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Biomedical subjects

N Frenkel

Publications and source records attributed to N Frenkel.

68 records · Page 4Linked to original sources
PubMed1975

Anatomy of herpes simplex virus DNA. III. Characterization of defective DNA molecules and biological properties of virus populations containing them.

We have characterized the virus progeny and its DNA from plaque-purified and undiluted passages of herpes simplex virus 1 in HEp-2 cells. Secifically, (i) infectious virus yields declined progressively in passages 1 through 10 and gradually increased at passages 11 through 14. The yields correlated with PFU/particle ratios. (ii) In cells infected with virus from passages 6 through 10, there was an overproduction of an early viral polypeptide (no. 4) and a delay in the synthesis of late viral proteins. In addition, the virus in these passages interfered with the replication of a nondefective marker virus. Cells infected with passage 14 virus produced normal amounts of polypeptide 4 and, moreover, this virus showed minimal interfering capacity. (iii) In addition to DNA of density 1.726 g/cm-3, which was the sole component present in viral progeny of passage 0, passages 6 through 14 contained one additional species (p 1.732) and in some instances (passages 6 and 10) also DNA of an intermediate buoyant density. The ratio of p 1.732 to p 1.726 DNA increased to a maximum of 4 in passages 6 through 9 and gradually decreased to 1 in passages 10 through 14. (iv) p 1.732 DNA cannot be differentiated from p 1.726 DNA with respect to size; however, it has no Hin III restriction enzyme cleavage sites and yields only predominantly two kinds of fragments with molecular weights of 5.1 x 10-6 and 5.4 x 10-6 upon digestion with EcoRI enzyme. (v) Partial denaturation profiles of purified p 1.732 DNA from passage 14 revealed the presence of two types of tandemly repeated units corresponding roughly in size to the EcoRI fragments and situated in different molecules. (vi) In addition to the two kinds of p 1.732 molecules consisting of tandem repaeat units of different sizes, other evidence for the diversity of defective DNA molecules emerged from comparisons of specific infectivity and interfering capacity of the progeny from various passages. The data suggest that some of the particles with DNA of normal buoyant density (1.726) must also be defective since the capacity to interfere and to produce an excess of polypeptide 4 did not appear to be proportional to the amount of high-buoyant-density defective DNA. The data suggest that defective interfering particles are replaced by defective particles with diminished capacity to interfere and that more than one species of defective DNA molecules evolves on serial preparation of HSV.

Carcinoma, Squamous Cell↗
PubMed1973

Relationship between post-transcriptional adenylation of herpes virus RNA and messenger RNA abundance.

Analysis of the hybridization kinetics of labeled DNA of herpes simplex virus with unlabeled excess RNA from infected cells showed that viral RNA sequences form two classes differing in molar concentration. The abundant class constituted 93.5-99.3% of total virus-specific RNA and was complementary to 14-16% of the early DNA (2 hr after infection) and to 19-22% of the late DNA (8 hr after infection) in the reproductive cycle of the virus. The early RNA sequences were found to be a subset of the late sequences. The scarce sequences constituted 0.7-6.5% of total virus-specific RNA and were complementary to 28-30% of DNA both early and late in the reproductive cycle. In this study, abundant and scarce sequences were quantitatively separated on the basis of the finding that abundant species are adenylated, i.e., contain post-transcriptionally added poly(A), whereas the scarce RNA is not. Thus, nuclear and polyribosomal adenylated RNA were complementary to 24 and 22%, respectively, of viral DNA and, in abundance competition tests, were found to compete with each other and with abundant RNA from infected cells after 8 hr. The nonadenylated polyribosomal RNA was complementary to 27% of total viral DNA of which 6% was also complementary to adenylated polyribosomal RNA. Hybridization kinetics indicated that each of the fractionated adenylated RNA formed two classes complementary to 6 and 21% of viral DNA.

Adenine Nucleotides↗
PubMed1973

RNA synthesis in cells infected with herpes simplex virus. VII. Control of transcription and of transcript abundancies of unique and common sequences of herpes simplex virus 1 and 2.

Analysis of the kinetics of hybridization in liquid of labeled herpes simplex virus (HSV) 1 and 2 DNAs with excess unlabeled RNA extracted at 2 (early) and 8 (late) h postinfection revealed the following. (i) The RNA transcripts present in the HSV-1-infected cells at 2 and 8 h postinfection are complementary to 44 and 48% of HSV-1 DNA. The RNA transcripts present in the HSV-2-infected cells at 2 and 8 h postinfection are complementary to 21 and 50% of HSV-2 DNA. (ii) The transcripts present in 2-h HSV-1- or HSV-2-infected cells treated with cycloheximide are complementary to 44 and 45% of the respective DNAs. (iii) The RNA transcripts present in the HSV-1-infected cells at 2 h postinfection and in HSV-2-infected cells at 8 h postinfection form 2 classes, abundant and scarce, differing in molar concentrations. The RNA transcripts present in the HSV-2-infected cells at 2 h postinfection form only one abundance class. (iv) The transcripts present in the HSV-1-infected cells at 8 h postinfection are complementary to 24% of HSV-2 DNA and therefore 50% of the transcribed HSV-1 sequences are shared by the two viruses. Of the RNA sequences complementary to HSV-2 DNA, 13% arise from HSV-1 templates specifying abundant RNA and 11% arise from HSV-1 templates specifying scarce RNA. Thus, the DNA sequences shared in common by HSV-1 and HSV-2 DNAs constitute 71% of the HSV-1 templates specifying abundant RNA and 39% of sequences specifying scarce RNA.

