Andrews, Nancy C. and Levin, Daniel and Baltimore, David (1985) Poliovirus replicase stimulation by terminal uridylyl transferase. Journal of Biological Chemistry, 260 (12). pp. 7628-7635. ISSN 0021-9258 http://resolver.caltech.edu/CaltechAUTHORS:ANDjbc85
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In an in vitro poliovirus replication system, purified viral polymerase, plus sense virion RNA, and a host factor have been previously shown to be necessary for the transcription of minus strands. We have found that a partially purified eukaryotic initiation factor-2 (eIF-2) fraction from rabbit reticulocytes can replace HeLa host factor in the replicase reaction. This enzyme preparation contains eIF-2 and two other major proteins. In addition to eIF-2 activity, which does not appear to play a role in the replicase reaction, we find that the fraction contains terminal uridylyl transferase activity. The enzyme adds UMP moieties to the 3' end of primer RNA molecules. The number of UMP residues added depends on the primer. Although long tails of heterogeneous lengths (50 to 100 nucleotides) can be polymerized on the 3' end of oligo(U), a poly(A) primer accepts only four U's. The terminal uridylyl transferase activity requires only UTP, Mg2+, a sulfhydryl reagent, and an RNA primer for activity. It is partially associated with ribosomes. We provide preliminary evidence that it may be responsible for host factor-like activity. We present a model for minus strand synthesis by poliovirus replicase, based on the hypothesis that a terminal uridylyl transferase can participate in initiation.
|Additional Information:||Copyright © 1985 by the American Society for Biochemistry and Molecular Biology. (Received for publication, December 20, 1984) We would like to thank Nigel Crawford for his gift of partially purified viral polymerase used in the early stages of this work; Bernard Mathey-Prevot for many constructive suggestions; Magda Konarska for advice in the nearest neighbor analysis; Pim Zabel and Bert Flanegan for communication of results prior to publication; Karla Kirkegaard for critical reading of the manuscript; and Barbara Nelsen for helping to prepare virion RNA. This investigation was partially supported by United States Public Health Research Service Award 2T 32 GM07753-06 (to N.C.A., a Medical Scientist Training Program trainee) and Grant GM24825 (to D.L.) from the National Institute of General Medical Sciences, and by Grant A122346 from the National Institute for Allergy and Infectious Diseases (to D.B.). The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.|
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