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Nucleic Acids Research, 2002, Vol. 30, No. 11 2290-2298
© 2002 Oxford University Press

RNA polymerase II complexes in the very early phase of transcription are not susceptible to TFIIS-induced exonucleolytic cleavage

Robert Sijbrandi, Ulrike Fiedler and H. Th. Marc Timmers*

Laboratory for Physiological Chemistry, UMCU, Stratenum, Universiteitsweg 100, 3584 CG Utrecht, The Netherlands

TFIIS is a transcription elongation factor for RNA polymerase II (pol II), which can suppress ribonucleotide misincorporation. We reconstituted transcription complexes in a highly purified pol II system on adenovirus Major-Late promoter constructs. We noted that these complexes have a high propensity for read-through upon GTP omission. Read-through occurred during the early stages at all registers analyzed. Addition of TFIIS reversed read-through of productive elongation complexes, which indicated that it was due to misincorporation. However, before register 13 transcription complexes were insensitive to TFIIS. These findings are discussed with respect to the structural models for pol II and we propose that TFIIS action is linked to the RNA:DNA hybrid.

* To whom correspondence should be addressed. Tel: +31 30 253 8981; Fax: +31 30 253 9035; Email: h.t.m.timmers{at}med.uu.nlPresent addresses:Robert Sijbrandi, Department of Molecular Microbiology, Vrije Universiteit, De Boelelaan 1085, 1081 HV Amsterdam, The NetherlandsUlrike Fiedler, Institute for Molecular Oncology, Department of Vascular Biology and Angiogenesis Research, Breisacher Strasse 117, D-79106 Freiburg, Germany


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