Base Sequence↗
PubMed1972

A DNA fragment of Herpes simplex 2 and its transcription in human cervical cancer tissue.

A human cervical tumor, free of detectable infectious herpes simplex 2 virus, contained a fragment comprising 39% of herpes viral DNA. Renaturation kinetics indicate that an average of 1 to 3.5 DNA fragments of herpes simplex virus are present per cell, depending on the ploidy of the cells in this particular tumor. Virus-specific sequences were found linked to highly repetitive sequences of host DNA, which reassociated under conditions designed to preclude reassociation of viral sequences. The tumor also contained RNA transcripts complementary to 5% of the viral DNA. The fraction of viral DNA template transcribed in the cervical tumor is considerably less than that transcribed in productively infected cells (50%).

DNA, Neoplasm↗
PubMed1972

Ribonucleic acid synthesis in cells infected with herpes simplex virus: controls of transcription and of RNA abundance.

Analysis of the kinetics of hybridization in liquid of labeled herpes simplex virus-1 DNA and excess viral RNA revealed the following: (i) Cells infected by herpes simplex virus-1 for 2 hr (before DNA synthesis) contain two classes of RNA molecules differing 140-fold in molar concentrations. The abundant and scarce RNAs are transcribed from 14 and 30% of the DNA, respectively. RNA extracted at 8 hr after infection (late RNA) also contains abundant and scarce classes differing 40-fold in molar concentrations; these are transcribed from 19 and 28% of viral DNA, respectively. Abundance competition hybridization tests indicate that the abundant RNA at 2 hr is a subset of the 8-hr abundant RNA. (ii) The abundant RNAs probably specify structural proteins, as indicated by estimates of DNA template required for structural proteins and by experiments showing that 19 of 24 proteins (corresponding to 68% of genetic information for structural proteins) are already made between 0.5 and 2 hr after infection. We conclude that there are two types of transcriptional controls, i.e., on-off and abundance controls, and that the synthesis of most structural components is an early viral function.

Amino Acids↗
PubMed1972

Separation of the herpesvirus deoxyribonucleic acid duplex into unique fragments and intact strand on sedimentation in alkaline gradients.

Deoxyribonucleic acid (DNA) extracted from herpes simplex virions forms multiple partially overlapping bands upon denaturation and centrifugation in alkaline sucrose density gradients. The most rapidly sedimenting DNA corresponds to an intact strand 48 x 10(6) daltons in molecular weight. In this study, we analyzed the DNA fragments generated in alkaline sucrose gradients with respect to size and uniqueness of base sequences. The distribution of sedimentation constants of the various fragments obtained in numerous gradients showed that the fragments smaller than the whole strand fall into six distinct classes ranging in molecular weight from 10 x 10(6) to 39 x 10(6) daltons. Four types of DNA strands can be reconstructed from the whole strand and six fragments on the basis of their molecular weights. DNA from each of the bands self-hybridizes to a lower extent than unfractionated viral DNA, indicating that each of the bands preferentially contains sequences from one unique strand. The data permit reconstruction of four possible types of DNA duplexes differing in the positions of the strand interruptions. Analysis of viral DNA extracted from nuclei of cells labeled with (3)H-thymidine for intervals from 3 to 120 min showed that nascent DNA is invariably attached to small fragments and that the fragments become elongated only upon prolonged incubation of cells. The experiments suggest that viral DNA replication begins at numerous initiation sites along each strand and that the elongation beyond the size of the replication unit involves repair or ligation, or both. Since newly made DNA yields more fragments than viral DNA extracted from mature virions, it is suggested that the fragmentation of mature DNA on denaturation with alkali arises from incomplete processing of specific initiation sites. Comparison of viral DNA extracted from nuclei with that extracted from mature cytoplasmic virions in cells labeled for 120 min indicates that packaged DNA is not randomly selected from among the nuclear DNA population but rather represents DNA molecules which in alkaline gradients yield a minimal number of fragments.

Base Sequence↗
PubMed1971

Herpes vimplex virus: genome size and redundancy studied by renaturation kinetics.

Herpes simplex virus subtype 1 deoxyribonucleic acid (DNA) was sheared in a French press to uniform fragments, denatured by heating, then allowed to reassociate. The renaturation reaction followed second-order kinetics with a single rate constant indicating that at least 95% of the genome was unique and that repetitive sequences, if present, were not detectable by this technique. The kinetic complexity of the herpes simplex genome was determined by DNA renaturation kinetics to be (95 +/- 1) x 10(6) daltons. Since this value is in excellent agreement with the molecular weight of viral DNA [(99 +/- 5) x 10(6) daltons] obtained from velocity sedimentation studies, it is concluded that virions contain only one species of double-stranded DNA molecules 95 x 10(6) to 99 x 10(6) daltons in molecular weight.

Base Sequence↗