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Nucleic Acids Research, 1993, Vol. 21, No. 15 3451-3457
© 1993

METHODS

Efficient expression of a protein coding gene under the control of an RNA polymerase I promoter

Theo D. Palmer, A.Dusty Miller, Ronald H. Reeder and Brian McStay*

Hutchinson Cancer Research Center 1124 Columbia Street, Seattle, WA 98104, USA

* To whom correspondence should be addressed at: Biomedical Research Centre, University of Dundee, Ninewells Hospital and Medical School, Dundee DD1 9SY, UK

Received April 22, 1993. Revised June 17, 1993. Accepted June 17, 1993.

In mammalian cells, RNA polymerase I transcripts are uncapped and retain a polyphosphate 5' terminus. It is probably for this reason that they are poorly translated as messenger RNA. We show in this report that insertion of an internal Ribosome Entry Site (IRES) into the 5' leader of an RNA polymerase I transcript overcomes the block to translation, presumably by substituting for the 5' trimethyl G cap. Addition of an SV40 polyA addition signal also enhances protein production from the RNA polymerase I transcript. RNA Polymerase I driven expression vectors containing both elements produce protein at levels comparable to that produced from RNA polymerase II driven expression vectors which utilize a retroviral LTR. RNA Polymerase I driven expression vectors may have a variety of uses both for basic research and for practical expression of recombinant proteins.


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