Retroviruses and insect cells

• Main Author: Heine, Christopher
• Published: University of Warwick 1982
• Subjects: 579; QR355 Virology
• Online Access: https://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.330499

Several Invertebrate cell lines were examined for the presence of retrovirus particles. When cells of a Drosophila melanogaster cell line were disrupted and analysed on sucrose density gradients, a subcellular fraction with a density of 1.22 g/ml was found to possess endogenous DNA polymerase activity and could catalyse polymerization of deoxynucleotide triphosphates in response to added template primers. The latter activity had the cation and template, primer responses expected for reverse transcriptase. A high molecular weight polyadenylic acid-containing RNA was also purified from this fraction and could be dissociated by heat treatment into 30 to 35S and smaller species. Electron microscopy revealed the presence of toroidal forms reminiscent of intracytoplasmic A-type retrovirus particles within the Drosophila cells. Similar forms were found associated with the subcellular fraction of 1.22 g/ml. Drosophila cells could be induced by halogenated pyrimidines to release a polymerase activity capable of utilizing the template, primer poly(rA).oligo(dT). Pellets were prepared from induced cell culture fluids and analysed on sucrose gradients. Two peaks of poly(rA).oligo(dT) utilization were obtained of density 1.14 and 3,.20 g/ml. Radiolabelled RNA was prepared from these fractions. Radiolabelled material from both 1.14 and 1.20 g/ml density fractions sedimented at a position corresponding to 60-70S molecular weight RNA. Those high molecular weight RNAs could be dissociated by heat treatment into 30-35S species. Infection of insect cells with known Avian and Murine retroviruses was carried out. No positive results could be obtained by using polymerase activity as a measure of replication. By using Southern blot analysis, it was found that Drosophila cells could acquire exogenous viral sequences from XLV (KMSV). A number of cell lines were treated with the induced agent from Drosophila cells in an attempt to demonstrate infectivity. This could not be detected by the use of poly (rA) .oligo (dT) utilization as a measure of replication. These results are discussed and conclusions are drawn.