were present in the spectra. The dose equivalents due to the induced activation were measured by means of an ionization chamber type survey meter at six locations in the room. Irradiations of 120 monitor units were given at 15-min intervals, thus simulating the clinical situation. The measurements were made between the irradiations. The highest levels were registered around the treatment head. Two patterns are clearly distinguishable in these measurements. A fast decaying component with approximately 2 min half-life can be ascribed predominantly to 28Al and a slow growing component reaching saturation after about 4-5 treatments is associated with the presence of 56Mn. For uniform treatment load the activation build-up in each location was similar every day of the week with minimal values measured after the week end shut down. Personnel monitoring is achieved with dosimeters capable of detecting an extended range of neutron energies as well as beta rays and photons. Correlation between the number of fields treated and the doses to the radiation therapy technologists was shown. The mean dose equivalent received by the therapists is 7.1 +/- 0.2 microSv per treatment field. Means of reducing personnel dose equivalent levels are proposed.
Subject(s)
Cyclotrons , Fast Neutrons , Occupational Exposure , Radiation Monitoring/methods , Hospitals , Humans , Neutron Capture Therapy/instrumentation , Personnel, Hospital , Radiation Dosage , Time FactorsABSTRACT
We have studied the reactions of hamster embryo cells transformed by ultraviolet-inactivated herpes simplex type 2 (333-8-9 T cells) to infections with adeno-associated satellite virus (AAV) and its adenovirus helpers. Resident HSV structural antigens were not detectable in early or late passage of 333-8-9 T cells. AAV structural antigens were not detected in these cells unless the cells were coinfected with a helper adenovirus. In early passage 333-8-9 T cells were permissive to infections with simian adenovirus SV15 whereas normal hamster cell line LSH was nonpermissive. In some late passages of 333-8-9 T cells infections with SV15 adenovirus led to the production of viruslike particles whose morphology was identical with reoviruses.
Subject(s)
Adenoviridae , Cell Transformation, Neoplastic , Dependovirus , Simplexvirus , Adenoviruses, Human , Adenoviruses, Simian , Animals , Antigens, Viral , Cricetinae , Dependovirus/immunology , Genes, Viral , Helper Viruses , Inclusion Bodies, Viral , Microscopy, Electron , Simplexvirus/genetics , Simplexvirus/immunologyABSTRACT
The replication of rodent parvovirus X14 DNA has been studied in rat embryo tissue culture cells. Virus DNA was isolated from I M-NaCl-SDS-pronase supernatant fluids from 24 h after infection. The majority of this DNA was 1-7 mum in length and double-stranded, indicating that it was an intermediate in the replication cycle of this single-stranded DNA virus. Single-stranded DNA of equivalent length was isolated directly from X14 virions. The buoyant density of this DNA was 1-728 g/ml whereas the double-stranded form banded at 1-714 g/ml in caesium chloride gradients. Difficulties in detecting significant amounts of single-stranded viral DNA directly from infected cells would appear to indicate that progeny single-stranded DNA is rapidly encapsidated after synthesis.
Subject(s)
DNA Replication , DNA, Single-Stranded/biosynthesis , DNA, Viral/biosynthesis , Parvoviridae/metabolism , Culture Techniques , DNA, Single-Stranded/analysis , DNA, Viral/analysis , Parvoviridae/analysis , Parvoviridae/growth & development , Virus ReplicationABSTRACT
Infectious DNA from adeno-associated satellite virus (ASV) has been isolated from cells coinfected with a temperature-sensitive mutant of herpes simplex virus (HSV) type 1 in the absence of contaminating HSV DNA. This satellite virus DNA does not appear to differ in its physical, chemical and biological properties from DNA isolated directly from virions or from cells co-infected with adenovirus. The DNA is double-stranded with a buoyant density of 1.718 gm/cm(3). It sediments at 16S in both neutral and alkaline sucrose gradients. Single-stranded DNA from alkaline sucrose gradients has a modal length of 1.5 mum and demonstrates evidence of internal redundancies in the electron microscope.
Subject(s)
DNA, Viral/isolation & purification , Dependovirus/chemistry , Simplexvirus/physiology , Animals , Centrifugation, Density Gradient , Chlorocebus aethiops , DNA, Single-Stranded/analysis , DNA, Single-Stranded/isolation & purification , DNA, Viral/analysis , Dependovirus/physiology , Simplexvirus/genetics , Vero Cells , Virion/chemistryABSTRACT
Adeno-associated satellite virus type 4, obtained by repeated undiluted passage, failed to produce distinct bands at the expected density of 1.43 g/cm(3) after density gradient centrifugation in CsCl. This phenomenon occurred regardless of the hemagglutinating activity of the starting material. Sharp bands were found at a density of 1.34 to 1.35 g/cm(3). These bands contained adenovirions and numerous satellite particles. These latter particles could be distinguished by electron microscopy from standard dense satellite particles by their flattened profiles and deep penetration of negative stains. Dense bands of satellite virus at 1.43 g/cm(3) were constantly observed when the inoculum was comprised of highly diluted seed virus. Light satellite particles had a particle to HA ratio comparable with dense particles, but possessed low infectivity. Measurements of contour lengths of extracted deoxyribonucleic acid (DNA) indicate that light particles contain only a small amount of DNA, possibly less than 0.5 x 10(6) daltons, compared to 1.4 x 10(6) for the complete satellite DNA molecule.
Subject(s)
Adenoviridae/growth & development , Animals , Cell Line , Centrifugation, Density Gradient , Culture Techniques , DNA, Viral/metabolism , Freezing , Haplorhini , Hemagglutination Tests , Kidney , Microscopy, Electron , Satellite Viruses/growth & development , Virus CultivationABSTRACT
A microdiffusion technique, developed for visualization of nucleic acid molecules in the electron microscope, requires less than 0.01 microgram of nucleic acid. Although originally developed for free nucleic acids, the method can be applied to virion suspensions for direct visualization of their genomes; less than 10(10) virions per milliliter are required. Results agree well with those yielded by the diffusion technique of Lang, Kleinschmidt, and Zahn.