Abstract
Previous studies have shown that endogenous class III RNA polymerase(s) in nuclei from adenovirus 2-infected cells synthesize virus-coded RNA species which are approximately 200 (V200), 156 (V156), and 140 (V140) nucleotides in length (Weinmann, R., Brendler, T. G., Raskas, H.J., and Roeder, R. G. (1976) Cell 7, 557-566). The V156 nuclear RNA is identical in sequence to the major virus-associated RNA (VA RNA1 or 5.5 S RNA) synthesized in intact cells (Ohe, K., and Weissman, S. M. (1971) J. Biol. Chem. 246, 6991-7009). The V140 RNA contains several components, one of which appears identical to a minor virus-associated RNA (VA RNAII) which is synthesized in infected cells (Mathews), M. B. (1975) Cell 6, 223-229). Thus transcription of the VA RNAI and VA RNAII genes in vitro accurately reflects the in vivo transcription of these genes. The V200 RNA contains all the nucleotide sequences found in V156 RNA plus an additional 38 to 40 nucleotides on the 3' terminus. Transcription of the gene encoding this RNA species terminates within a stretch of 6 deoxythymidylic acid residues which are located 38 nucleotides beyond the predicted termination site for VA RNAI and which are preceded by a GC-rich sequence of nucleotides. These data suggest either that the V200 RNA is a precursor to the VA RNAI or that the RNA polymerase III occasionally reads-through the presumptive VA RNAI gene termination signal and stops at a potentially stronger downstream termination site.
Highlights
RNAII) which is synthesized in infected cells
In nuclei isolated from adenovirus 2-infected cells, RNA polymerase III synthesizes viral RNA species of approximately 200 (V,), 156 (V,; or 5.5 S RNA), and 140 (V,) nucleotides [3]
Subsequent to its isolation by hybridization, VA RNA11 was characterized by fingerprint analysis and shown to be distinct from VA RNA
Summary
In nuclei isolated from adenovirus 2-infected cells, RNA polymerase III synthesizes viral RNA species of approximately 200 (V,,,,), 156 (V,,,; or 5.5 S RNA), and 140. The VI56 RNA is the major transcript and appears similar [3, 4, 9, 10] to the major low molecular weight viral RNA (5.5 S RNA or VA RNA,) isolated from infected cells and sequenced by Ohe and Weissman [11]. The presence of other minor viral RNA species in intact adenovirus-infected cells has been reported. A major objective of the work reported here was to examine in greater detail the structural relationships of the various in vitro and in uivo adenovirus 2 transcripts These studies demonstrate that the in vitro VISfi and V141) transcripts are distinct and that they are indistinguishable, respectively, from the VA RNA, (5.5 S RNA) and VA RNA11 species observed in vivo. The cells were resuspended in 5 volumes of isotonic buffer (above) and disrupted
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