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Hans Weber

Replication of RNA phages

Hans Weber
(Professor emeritus as of Jan. 2000)

Institut für Molekularbiologie
Universität Zürich
Winterthurerstrasse 190
8057 Zürich
phone: +41-1-63 53120
fax: +41-1-63 56864
 contact - research - publications


RNA-Protein Interactions in the Replication System of Bacteriophage Qbeta RNA

     Although the bacteriophage Qbeta RNA replication system has been known for over 30 years, it today still is of unique interest, because it represents a prototypical enzyme assembly that allows the specific and efficient amplification of an active viral RNA in vitro by soluble and defined molecular components.
     Intriguingly, only one of the subunits of the replicase enzyme is coded for by the viral genome. The other three are the host proteins EF-Tu and EF-Ts and the ribosomal protein S1. In order to use Qbeta RNA as a template, replicase requires an additional RNA binding protein, the Qbeta host factor, which was recently reported to have a role in the regulation of the stationary phase of the host cell.
     Our own recent results indicate that, paradoxically, it is two host proteins, namely the S1 protein and host factor, that are responsible for mediating the recognition of the phage RNA as a template, by binding the RNA at two internal sites and at the 3'-end. This conclusion comes mainly from studies in which we observed the effects of deletions in the RNA on template activity, as well as on the structure of the RNA-protein complexes as visualized by electron microscopy.
     In another approach, we have recently been able to adapt Qbeta to an E. coli strain devoid of host factor. All host factor-independent isolates were found to contain the same four point mutations, three of which affect the 3'-terminal secondary structure. Interestingly, a long-range base-pairing interaction involving the nucleotides at the very 3'-end is disrupted in the adapted mutants. This suggests that the host factor might function on wild type RNA as an "RNA chaperone", making the 3'-end available to replicase by melting out its 3'-terminal base-pairing interaction.
     Recognition of the Qbeta minus strand by replicase takes place by a completely different mechanism, for which neither S1 protein nor host factor are required. We found that on the RNA two specific secondary structure elements were important, one near the 5'-, the other near the 3'-end.
     RNA-protein interactions are of fundamental importance for the processes involved in genetic information transfer, but today they may well be the least understood of the different classes of macromolecular interactions. In view of the fact that only few RNA-protein complexes have been resolved by structural techniques, we attempt to describe the processes involved in phage RNA/replicase recognition at the highest possible resolution by molecular genetic/biochemical methods.


Recent publications

Schuppli, D., Miranda, G., Tsui H.-C. T., Winkler, M.E., Sogo, J.M. and Weber, H. (1997). Altered 3'-terminal RNA structure in phage Qbeta adapted to host factor-less Escherichia coli. Proc. Natl. Acad. Sci. USA 94, 10239-10242.

Miranda, G., Schuppli, D., Barrera, I., Hausherr, C., Sogo, J. M. and Weber, H. (1997). Recognition of bacteriophage Qbeta plus strand RNA as a template by Qbeta replicase: Role of RNA interactions mediated by ribosomal protein S1 and host factor. J. Mol. Biol. 267, 1089-1103.

Qiu, S., Schuppli, D., Tsui, H.-C. T., Winkler, M. E. and Weber, H. (1997). Strongly reduced phage Qbeta replication, but normal phage MS2 replication in an Escherichia coli K12 mutant with inactivated Qbeta host factor (hfq) gene. Virology 227, 211-214.

Schuppli, D., Barrera, I. and Weber, H. (1994). Identification of recognition elements on bacteriophage Qbeta minus strand RNA that are essential for template activity with Qbeta replicase. J. Mol. Biol. 243, 811-815.

Barrera, I., Schuppli, D., Sogo, J. M., and Weber, H. (1993). Different mechanisms of recognition of bacteriophage Qbeta plus and minus strand RNAs by Qbeta replicase. J. Mol. Biol. 232, 512-521.


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