Penetration and intracellular release of the genomes of avian RNA tumor viruses

Abstract

Attachment and penetration of avian sarcoma and leukosis viruses into chick embryo fibroblasts was examined by a combination of electron microscopy, autoradiography, and physical-chemical procedures. Adsorption occurred for 60 min at low temperature and, upon warming to 37°, disposition of virions in cells was studied in samples taken at intervals over a 120-min period. Monolayer cultures were either preserved in their undisturbed state or cells were scraped and centrifuged into pellets. In both preparations, adsorption of most viruses occurred at surfaces coated by a dense reticulum of material, termed attachment sites. Individual virions or groups of them were internalized by viropexis and moved within vacuoles to the vicinity of nuclear envelopes. Ten minutes after warming, at the time that inoculum particles were first observed in the perinuclear zone, the radioautograms indicated the occurrence of 3H-uridine-labeled viral RNA within the nucleus. Maximum concentration of this tracer was reached by 60–120 min after warming, although only about 5–10% of all cell-associated inoculum radioactivity became intranuclear. The phenol extracted nuclear labeled RNA possessed a sedimentation value of 60 S, the same as that of parental virions. Experiments in which 3H-thymidine was employed to examine by autoradiography DNA synthesis of uninfected and virus-infected cells failed to demonstrate “reverse transcriptase” activity in the cytoplasm. These observations, together with the evidence for rapid transfer of inoculum genomes into the nucleus, are best explained by assuming that virion-directed DNA transcription and reduplication occurs in the nucleus.