DNA and histone synthesis of butyrate-inhibited BSC-1 cells infected with SV40.
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Acetylation↗
Biomedical subjects
M J Fedor
Publications and source records attributed to M J Fedor.
Transcription of adenovirus cores in vitro: major RNA products differ from those made from a DNA template.
Intact cores of adenovirus type 5 were transcribed in vitro in extracts derived from uninfected HeLa cells, and the products were analyzed by hybridization to restricted viral DNA. Predominant transcripts from cores were distinctly different from transcripts generated from deproteinized DNA. The extracts contained RNA polymerases II and III. Transcription from cores was completely inhibited by low levels of alpha-amanitin, and there was little or no hybridization of products to the region of the genome encoding the small polymerase III-dependent virus-associated RNA that was a major transcript from the DNA template. The major transcripts from cores hybridized to the rightward-transcribed strand of the region between 75 and 89 map units, early region 3 of the adenovirus genome. There was several-fold more transcription of this region in cores than of the region downstream from the major late promoter, which predominates among polymerase II transcripts from DNA.
Adenoviruses, Human↗
Adenovirus chromatin structure at different stages of infection.
We investigated the structure of adenovirus deoxyribonucleic acid (DNA)-protein complexes in nuclei of infected cells by using micrococcal nuclease. Parental (infecting) DNA was digested into multimers which had a unit fragment size that was indistinguishable from the size of the nucleosomal repeat of cellular chromatin. This pattern was maintained in parenteral DNA throughout infection. Similar repeating units were detected in hamster cells that were nonpermissive for human adenovirus and in cells pretreated with n-butyrate. Late in infection, the pattern of digestion of viral DNA was determined by two different experimental approaches. Nuclear DNA was electrophoresed, blotted, and hybridized with labeled viral sequences; in this procedure all virus-specific DNA was detected. This technique revealed a diffuse protected band of viral DNA that was smaller than 160 base pairs, but no discrete multimers. All regions of the genome were represented in the protected DNA. To examine the nuclease protection of newly replicated viral DNA, infected cells were labeled with [3H]thymidine after blocking of cellular DNA synthesis but not viral DNA synthesis. With this procedure we identified a repeating unit which was distinctly different from the cellular nucleosomal repeat. We found broad bands with midpoints at 200, 400, and 600 base pairs, as well as the limit digest material revealed by blotting. High-resolution acrylamide gel electrophoresis revealed that the viral species comprised a series of closely spaced bands ranging in size from less than 30 to 250 base pairs.
Adenoviruses, Human↗
Acetylation of histone-like proteins of adenovirus type 5.
We examined acetylation of the histone-like adenovirus core proteins VII and V and the precursor of the major core protein, pVII, by measuring the incorporation of [14C]acetate. Adenovirus proteins pVII and V appeared to be acetylated, whereas protein VII was not. Label incorporated into these viral proteins in the form of acetate was metabolically stable, and labeling was not enhanced by treatment with sodium butyrate, an inhibitor of histone deacetylases. Viral protein acetylation therefore differs from the reversible acetylation of histones that has been implicated in transient alterations of chromatin structure. Inhibition of protein synthesis in infected cells resulted in a proportional reduction in [14C]acetate uptake into pVII and V, suggesting that these proteins undergo acetylation during protein synthesis and not as a post-translational modification. Therefore, these viral proteins are probably acetylated amino-terminally, a characteristic shared by three of the five major histone classes.
Acetates↗